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    Comparison of forage yield, silage fermentative quality, anthocyanin stability, antioxidant activity, and in vitro rumen fermentation of anthocyanin-rich purple corn (Zea mays L.) stover and sticky corn stover
    TIAN Xing-zhou, Pramote Paengkoum, Siwaporn Paengkoum, Sorasak Thongpea, BAN Chao
    2018, 17 (09): 2082-2095.   DOI: 10.1016/S2095-3119(18)61970-7
    Abstract406)      PDF in ScienceDirect      
    The objective of this study was to observe the forage yield, silage fermentative quality, anthocyanin stability, and antioxidant activity during the storage period and in vitro rumen fermentation of anthocyanin-rich purple corn (Zea mays L.) stover (PS) and sticky corn stover (SS).  Forage yield of corn stover was weighed and ensiled with two treatments: (1) hybrid sticky waxy corn stover (control), and (2) hybrid purple waxy corn stover (treatment).  Samples were stored in mini-silos for periods of 0, 7, 14, 21, 42, 63, 84, and 105 d.  The results showed that PS had significantly higher (P<0.05) yields of dry matter (DM), organic matter (OM), gross energy (GE), crude protein (CP), neutral detergent fiber (NDF), acid detergent fiber (ADF), and total anthocyanins than that of the SS.  Anthocyanin-rich purple corn stover silage (PSS) showed higher (P<0.05) levels of DM and CP relative to the sticky corn stover silage (SSS).  Although anthocyanin-rich PSS displayed a lower (P<0.05) level of pelargonidin-3-glucoside (P3G), it had higher (P<0.05) levels of peonidin (Peo) and pelargonidin (Pel) compared to the control.  Delphinidin (Del) and malvidin (Mal) were not detected in SSS during the ensilage period; in PSS, Del was no longer detected after 7 d of ensilage.  Specifically, total anthocyanins in anthocyanin-rich PSS decreased rapidly (P<0.05) prior to 7 d of ensilage, and then remained at relatively stable (P>0.05) constants.  Compared to the anthocyanin-rich PSS, SSS displayed significantly higher P<0.05) pH value and ammonia nitrogen (NH3-N) content.  Propionic acid (PA) at 0 d and butyric acid (BA) during the entire study period were not detected, whereas anthocyanin-rich PSS showed a higher (P<0.05) level of lactic acid (LA) than that of the SSS.  Compared with the SSS extract, anthocyanin-rich PSS extract showed a higher (P<0.05) level of 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging activity and displayed a lower (P<0.05) half maximal inhibitory concentration (IC50) value.  Moreover, anthocyanin-rich PSS reduced (P<0.05) gas production (GP), and displayed lower levels of immediately soluble fraction and ratio of acetic acid (AA) to PA at 12 h, but the other parameters were unaffected (P>0.05) relative to the control.  Taken together, the results indicated that: (1) anthocyanins could be stable in silage; (2) anthocyanin-rich PSS showed better silage fermentative quality and stronger antioxidant activity; and (3) anthocyanin-rich PSS had no negative effect on rumen fermentation parameters.
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    MicroRNA-22 inhibits proliferation and promotes differentiation of satellite cells in porcine skeletal muscle
    Hong Quyen Dang, XU Gu-li, HOU Lian-jie, XU Jian, HONG Guang-liang, Chingyuan Hu, WANG Chong
    2020, 19 (1): 225-233.   DOI: 10.1016/S2095-3119(19)62701-2
    Abstract84)      PDF in ScienceDirect      
    Pig is an important economic animal in China.  Improving meat quality and meat productivity is a long time issue in animal genetic breeding.  MicroRNAs (miRNAs) are short non-coding RNAs that participate in various biological processes, such as muscle development and embryogenesis.  miR-22 differentially expresses in embryonic and adult skeletal muscle.  However, the underlying mechanism is unclear.  In this study, we investigated miR-22 function in proliferation and differentiation of porcine satellite cells (PSCs) in skeletal muscle.  Our data show that miR-22 expressed in both proliferation and differentiated PSCs and is significantly upregulated (P<0.05) during differentiation.  After treated with the miR-22 inhibitor, PSCs proliferation was significantly increased (P<0.05), as indicated by the up-regulation (P<0.01) of cyclin D1 (CCND1), cyclin B1 (CCNB1) and down-regulation (P<0.05) of P21.  Conversely, over-expression of miR-22 resulted in opposite results.  Differentiation of PSCs was significantly suppressed (P<0.05), evidenced by two major myogenic markers: myogenin (MyoG) and myosin heavy chain (MyHC), after transfecting the PSCs with miR-22 inhibitor.  Opposite results were demonstrated in the other way around by transfection with miR-22 mimics.  In conclusion, the data from this study indicated that miR-22 inhibited the PSCs proliferation but promoted their differentiation. 
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    Genetic parameter estimation and genome-wide association study (GWAS) of red blood cell count at three stages in a Duroc×Erhualian pig population
    NAN Jiu-hong, YIN Li-lin, TANG Zhen-shuang, CHEN Jian-hai, ZHANG Jie, WANG Hai-yan, DU Xiao-yong, LIU Xiang-dong
    2020, 19 (3): 793-799.   DOI: 10.1016/S2095-3119(19)62773-5
    Abstract81)      PDF in ScienceDirect      
    Red blood cells play an essential role in the immune system.  Moreover, red blood cell count (RBC) is an important clinical indicator of various diseases, including anemia, type 2 diabetes and the metabolic syndrome.  Thus, it is necessary to reveal the genetic mechanism of RBC for animal disease resistance breeding.  However, quite a few studies had focused on porcine RBC, especially at different stages.  Thus, studies on porcine RBC at different stages are needed for disease resistant breeding.  In this study, the porcine RBC of 20-, 33-, and 80-day old were measured, and genetic parameter estimation and genome-wide association study (GWAS) were both performed.  As a result, the heritability was about 0.6 at the early stages, much higher than that at 80 days.  Nine novel genome wide significant single nucleotide polymorphisms (SNPs), located at Sus scrofa chromosome (SSC)3, 4, 8, 9, 10 and 15, respectively, were identified.  Further, TGFβ2, TMCC2 and PPP1R15B genes were identified as important candidate genes of porcine red blood cell count.  So different SNPs and candidate genes were found significantly associated with porcine RBC at different stages, suggesting that different genes might play key roles on porcine RBC at different stages.  Overall, new evidences were offered in this study for the genetic bases of animal RBC, and that the SNPs and candidate genes would be useful for disease resistant breeding of pig.
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    Expression and contribution of microphthalmia-associated transcription factor to the melanin deposition in Liancheng white ducks
    XIN Qing-wu, MIAO Zhong-wei, LIU Zhao-yuan, LI Li, ZHANG Lin-li, ZHU Zhi-ming, ZHANG Zheng-hong, ZHENG Nen-zhu, WANG Zheng-chao
    2020, 19 (3): 800-809.   DOI: 10.1016/S2095-3119(19)62736-X
    Abstract66)      PDF in ScienceDirect      
    The present study investigates the expression of microphthalmia-associated transcription factor (MITF) and its contribution to the melanin deposition in Liancheng white ducks.  Nested PCR was used to clone the MITF gene sequence from the skin tissue of female Liancheng white ducks.  Ultraviolet spectrophotometry was used to detect the melanin deposition.  MITF mRNA expression and melanin deposition in different tissues and organs were detected and their correlation was analyzed.  The MITF gene (GenBank number: MG516570) was 1 323 bp in length, contains a complete CDS region (34–1 323 bp) and codes 429 amino acids with 100% homology to the MITF of Anas platyrhynchos and over 95% homology to those of Gallus gallus and Coturnix japonica.  Genetic evolution analysis reveals a close relationship of Liancheng white ducks with A. platyrhynchos, and also to lesser extents with Anser cygnoides, silky fowl and G. gallus, as well as Sus scrofa, Ovis aries and other mammals.  Real-time quantitative PCR (qPCR) analysis demonstrated that MITF was expressed in skin, gizzard, liver, kidney and muscle, and of these tissues, its expression was the highest in the skin tissue (skin>gizzard>liver>kidney>muscle).  Ultraviolet spectrophotometry showed that melanin deposition was positively correlated with the MITF expression level in these five tissues and organs (P<0.05).  Together, these results demonstrated a tissue-specific pattern of MITF expression and a positive correlation between MITF expression and melanin deposition, indicating that MITF expression may contribute to the melanin deposition in Liancheng white ducks.
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    Tissue-specific expression and correlation with promoter DNA methylation of the LBP gene in pigs
    CAO Yue, GAO Zhong-cheng, WU Zheng-chang, WANG Hai-fei, BAO Wen-bin
    2020, 19 (4): 1055-1064.   DOI: 10.1016/S2095-3119(19)62749-8
    Abstract56)      PDF in ScienceDirect      
    Lipopolysaccharide binding protein (LBP) is a key factor in the recognition of lipopolysaccharide (LPS) and the initiation of immune response, thus regulating the body’s resistance to pathogenic infection.  To investigate the tissue-specific expression characteristics of the LBP gene and its transcriptional regulation in pigs, we detected LBP expression in different tissues of 35-day-old Meishan weaned piglets, determined LBP core promoter region using bioinformatics prediction combined with dual luciferase activity assay, and finally detected methylation levels by pyrosequencing.  The results showed that LBP expression in the liver tissue was significantly higher (P<0.01) than that in other tissues, followed by the intestinal tissues.  The core promoter region of LBP was located at –500–(–206) bp (chr.17: g.46837534–g.46837828), containing three CpG sites (CpG1, CpG2 and CpG3).  Of the three CpG sites, CpG2 and CpG3 were variously methylated (P<0.01) in different tissues.  Moreover, LBP mRNA levels were negatively correlated (P<0.01) with methylation levels of the CpG2 and CpG3 sites in the YY1 transcription factor binding sequence.  It is speculated that the methylation of CpG2 and CpG3 sites might inhibit YY1 binding to the promoter sequences, thereby regulating the tissue-specific expression of LBP.  This study demonstrated the distinct patterns of LBP expression and promoter methylation in the tissues of Meishan pigs and indicated the potential roles of DNA methylation in regulating LBP expression, which may contribute to further investigations on pig LBP gene expression and function. 
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    The CRISPR/Cas9 induces large genomic fragment deletions of MSTN and phenotypic changes in sheep
    DING Yi, ZHOU Shi-wei, DING Qiang, CAI Bei, ZHAO Xiao-e, ZHONG Shu, JIN Miao-han, WANG Xiao-long, MA Bao-hua, CHEN Yu-lin
    2020, 19 (4): 1065-1073.   DOI: 10.1016/S2095-3119(19)62853-4
    Abstract140)      PDF in ScienceDirect      
    The CRISPR/Cas9 system has been extensively used to engineer genetic loci for the generation of knockouts, insertions, and point mutations in animal models.  However, many mutations that have been reported in animals are small insertions or deletions.  This study used the CRISPR/Cas9 system to induce large DNA fragment deletions in MSTN via three guide RNAs in sheep.  This successfully achieved the precise gene editing of the ovine MSTN gene by injecting both Cas9 mRNA and sgRNAs into embryos at the one-cell stage.  Of 10 edited animals, 3 animals (30%) exhibited large genomic fragment deletions (~5 kb).  Furthermore, the body weights of these 3 animals were significantly different (P0<0.0001, P15=0.001, P30=0.005, P60=0.027) between lambs with large deletions and wildtype lambs.  In addition, the edited lambs were also significantly different (P0<0.0001, P15<0.0001, P30=0.002, P60=0.011) compared with wildtype.  These results suggest that the generated MSTN knockout sheep is a reliable and effective animal model for further study.  Furthermore, this method is time- and labor-saving, and efficient for the creation of animal models for agriculture, biology, and medicine.
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    Genome-wide detection of selective signatures in a Jinhua pig population
    XU Zhong, SUN Hao, ZHANG Zhe, Zhao Qing-bo, Babatunde Shittu Olasege, Li Qiu-meng, Yue Yang, Ma Pei-pei, Zhang Xiang-zhe, Wang Qi-shan, Pan Yu-chun
    2020, 19 (5): 1314-1322.   DOI: 10.1016/S2095-3119(19)62833-9
    Abstract70)      PDF in ScienceDirect      
    The aim of this study was to detect evidence for signatures of recent selection in the Jinhua pig genome.  These results can be useful to better understand the regions under selection in Jinhua pigs and might shed some lights on groups of genes that control production traits.  In the present study, we performed extended haplotype homozygosity (EHH) tests to identify significant core regions in 202 Jinhua pigs.  A total of 26 161 core regions spanning 636.42 Mb were identified, which occupied approximately 28% of the genome across all autosomes, and 1 158 significant (P<0.01) core haplotypes were selected.  Genes in these regions were related to several economically important traits, including meat quality, reproduction, immune responses and exterior traits.  A panel of genes including ssc-mir-365-2, KDM8, RABEP2, GSG1L, RHEB, RPH3AL and a signal pathway of PI3K-Akt were detected with the most extreme P-values.  The findings in our study could draw a comparatively genome-wide map of selection signature in the pig genome, and also help to detect functional candidate genes under positive selection for further genetic and breeding research in Jinhua and other pigs.
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    Investigating structural impact of a valine to isoleucine substitution on anti-Müllerian hormone in silico and genetic association of the variant and AMH expression with egg production in chickens
    DANG Li-ping, LIU Rui-fang, ZHAO Wen-yan, ZHOU Wen-xin, MIN Yu-na, WANG Zhe-peng
    2020, 19 (6): 1635-1643.   DOI: 10.1016/S2095-3119(20)63176-8
    Abstract95)      PDF in ScienceDirect      
    Anti-Müllerian hormone (AMH) acts in maintaining orderly cyclic recruitment of early follicles, suggesting that it is a promising candidate for influencing animal reproductive efficiency.  This study aimed to elucidate the effect of a missense mutation of Val566Ile on the structure of AMH protein and the genetic association of Val566Ile and AMH expression with egg production in chickens.  Structural perturbations of Val566Ile were predicted by homology modeling.  The association of the variant with the number of eggs was tested using a quantitative trait transmission disequilibrium test model. AMH expression in granulosa cells in Lueyang black-boned chickens was compared with that in Nick chickens.  The Val566 of AMH is a non-conservative amino acid among mammals and birds, but its hydrophobicity is completely conservative.  The substitution of Val566 for Ile566 potentially disrupted hydrogen bonds and solvent accessibility of 22 residues and created a short α-helix in the C terminus of AMH.  Despite having striking structure-disrupting potential, the variant was not statistically associated with the number of eggs (P>0.05) in the Lueyang black-boned chickens.  We did not detect differential expression of AMH between Lueyang black-boned chickens and Nick chickens (P>0.05).  These results confirmed the structural impact of Val566Ile, but suggested that Val566Ile and AMH expression might not be the major genetic determinants for egg production in Lueyang black-boned chickens.
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    The untold story between enhancers and skeletal muscle development
    ZHANG Yong-sheng, LU Dan, LIU Yu-wen, YI Guo-qiang, TANG Zhong-lin
    2020, 19 (9): 2137-2149.   DOI: 10.1016/S2095-3119(20)63235-X
    Abstract97)      PDF in ScienceDirect      
    Currently, enhancers have key transcriptional regulatory roles in muscle development.  Skeletal muscle formation involves various molecules, and in animals, enhancers are one of the main types of transcriptional regulatory regions that are of great importance to regulate myogenic gene expression.  In muscle development, enhancers can generate enhancer RNAs (eRNAs) that are involved in the regulation of gene transcription.  The regulation of gene expression by eRNAs offers great potential in improving animal production traits.  Herein we comprehensively review the roles of enhancers in muscle formation and its potential function in skeletal muscle development.  This review will describe the future application of enhancers in skeletal muscle development and discuss the prospects that enhancer studies offer for agriculture, biotechnology, and animal breeding.
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    Two new SINE insertion polymorphisms in pig Vertnin (VRTN) gene revealed by comparative genomic alignment
    ZHENG Yao, CHEN Cai, CHEN Wei, WANG Xiao-yan, WANG Wei, GAO Bo, Klaus WIMMERS, MAO Jiu-de, SONG Cheng-yi
    2020, 19 (10): 2514-2522.   DOI: 10.1016/S2095-3119(20)63255-5
    Abstract72)      PDF in ScienceDirect      
    Despite one SINE retrotransposon insertion polymorphism (sRTIP) in the vertebrae development-associated (VRTN) gene was identified in pigs, the structural variations (SVs) in VRTN gene and its proximal flank regions were largely unknown.  VRTN genic and flanking sequences from 14 breeds were assembled or downloaded from whole-genome shotgun contings (WGS) database, and aligned to identify the SVs with Clustalx, and retrotransposons in VRTN gene were annotated by RepeatMasker, the splicing patterns of VRTN gene were predicted by Genescan, and large SVs were evaluated by PCR.  A total of 12 small SVs and three large SVs in intron of VRTN, derived from SINE insertion polymorphisms, were identified, and two of them (VRTN-sRTIP2 and VRTN-sRTIP3) were not reported before.  These VRTN-sRTIPs may affect the splicing patterns of VRTN.  They displayed polymorphisms in most detected eight breeds.  VRTN-sRTIP2 and VRTN-sRTIP3 showed Hardy-Weinberg equilibrium distributions in most populations except the Chinese local Erhualian pigs, while VRTN-sRTIP1 showed genetic equilibrium in Erhualian pigs.  Three VRTN-sRTIPs were identified, and displayed polymorphisms in pigs, and two of them were not reported before.  These SVs provide a useful molecular markers for genetic analysis in pigs, and offer new information to facilitate the understanding the SVs of VRTN gene and their putative roles in the variation of vertebral number.
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    The expression of Lin28B was co-regulated by H3K4me2 and Wnt5a/β-catenin/TCF7L2
    ZHANG Ya-ni, HU Cai, WANG Ying-jie, ZUO Qi-sheng, LI Bi-chun
    2020, 19 (12): 3054-3064.   DOI: 10.1016/S2095-3119(20)63441-4
    Abstract78)      PDF in ScienceDirect      
    Lin28A and Lin28B are homologous RNA-binding proteins that participate in the development of primordial germ cells.  The mechanisms underlying expression and regulation of Lin28A have been well documented, but such information for Lin28B is limited.  In this study, a fragment of the Lin28B promoter was cloned, the pEGFP-pLin28B vector was constructed.  DF-1 chicken fibroblasts were transfected and the expression of green fluorescent protein (GFP) was measured.  Furtherly, Lin28B promoter of different lengths fragments was cloned using the chromosome-walking method and the fragments were ligated into the PGL3-Basic vector, and transfected into DF-1 cells.  Results of dual-luciferase reporter assay showed that the core of the Lin28B promoter was included in the sequence from –1 431 to –1 034 bp.  The binding sites of the transcription factor TCF7L2 was showed within this sequence by bioinformatics analysis.  The promoter activity of Lin28B was downregulated (P<0.05) when the TCF7L2 binding site was mutated.  Further experiments suggested that Lin28B promoter activity responded to the activation or inhibition of Wnt signaling.  Results of chromatin immunoprecipitation and quantitative PCR showed that β-catenin-TCF7L2 may be enriched in the Lin28B promoter core area.  In vivo and in vitro activation or inhibition of Wnt signaling significantly up- or down-regulated (P<0.05) Lin28B expression.  H3K4me2 enriched in the promoter of Lin28B, which affected the regulation of Wnt signaling to Lin28B.  In conclusion, our results showed that H3K4me2 and Wnt5a/β-catenin/TCF7L2 were the positive regulators of Lin28B expression.  Findings of this study may lay a theoretical foundation for illuminating the mechanism underlying Lin28B expression.
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    The tissue expression level of BPI gene in piglets from newborn to weaning and its relationship with Gram-negative bacterial infection
    DAI Chao-hui, CAO Yue, GAO Zhong-cheng, ZHU Guo-qiang, WU Sheng-long, BAO Wen-bin
    2020, 19 (12): 3065-3073.   DOI: 10.1016/S2095-3119(20)63369-X
    Abstract50)      PDF in ScienceDirect      
    The bactericidal/permeability increasing protein (BPI) has an important function of nonspecific killing of Gram-negative bacteria.  In this study, qPCR was used to detect the expression of the BPI gene in twelve tissues of Meishan piglets from birth to weaning.  BPI gene overexpression, bacterial adhesions count and indirect immunofluorescence were applied to analyze the relationship between BPI gene expression and the infectivity of Escherichia coli and Salmonella.  The results showed that the BPI gene was expressed highly in duodenum, jejunum and ileum (fold changes of relative expression levels were more than 10 000, 500 and 200, respectively).  The expression of the BPI gene at 35 days of age was significantly higher (P<0.01) than that at all other days.  Transcription of the BPI gene was up-regulated 2 401-fold in porcine intestinal epithelial (IPEC-J2) cells that were transfected with the BPI gene overexpression lentivirus (IPEC-J2-BPI), and significantly higher (P<0.01) than that in negative control cells (IPEC-J2-NC).  Protein expression levels in IPEC-J2-BPI cells were also increased.  When IPEC-J2 cells were incubated with E. coli and Salmonella, respectively, for 2, 4, 6, 8, 10 and 12 h, the number of bacterial adhesions in IPEC-J2-BPI cells was significantly less (P<0.05) than that in IPEC-J2-NC cells.  The results of indirect immunofluorescence analysis showed that the number of bacterial adhesions in IPEC-J2-BPI cells was significantly less (P<0.01) than that in IPEC-J2-NC cells.  These results demonstrated that the BPI gene might play an important role in regulating weaning stress especially intestinal-mediated immune response.  Overexpression of the BPI gene at the cellular level could significantly enhance the anti-bactericidal ability against Gram-negative bacteria such as E. coli and Salmonella.  This has important biological significance in piglet resistance to bacterial diarrhea. 
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    Infectious disease-resistant pigs: Will they fly?
    2021, 20 (1): 1-3.   DOI: 10.1016/S2095-3119(20)63468-2
    Abstract41)      PDF in ScienceDirect      
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    Identification and functional prediction of long intergenic noncoding RNAs in fetal porcine longissimus dorsi muscle
    LI Cen-cen, YU Shu-long, REN Hai-feng, WU Wei, WANG Ya-ling, HAN Qiu, XU Hai-xia, XU Yong-jie, ZHANG Peng-peng
    2021, 20 (1): 201-211.   DOI: 10.1016/S2095-3119(20)63261-0
    Abstract75)      PDF in ScienceDirect      
    Pigs are globally farmed animals which provide protein for human consumption in the form of skeletal muscle.  To better understand the function of long intergenic noncoding RNAs (lincRNAs) in porcine skeletal muscle growth and development, we collected RNA-seq data from porcine longissimus dorsi muscle (LDM) during embryonic development.  We identified a total of 739 lincRNA transcripts, which were distributed on all chromosomes except the chromosome Y, and analyzed their molecular characteristics.  Compared to protein-coding genes, lincRNAs showed shorter transcripts, longer exons, fewer exons and higher tissue specificity.  In addition, the abundance of lincRNAs in five embryonic development stages were analyzed and 45 differentially expressed lincRNAs were screened, three of which were highly expressed in LDM during porcine embryonic development.  Finally, we predicted the potential target genes and functions of the lincRNAs, and identified 1 537 cis-target genes and 8 571 trans-target genes.  Furthermore, we identified two key candidate lincRNAs involved in muscle development, XLOC_024652 and XLOC_001832, for post-trial validation.  Our results provide a genome-wide resource of lincRNAs which are potentially involved in porcine embryonic skeletal muscle development and lay a foundation for the further study of their functions.
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    Switches in transcriptome functions during seven skeletal muscle development stages from fetus to kid in Capra hircus
    LING Ying-hui, ZHENG Qi, JING Jing, SUI Meng-hua, ZHU Lu, LI Yun-sheng, ZHANG Yun-hai, LIU Ya, FANG Fu-gui, ZHANG Xiao-rong
    2021, 20 (1): 212-226.   DOI: 10.1016/S2095-3119(20)63268-3
    Abstract111)      PDF in ScienceDirect      
    Skeletal muscle accounts for about 40% of mammalian body weight, the development of which is a dynamic, complex and precisely regulated process that is critical for meat production. We here described the transcriptome expression profile in 21 goat samples collected at 7 growth stages from fetus to kid, including fetal 45 (F45), 65 (F65), 90 (F90), 120 (F120), and 135 (F135) days, and birth 1 (B1) day and 90 (B90) days kids.  Paraffin sections combined with RNA-seq data of the 7 stages divided the transcriptomic functions of skeletal muscle into 4 states: before F90, F120, F135 and B1, and B90.  And the dynamic expression of all 4 793 differentially expressed genes (DEGs) was identified.  Furthermore, DEGs were clustered by weighted gene correlation network analysis into 4 modules (turquoise, grey, blue and brown) that corresponded to these 4 states.  Functional and pathway analysis indicated that the active genes in the stages before F90 (turquoise) were closely related to skeletal muscle proliferation.  The DEGs in the F120-related module (grey) were found to participate in the regulation of skeletal muscle structure and skeletal muscle development by regulating tRNA.  The brown module (F135 and B1) regulated fatty acid biological processes to maintain the normal development of muscle cells.  The DEGs of B90 high correlation module (blue) were involved the strengthening and power of skeletal muscle through the regulation of actin filaments and tropomyosin.  Our current data thus revealed the internal functional conversion of the goat skeletal muscle in the growth from fetus to kid.  The results provided a theoretical basis for analyzing the involvement of mRNA in skeletal muscle development.
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    Jasmonic acid and ethylene signaling pathways participate in the defense response of Chinese cabbage to Pectobacterium carotovorum infection
    CHEN Chang-long, YUAN Fang, LI Xiao-ying, MA Rong-cai, XIE Hua
    2021, 20 (5): 1314-1326.   DOI: 10.1016/S2095-3119(20)63267-1
    Abstract49)      PDF in ScienceDirect      
    Chinese cabbage (Brassica rapa subsp. pekinensis) suffers from soft rot disease caused by Pectobacterium carotovorum (Pc).  To uncover the mechanisms underlying the defense response of Chinese cabbage to Pc, we constructed a suppression subtractive hybridization (SSH) library from Pc-infected cabbage and obtained 1 919 non-redundant expressed sequence tags (ESTs), which were used for cDNA microarray.  We detected 800 differentially expressed genes (DEGs) in cabbage at different time points post-Pc inoculation, which were further confirmed by quantitative real-time PCR.  One quarter of these DEGs were involved in the biotic stress pathways visualized by MapMan.  Among them, 8, 8, 1, 3, and 2 DEGs were related to jasmonic acid (JA), ethylene (ET), JA+ET, auxin, and abscisic acid (ABA) signaling pathways, respectively, while no DEG was detected for salicylic acid (SA) signaling.  Assessment of phytohormone production in the Pc-infected leaves showed that JA and ET production was increased, while SA production was decreased.  Treatment with JA, methyl jasmonate (MeJA), the ET precursor 1-aminocyclopropane-1-carboxylate (ACC), or combinations thereof, reduced the disease severity, and the JA and JA+ACC treatments were superior and performed equally well.  Our findings suggest that JA and ET may act synergistically against Pc infection in Chinese cabbage, and JA-mediated signaling might be the most significant. 
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    circRNA landscape of non-pregnant endometrium during the estrus cycle in dairy goats
    LIU Xiao-rui, ZHANG Lei, CUI Jiu-zeng, YANG Li-chun, HAN Jin-cheng, CHE Si-cheng, CAO Bin-yun, LI Guang, SONG Yu-xuan
    2021, 20 (5): 1346-1358.   DOI: 10.1016/S2095-3119(20)63464-5
    Abstract88)      PDF in ScienceDirect      
    Endometrial development is a complicated process involving numerous regulatory factors.  Circular RNAs (circRNAs) have been known as a member of the naturally occurring non-coding RNA family, and are reportedly crucial for a variety of physiological processes.  This study investigated the circRNA landscape of non-pregnant endometrium of dairy goats during estrus.  Non-pregnant endometrial samples of goats at estrus day 5 (Ed5) and estrus day 15 (Ed15) were used to methodically analyze the circRNA landscape using strand-specific Ribo-Zero RNA-Seq.  A total of 2 331 differentially expressed (P<0.05) circRNAs (DEciRs) between Ed5 and Ed15 were discovered in the goat endometrium.  It was found that Nipped-B-like (NIPBL) and calcium responsive transcription factor (CARF) may participate in the development of the endometrium by decreasing (P<0.05) the levels of their circRNA-transcript forms.  Furthermore, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses of DEciR host genes (hgDEciRs) revealed that tight junctions and GTPases may be involved in endometrial development during the estrus cycle.  A total of 2 331 DEciRs were discovered in the endometrium at Ed5 and Ed15.  Based on GO and KEGG enrichment analyses, it could be inferred that tight junctions and GTPases are likely to play an important role in the development of goat endometrium during the estrus cycle.  This circRNA study greatly enhances our knowledge of global trends in the development of non-pregnant endometrium during the estrus cycle in goats; these results help us to better understand the molecular regulation of endometrial development in dairy goats.
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    Exploring the genetic features and signatures of selection in South China indigenous pigs
    DIAO Shu-qi, XU Zhi-ting, YE Shao-pan, HUANG Shu-wen, TENG Jin-yan, YUAN Xiao-long, CHEN Zan-mou, ZHANG Hao, LI Jia-qi, ZHANG Zhe
    2021, 20 (5): 1359-1371.   DOI: 10.1016/S2095-3119(20)63260-9
    Abstract81)      PDF in ScienceDirect      
    To explore the genetic features and signatures of selection in indigenous pigs from South China and Duroc pigs, 259 pigs from six populations were genotyped using single-nucleotide polymorphism (SNP) BeadChips.  Principal component analysis (PCA), effective population size (Ne), linkage disequilibrium (LD), and signatures of selection were explored and investigated among the six pig populations.  The results showed the Ne of five South China indigenous pig populations has been decreasing rapidly since 100 generations ago.  The LD between pairwise SNP distance at 100 kb ranged from 0.16 to 0.20 for the five indigenous pig populations, while it was 0.32 for the Duroc population.  However, the LD of all six pig populations showed the opposite order at long distances (>5 Mb).  Furthermore, 15 potential signatures of selection associated with meat quality and age at puberty were exclusively detected in South China indigenous pigs, while eight potential signatures of selection associated with growth traits were detected in Duroc pigs.  Our work provides valuable insights for the utilization and conservation of South China indigenous pigs.
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    Identifying the complex genetic architecture of growth and fatness traits in a Duroc pig population
    ZHANG Zhe, CHEN Zi-tao, DIAO Shu-qi, YE Shao-pan, WANG Jia-ying, GAO Ning, YUAN Xiao-long, CHEN Zan-mou, ZHANG Hao, LI Jia-qi
    2021, 20 (6): 1607-1614.   DOI: 10.1016/S2095-3119(20)63264-6
    Abstract88)      PDF in ScienceDirect      
    In modern pig breeding programs, growth and fatness are vital economic traits that significantly influence porcine production.  To identify underlying variants and candidate genes associated with growth and fatness traits, a total of 1 067 genotyped Duroc pigs with de-regressed estimated breeding values (DEBV) records were analyzed in a genome wide association study (GWAS) by using a single marker regression model.  In total, 28 potential single nucleotide polymorphisms (SNPs) were associated with these traits of interest.  Moreover, VPS4B, PHLPP1, and some other genes were highlighted as functionally plausible candidate genes that compose the underlying genetic architecture of porcine growth and fatness traits.  Our findings contribute to a better understanding of the genetic architectures underlying swine growth and fatness traits that can be potentially used in pig breeding programs. 
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    Genome-wide scan for selection signatures based on whole-genome re-sequencing in Landrace and Yorkshire pigs
    WANG Kai, WU Ping-xian, CHEN De-juan, ZHOU Jie, YANG Xi-di, JIANG An-an, MA Ji-deng, TANG Qian-zi, XIAO Wei-hang, JIANG Yan-zhi, ZHU Li, QIU Xiao-tian, LI Ming-zhou, LI Xue-wei, TANG Guo-qing
    2021, 20 (7): 1898-1906.   DOI: 10.1016/S2095-3119(20)63488-8
    Abstract93)      PDF in ScienceDirect      
    We performed a genome-wide scan to detect selection signatures that showed evidence of positive selection in the domestication process by re-sequencing the whole genomes of Landrace and Yorkshire pigs.  Fifteen annotated elements with 13 associated genes were identified using the Z-transformed FST (Z(FST)) method, and 208 annotated elements with 140 associated genes were identified using the Z-transformed heterozygosity (ZHp) method.  The functional analysis and the results of previous studies showed that most of the candidate genes were associated with basic metabolism, disease resistance, cellular processes, and biochemical signals, and several were related to body morphology and organs.  They included PPP3CA, which plays an essential role in the transduction of intracellular Ca2+-mediated signals, and WWTR1, which plays a pivotal role in organ size control and tumor suppression.  These results suggest that genes associated with body morphology were subject to selection pressure during domestication, whereas genes involved in basic metabolism and disease resistance were subject to selection during artificial breeding.  Our findings provide new insights into the potential genetic variation of phenotypic diversity in different pig breeds and will help to better understand the selection effects of modern breeding in Landrace and Yorkshire pigs.
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    Bioinformatic analysis and functional characterization of CFEM proteins in Setosphaeria turcica
    WANG Jian-xia, LONG Feng, ZHU Hang, ZHANG Yan, WU Jian-ying, SHEN Shen, DONG Jin-gao, HAO Zhi-min
    2021, 20 (9): 2438-2449.   DOI: 10.1016/S2095-3119(20)63342-1
    Abstract139)      PDF in ScienceDirect      
    Common in Fungal Extracellular Membrane (CFEM) domains are uniquely found in fungal extracellular membrane proteins which are important for pathogens.  This study identified 13 StCFEM proteins in the genome of Setosphaeria turcica, the hemibiotrophic fungus that causes northern corn leaf blight.  Sequence alignment and WebLogo analysis of their CFEM domains indicated that the amino acids were highly conserved and that, with the exception of StCFEM1, 2, 3, and 6, they contained eight cysteines.  Phylogenic analysis suggested that these 13 proteins (StCFEM1–13) could be divided into 2 clades based on the presence of the trans-membrane domain.  Six StCFEM proteins with a signal peptide and without a trans-membrane domain were considered as candidate effector proteins.  The CFEM domain in the candidate effector proteins could form a helical-basket structure homologous to Csa2 in Candida albicans.  Transcriptome analysis suggested that the 13 genes were expressed during fungal infection and a yeast secretion assay revealed that these candidate effectors were secreted proteins.  It was also found that StCFEM3, 4, and 5 couldn’t affect BAX/INF1-induced programmed cell death (PCD) in Nicotiana benthamiana and while StCFEM12 could suppress INF1-induced PCD, it showed no effect on BAX-induced PCD.  This study found that there were 13 members of the S. turcica CFEM protein family and that StCFEM12 was a candidate effector.  This study laid the foundation for illustrating the roles of CFEM proteins during the pathogenic processes of phytopathogens.
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    Identifying SNPs associated with birth weight and days to 100 kg traits in Yorkshire pigs based on genotyping-by-sequencing
    WU Ping-xian, ZHOU Jie, WANG Kai, CHEN De-juan, YANG Xi-di, LIU Yi-hui, JIANG An-an, SHEN Lin-yuan, JIN Long, XIAO Wei-hang, JIANG Yan-zhi, LI Ming-zhou, ZHU Li, ZENG Yang-shuang, XU Xu, QIU Xiao-tian, LI Xue-wei, TANG Guo-qing
    2021, 20 (9): 2483-2490.   DOI: 10.1016/S2095-3119(20)63474-8
    Abstract76)      PDF in ScienceDirect      
    Birth weight (BW) and days to 100 kg (D100) are important economic traits that are both affected by polygenes.  However, the genetic architecture of these quantitative traits is still elusive.  Genotyping-by-sequencing (GBS) data containing a large number of single nucleotide polymorphisms (SNPs) have become a powerful tool in genomic analysis.  To better understand their complex genetic structure, a total of 600 Yorkshire pigs were sequenced using GBS technology.  After quality control, 279 787 SNPs were generated for subsequent genome-wide association study (GWAS).  A total of 30 genome-wide SNPs (P<1.79E–07) were identified for D100.  Furthermore, a total of 22 and 2 suggestive SNPs (P<3.57E–06) were detected for D100 and BW, respectively.  Of these, one locus located on SSC12 (position: 46 226 512 bp) were evaluated to affect both BW and D100 in Yorkshire pigs, indicating the pleiotropism in different traits.  Considering the function of candidate genes, two genes, NSRP1 and DOCK7, were suggested as the most promising candidate genes involved in growth traits.  Thus, use of GBS is able to identify novel variants and potential candidate genes for BW and D100, and provide an opportunity for improving pig growth traits using genomic selection in pigs.
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    Integration of association and computational methods reveals functional variants of LEPR gene for abdominal fat content in chickens
    LI Yu-dong, WANG Wei-jia, LI Zi-wei, WANG Ning, XIAO Fan, GAO Hai-he, GUO Huai-shun, LI Hui, WANG Shou-zhi
    2021, 20 (10): 2734-2748.   DOI: 10.1016/S2095-3119(20)63575-4
    Abstract73)      PDF in ScienceDirect      
    Leptin receptor (LEPR) plays a vital role in obesity in humans and animals.  The objective of this study is to assess LEPR functional variants for chicken adipose deposition by integration of association and in-silico analysis using a unique chicken population, the Northeast Agricultural University broiler lines divergently selected for abdominal fat content (NEAUHLF).  Five online bioinformatics tools were used to predict the functionality of the single nucleotide polymorphisms (SNPs) in coding region.  Further, the possible structure–function relationship of high confidence SNPs was determined by bioinformatics analyses, including the conservation and stability analysis based on amino acid residues, prediction of protein ligand-binding sites, and the superposition of protein tertiary structure.  Meanwhile, we analyzed the association between abdominal fat traits and 20 polymorphisms of chicken LEPR gene.  The integrated results showed that rs731962924 (N867I) and rs13684622 (C1002R) could lead to striking changes in the structure and function of proteins, of which rs13684622 (C1002R) was significantly associated with abdominal fat weight (AFW, P=0.0413) and abdominal fat percentage (AFP, P=0.0260) in chickens.  Therefore, we are of the opinion that rs13684622 (C1002R) may be an essential functional SNP affecting chicken abdominal fat deposition, and potentially applied to improvement of broiler abdominal fat in molecular marker-assisted selection (MAS) program.  Additionally, the coupling of association with computer electronic predictive analysis provides a new avenue to identify important molecular markers for breeders.
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    Follicle-stimulating hormone is expressed in ovarian follicles of chickens and promotes ovarian granulosa cell proliferation
    BI Yu-lin, YANG Shu-yan, WANG Hai-yan, CHANG Guo-bin, CHEN Guo-hong
    2021, 20 (10): 2749-2757.   DOI: 10.1016/S2095-3119(21)63606-7
    Abstract95)      PDF in ScienceDirect      
    Follicle-stimulating hormone (FSH), an important hypothalamic-pituitary-gonadal axis (HPG) hormone, is secreted by the pituitary gland.  This study confirms that FSH is expressed in chicken follicles at different stages, and positive FSHβ mRNA signals were stronger (P<0.05) in granulosa cells than in oocytes.  The 369 bp coding sequence of FSHβ in ovaries is 100% identical to that in the pituitary gland.  The experiment in vitro revealed that the ovary possessed FSH secretory capacity.  Further, FSHβ mRNA was significantly upregulated (P<0.05) in follicles and significantly higher (P<0.05) than that in the pituitary gland by approximately 2–23 times with the development.  The number of granulosa cells decreased significantly (P<0.05) in the cells with siRNA treatment, confirming that the ovarian FSH could promote granulosa cell proliferation.  This view was supported by cell cycle analysis and CCND2 and CCNE2 expression.  Further research indicated that no difference (P>0.05) was observed between the number of granulosa cells treated with FSHβ siRNA and in exogenous FSH. However, the number of granulosa cells without FSHβ siRNA transfection was significantly higher (P<0.05) for exogenous FSH.  This finding suggests that the proliferative effect of exogenous FSH on ovarian granulosa cells depend on endogenous FSH.  This study demonstrated that the FSH gene was expressed in chicken follicles and promoted ovarian granulosa cell proliferation, which enriched the theory on HPG axis.
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    UBE2I stimulates female gonadal differentiation in chicken (Gallus gallus) embryos
    JIN Kai, ZHOU Jing, ZUO Qi-sheng, LI Jian-cheng, Jiuzhou SONG, ZHANG Ya-ni, CHANG Guo-bing, CHEN Guo-hong, LI Bi-chun
    2021, 20 (11): 2986-2994.   DOI: 10.1016/S2095-3119(20)63486-4
    Abstract66)      PDF in ScienceDirect      
    Without known analogous sex-determining factors like SRY (sex determining region Y) in mammals, the chicken (Gallus gallus) sex determination mechanism still remains unclear, which highly restricts the biological research on chicken development and poultry single-sex reproduction.  Here we not only characterized a new female-biased gene UBE2I and identified the expression pattern by qRT-PCR, but also described the functional role of UBE2I in the gonadal development of chicken embryos.  Results showed that UBE2I exhibited a female-biased expression pattern in the early stage of PGCs (primordial germ cells) in embryonic gonads and robust expression in ovaries of newborn chickens.  Most importantly, we successfully developed an effective method to interfere or overexpress UBE2I in chicken embryos through the intravascular injection.  The qRT-PCR analysis showed that the sex-related genes (FOXL2, CYP19A1 and HINTW) in females were upregulated (P<0.05) under the overexpression of UBE2I and the sex-related genes (SOX9, DMRT1 and WT1) in females were downregulated (P<0.05) after interfering UBE2I.  Furthermore, the change of UBE2I expression was associated with the level of estradiol and its receptors (AR and ESR), which suggests that UBE2I is necessary to initiate the female-specific development in chickens.  In conclusion, this work demonstrates that UBE2I is a crucial sex differentiation-related gene in the embryonic development of chickens, which provides insights for further understanding the mechanism of sex determination in chickens.
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    Epigenome-wide DNA methylation analysis reveals differentially methylation patterns in skeletal muscle between Chinese Chenghua and Qingyu pigs
    WANG Kai, WU Ping-xian, WANG Shu-jie, JI Xiang, CHEN Dong, JIANG An-an, XIAO Wei-hang, JIANG Yan-zhi, ZHU Li, ZENG Yang-shuang, XU Xu, QIU Xiao-tian, LI Ming-zhou, LI Xue-wei, TANG Guo-qing
    2022, 21 (6): 1731-1739.   DOI: 10.1016/S2095-3119(21)63814-5
    Abstract138)      PDF in ScienceDirect      
    Chenghua (CH) pig and Qingyu (QY) pig are typical Chinese native fatty breeds.  CH pig is mainly distributed in Chengdu Plain, while QY pig is widely distributed throughout the mountain areas around the Sichuan Basin.  There are significant differences in their phenotypic traits, including body image, growth performance, and meat quality.  This study compared several meat quality traits of CH and QY pigs and conducted a genome-wide DNA methylation analysis using reduced representation bisulfite sequencing (RRBS).  It was observed that the pH at 45 min (pH45min, P=5.22e–13), lightness at 45 min (L*45min, P=4.85e–5), and lightness at 24 h (L*24h, P=3.57e–5) of CH pigs were higher than those of QY pigs.  We detected 10 699 differentially methylated cytosines (DMCs) and 2 760 differentially methylated genes (DMGs) associated with these DMCs.  Functional analysis showed that these DMGs were mainly enriched in the AMPK signaling pathway, Type II diabetes mellitus, Insulin signaling pathway, mTOR signaling pathway, and Insulin resistance.  Furthermore, 15 DMGs were associated with fat metabolism (ACACA, CAB39, CRADD, CRTC2, FASN, and GCK), muscle development (HK2, IKBKB, MTOR, PIK3CD, PPARGC1A, and RPTOR), or meat quality traits (PCK1, PRKAG2, and SLC2A4).  The findings may help to understand further the epigenetic regulation mechanisms of meat quality traits in pigs and provide new basic data for the study of local pigs.
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    Dietary threonine deficiency affects expression of genes involved in lipid metabolism in adipose tissues of Pekin ducks in a genotype-dependent manner
    JIANG Yong, MA Xin-yan, XIE Ming, ZHOU Zheng-kui, TANG Jing, CHANG Guo-bin, CHEN Guo-hong, HOU Shui-sheng
    2022, 21 (9): 2691-2699.   DOI: 10.1016/j.jia.2022.07.011
    Abstract106)      PDF in ScienceDirect      
    Dietary threonine (Thr) deficiency increases hepatic triglyceride content and reduces sebum and abdominal fat percentages in lean type (LT), but not in fatty type (FT) Pekin ducks.  However, the molecular changes regarding the role of Thr in lipid metabolism in LT and FT ducks induced by Thr deficiency remains unknown.  This study compared differential expression gene profiles related to lipid metabolism in FT and LT Pekin ducks affected by Thr deficiency.  We performed transcriptomic profiling and scanned the gene expression in the liver, sebum, and abdominal fat of Pekin ducks fed either Thr-deficient or Thr-adequate diet for 21 days from 14 to 35 days of age.  There were 187, 52, and 50 differentially expressed genes (DEGs) identified in the liver, sebum, and abdominal fat of LT ducks affected by Thr deficiency, of which 12, 9, and 5 genes were involved in lipid metabolism, respectively.  Thr deficiency altered the expression of 27, 6, and 3 genes in FT ducks’ liver, sebum, and abdominal fat, respectively.  None of the DEGs had a relationship with lipid metabolism in FT ducks.  KEGG analysis showed that the DEGs in the LT ducks’ livers were enriched in lipid metabolism pathways (linolenic acid metabolism, glycerophospholipid metabolism, and arachidonic acid metabolism) and amino acid metabolism pathways (biosynthesis of amino acids, phenylalanine metabolism, β-alanine metabolism, and glycine, serine and threonine metabolisms).  The DEGs in the sebum and abdominal fat of LT ducks were not enriched in lipid and amino acid metabolic pathways.  Additionally, DEGs involved in lipid metabolism were found to be upregulated by Thr deficiency in LT ducks, such as malic enzyme 3 (ME3), acyl-CoA synthetase short-chain family member 2 (ACSS2) in liver, and lipase member M (LIPM) in sebum.  In summary, dietary Thr deficiency regulated the gene expression involved in lipid metabolism in the liver, sebum, and abdominal fat of Pekin ducks in a genotype-dependent manner.

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    Regulation of bone phosphorus retention and bone development possibly by BMP and MAPK signaling pathways in broilers
    LIAO Xiu-dong, CAO Su-mei, LI Ting-ting, SHAO Yu-xin, ZHANG Li-yang, LU Lin, ZHANG Ri-jun, HOU Shui-sheng, LUO Xu-gang
    2022, 21 (10): 3017-3025.   DOI: 10.1016/j.jia.2022.07.037
    Abstract147)      PDF in ScienceDirect      

    The bone morphogenetic protein (BMP) and mitogen-activated protein kinase (MAPK) signaling pathways play an important role in regulation of bone formation and development, however, it remains unclear that the effect of dietary different levels of non-phytate phosphorus (NPP) on these signaling pathways and their correlations with bone phosphorus (P) retention and bone development in broilers.  Therefore, this experiment was conducted to investigate the effect of dietary P supplementation on BMP and MAPK signaling pathways and their correlations with bone P retention and bone development in broilers.  A total of 800 one-day-old Arbor Acres male broilers were randomly allotted to 1 of 5 treatments with 8 replicates in a completely randomized design.  The 5 treatments of dietary NPP levels were 0.15, 0.25, 0.35, 0.45 and 0.55% or 0.15, 0.22, 0.29, 0.36 and 0.43% for broilers from 1 to 21 days of age or 22 to 42 days of age, respectively.  The results showed that extracellular signal-regulated kinase 1 (ERK1) mRNA expression in the tibia of broilers on days 14 and 28, phosphorylated-ERK1 (p-ERK1) on day 14, and BMP2 protein expression on days 28 and 42 decreased linearly (P<0.04), while c-Jun N-terminal kinase 1 (JNK1) mRNA expression on day 42 increased linearly (P<0.02) with the increase of dietary NPP level.  At 14 days of age, total P accumulation in tibia ash (TPTA), bone mineral concentration (BMC), bone mineral density (BMD), bone breaking strength (BBS) and tibia ash were negatively correlated (r=–0.726 to –0.359, P<0.05) with ERK1 and JNK1 mRNA as well as p-ERK1; tibia alkaline phosphatase (ALP) and bone gal protein (BGP) were positively correlated (r=0.405 to 0.665, P<0.01) with ERK1 mRNA and p-ERK1.  At 28 days of age, TPTA, BMC, BMD, BBS and tibia ash were negatively correlated (r=–0.518 to –0.370, P<0.05) with ERK1 mRNA and BMP2 protein, while tibia ALP was positively correlated (r=0.382 to 0.648, P<0.05) with them.  The results indicated that TPTA, BMC, BMD, BBS or tibia ash had negative correlations, while tibia ALP and BGP had positive correlations with ERK1 and JNK1 mRNAs, BMP2 protein and p-ERK1, suggesting that bone P retention and bone development might be regulated by BMP and MAPK signaling pathways in broiler chickens.

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    Targeted myostatin loss-of-function mutation increases type II muscle fibers in Meishan pigs
    QIAN Li-li, XIE Jing-yi, GAO Ting, CAI Chun-bo, JIANG Sheng-wang, BI Han-fang, XIE Shan-shan, CUI Wen-tao
    2022, 21 (1): 188-198.   DOI: 10.1016/S2095-3119(21)63669-9
    Abstract156)      PDF in ScienceDirect      
    Myostatin (MSTN) is a negative regulator of skeletal muscle growth and development.  The skeletal muscle in MSTN–/– mice is significantly hypertrophied, with muscle fiber type II increasing significantly while muscle fiber type I decreasing.  However, it is still not clear how the skeletal muscle types change in MSTN–/– pigs, and how the mechanism for MSTN regulates fiber types, especially in large animals like pigs.  This study conducted a comprehensive analysis of the composition of skeletal muscle fibers in MSTN–/– pigs produced in our laboratory.  It was observed that, compared with wild-type (WT) pigs, both the total mass of skeletal muscle and type IIb muscle fibers increased significantly (P<0.01), while the type I and type IIa muscle fibers decreased significantly (P<0.01), in MSTN–/– Meishan pigs.  In addition, to explore the influence of MSTN on muscle fiber type and its regulation mechanism in the embryonic stage, this study selected a few genes (Myf5, Mef2d, MyoD and Six1) associated with muscle fiber type and validated their expression by quantitative RT-PCR.  Herein, it was found that Myh7, Myh2, Myh4 and Myh1 can be detected in the skeletal muscle of pigs at 65 days of gestation (dg).  Compared with WT pigs, in MSTN–/– Meishan pigs, Myh7 decreased significantly (P<0.01), while Myh4 (P<0.001) and Myh1 (P<0.05) increased significantly.  Meanwhile, the increased expression of Myf5 (P<0.05), Mef2d (P<0.01) and Six1 (P<0.05) in MSTN–/– Meishan pigs suggested that MSTN should regulate the directional development of muscle fiber types in the early stage of embryonic development.  Thus, at the embryonic stage, the type II muscle fibers began to increase in MSTN–/– pigs.  These results can provide valuable information not only for pig meat quality improvement, but also for the study of human skeletal muscle development and disease treatment.  
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    A comprehensive evaluation of factors affecting the accuracy of pig genotype imputation using a single or multi-breed reference population
    ZHANG Kai-li, PENG Xia, ZHANG Sai-xian, ZHAN Hui-wen, LU Jia-hui, XIE Sheng-song, ZHAO Shu-hong, LI Xin-yun, MA Yun-long
    2022, 21 (2): 486-495.   DOI: 10.1016/S2095-3119(21)63695-X
    Abstract122)      PDF in ScienceDirect      
    Genotype imputation has become an indispensable part of genomic data analysis.  In recent years, imputation based on a multi-breed reference population has received more attention, but the relevant studies are scarce in pigs.  In this study, we used the Illumina PorcineSNP50 Bead Chip to investigate the variations of imputation accuracy with various influencing factors and compared the imputation performance of four commonly used imputation software programs.  The results indicated that imputation accuracy increased as either the validation population marker density, reference population sample size, or minor allele frequency (MAF) increased.  However, the imputation accuracy would have a certain extent of decrease when the pig reference population was a mixed group of multiple breeds or lines.  Considering both imputation accuracy and running time, Beagle 4.1 and FImpute are excellent choices among the four software packages tested.  This work visually presents the impacts of these influencing factors on imputation and provides a reference for formulating reasonable imputation strategies in actual pig breeding.
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    C-type natriuretic peptide stimulates chicken myoblast differentiation through NPRB/NPRC receptors and metabolism pathway
    HUANG Hua-yun, LIANG Zhong, LIU Long-zhou, LI Chun-miao, HUANG Zhen-yang, WANG Qian-bao, LI Shou-feng, ZHAO Zhen-hua
    2022, 21 (2): 496-503.   DOI: 10.1016/S2095-3119(21)63694-8
    Abstract115)      PDF in ScienceDirect      
    Skeletal muscle development is closely related with the amount of meat production and its quality in chickens.  Natriuretic peptides (NPs) play an important role in myotube formation and fat oxidation of skeletal muscle in animals.  The effect of C-type natriuretic peptide (CNP), an important member of the NPs, and its underlying molecular mechanisms in skeletal muscle are incompletely understood.  Treatment of myoblasts with CNP led to enhanced proliferation/differentiation and significantly upregulated (P<0.05) mRNA expression of the CNP receptors natriuretic peptide receptor B (NPRB) and the clearance receptor C (NPRC).  In cells exposed to CNP, 142 differentially expressed genes (84 up-regulation and 58 down-regulation) (P<0.05) were identified by RNA-sequencing compared with those in control cells.  Sixteen genes were significantly enriched (P<0.05) in the metabolic pathway, and six of them (phospholipase C β4, phospholipase C β2, phosphoglycerate mutase 1, creatine kinase B, peroxiredoxin 6 and CD38) were closely related to skeletal muscle development and differentially expressed.  In conclusion, CNP stimulated differentiation of myoblasts by upregulating expression of the NPRB and NPRC receptors and enriching key genes in the metabolic pathway.  
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    Investigation of Mitochondrial DNA genetic diversity and phylogeny of goats worldwide
    GUO Yi, GONG Ying, HE Yong-meng, YANG Bai-gao, ZHANG Wei-yi, CHEN Bo-er, HUANG Yong-fu, ZHAO Yong-ju, ZHANG Dan-ping, MA Yue-hui, CHU Ming-xing, E Guang-xin
    2022, 21 (6): 1830-1837.   DOI: 10.1016/S2095-3119(21)63882-0
    Abstract151)      PDF in ScienceDirect      
    Genetic diversity, population structure, and population expansion of goats worldwide (4 165 individuals from 196 breeds) were analyzed using published mitochondrial DNA (mtDNA) D_loop hypervariable region sequences. Results showed that 2 409 haplotypes and 301 polymorphic sites were present within the 401-bp length D_loop region, the nucleotide diversity (Pi) was 0.03471, and the haplotype diversity (Hd) was 0.9983. Phylogenetic analysis revealed that 98.92% of haplotypes were divided into six obvious clusters, consistent with the classification of the known mitochondrial haplogroups of goats. Haplogroup A accounted for the largest proportion (86%). Interestingly, two unknown divisions (Unknown I and Unknown II) were discovered from goats in Southwest China, suggesting that Southwest China has unique maternal haplogroups. Analysis of molecular variance (AMOVA) and the average number of pairwise differences between populations (PiXY) indicated that geographical variation was small but significant. Neutrality tests (Tajima’s D and Fu’s FS tests) and mismatch distribution showed that haplogroups B, C, and G had expansion histories. In addition, the phylogenetic relationship between domestic and wild goats suggested that Capra aegagrus is the most likely wild ancestor and may have participated in the domestication of ancestral populations of A, B, C, and F haplogroups. A meta-analysis on the mtDNA sequences of goats from international databases was conducted to analyze goats’ genetic diversity, population structure, and matrilineal system evolution worldwide. The results may help further understand the domestication history and gene flow of goats worldwide.
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    HBP1 inhibits chicken preadipocyte differentiation by activating the STAT3 signaling via directly enhancing JAK2 expression
    CHEN Hong-yan, CHENG Bo-han, MA Yan-yan, ZHANG Qi, LENG Li, WANG Shou-zhi, LI Hui
    2022, 21 (6): 1740-1754.   DOI: 10.1016/S2095-3119(21)63895-9
    Abstract170)      PDF in ScienceDirect      

    Obesity presents a serious threat to human health and broiler performance.  The expansion of adipose tissue is mainly regulated by the differentiation of preadipocytes.  The differentiation of preadipocytes is a complex biological process regulated by a variety of transcription factors and signaling pathways.  Previous studies have shown that the transcription factor HMG-box protein 1 (HBP1) can regulate the differentiation of mouse 3T3-L1 preadipocytes by activating the Wnt/β-catenin signaling pathway.  However, it is unclear whether HBP1 involved in chicken preadipocyte differentiation and which signaling pathways it regulates.  The aim of the current study was to explore the biological function and molecular regulatory mechanism of HBP1 in the differentiation of chicken preadipocytes.  The expression patterns of chicken HBP1 in abdominal adipose tissue and during preadipocyte differentiation were analyzed by RT-qPCR and Western blot.  The preadipocyte stably overexpressing HBP1 or knockout HBP1 and their control cell line were used to analyze the effect of HBP1 on preadipocyte differentiation by oil red O staining, RT-qPCR and Western blot.  Cignal 45-Pathway Reporter Array was used to screen the signal pathways that HBP1 regulates in the differentiation of chicken preadipocytes.  Chemical inhibitor and siRNA for signal transducer and activator of transcription 3 (STAT3) were used to analyze the effect of STAT3 on preadipocyte differentiation.  The preadipocyte stably overexpressing HBP1 was transfected by the siRNA of STAT3 or treated with a chemical inhibitor of STAT3 for the rescue experiment.  The results of gene expression analysis showed that the expression of HBP1 was related to abdominal fat deposition and preadipocyte differentiation in chickens.  The results of function gain and loss experiments indicated that overexpression/knockout of HBP1 in chicken preadipocytes could inhibit/promote (P<0.05) lipid droplet deposition and the expression of adipogenesis-related genes.  Mechanismlly, HBP1 activates (P<0.05) the signal transducer and activator of transcription 3 (STAT3) signaling pathway by targeting janus kinase 2 (JAK2) transcription.  The results of functional rescue experiments indicated that STAT3 signaling mediated the regulation of HBP1 on chicken preadipocyte differentiation.  In conclusion, HBP1 inhibits chicken preadipocyte differentiation by activating the STAT3 signaling pathway via directly enhancing JAK2 expression.  Our findings provided new insights for further analysis of the molecular genetic basis of chicken adipose tissue growth and development.

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    Integration of genome-wide association study and selection signatures reveals genetic determinants for skeletal muscle production traits in an F2 chicken population
    LI Yu-dong, BAI Xue, LIU Xin , WANG Wei-jia, LI Zi-wei, WANG Ning, XIAO Fan, GAO Hai-he, GUO Huai-shun, LI Hui, WANG Shou-zhi
    2022, 21 (7): 2065-2075.   DOI: 10.1016/S2095-3119(21)63805-4
    Abstract175)      PDF in ScienceDirect      
    Improving the production of broiler chicken meat has been a goal of broiler breeding programs worldwide for many years.  However, the genetic architectures of skeletal muscle production traits in chickens have not yet been fully elucidated.  In the present study, a total of 519 F2 birds, derived from a cross of Arbor Acres broiler and Baier layer, were re-sequenced (26 F0 individuals were re-sequenced at a 10-fold depth; 519 F2 individuals were re-sequenced at a 3-fold depth) and the coupling of genome-wide association study (GWAS) and selection signatures (FST (fixation index) and θπ (nucleotide diversity)) was carried out to pinpoint the associated loci and genes that contribute to pectoral muscle weight (PMW) and thigh muscle weight (TMW).  A total of 7 890 258 single nucleotide polymorphisms (SNPs) remained to be analyzed after quality control and imputation.  The integration of GWAS and selection signature analyses revealed that genetic determinants responsible for skeletal muscle production traits were mainly localized on chromosomes 1 (168.95–172.43 Mb) and 4 (74.37–75.23 Mb).  A total of 17 positional candidate genes (PCGs) (LRCH1, CDADC1, CAB39L, LOC112531568, LOC112531569, FAM124A, FOXO1, NBEA, GPALPP1, RUBCNL, ARL11, KPNA3, LHFP, GBA3, LOC112532426, KCNIP4, and SLIT2) were identified in these regions.  In particular, KPNA3 and FOXO1 were the most promising candidates for meat production in chickens.  These findings will help enhance our understanding of the genetic architecture of chicken muscle production traits, and the significant SNPs identified could be promising candidates for integration into practical breeding programs such as genome-wide selection (GS) to improve the meat yield of chickens.

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    Incorporating genomic annotation into single-step genomic prediction with imputed whole-genome sequence data
    TENG Jin-yan, YE Shao-pan, GAO Ning, CHEN Zi-tao, DIAO Shu-qi, LI Xiu-jin, YUAN Xiao-long, ZHANG Hao, LI Jia-qi, ZHANG Xi-quan, ZHANG Zhe
    2022, 21 (4): 1126-1136.   DOI: 10.1016/S2095-3119(21)63813-3
    Abstract123)      PDF in ScienceDirect      
    Single-step genomic best linear unbiased prediction (ssGBLUP) is now intensively investigated and widely used in livestock breeding due to its beneficial feature of combining information from both genotyped and ungenotyped individuals in the single model.  With the increasing accessibility of whole-genome sequence (WGS) data at the population level, more attention is being paid to the usage of WGS data in ssGBLUP.  The predictive ability of ssGBLUP using WGS data might be improved by incorporating biological knowledge from public databases.  Thus, we extended ssGBLUP, incorporated genomic annotation information into the model, and evaluated them using a yellow-feathered chicken population as the examples.  The chicken population consisted of 1 338 birds with 23 traits, where imputed WGS data including 5 127 612 single nucleotide polymorphisms (SNPs) are available for 895 birds.  Considering different combinations of annotation information and models, original ssGBLUP, haplotype-based ssGHBLUP, and four extended ssGBLUP incorporating genomic annotation models were evaluated.  Based on the genomic annotation (GRCg6a) of chickens, 3 155 524 and 94 837 SNPs were mapped to genic and exonic regions, respectively.  Extended ssGBLUP using genic/exonic SNPs outperformed other models with respect to predictive ability in 15 out of 23 traits, and their advantages ranged from 2.5 to 6.1% compared with original ssGBLUP.  In addition, to further enhance the performance of genomic prediction with imputed WGS data, we investigated the genotyping strategies of reference population on ssGBLUP in the chicken population.  Comparing two strategies of individual selection for genotyping in the reference population, the strategy of evenly selection by family (SBF) performed slightly better than random selection in most situations.  Overall, we extended genomic prediction models that can comprehensively utilize WGS data and genomic annotation information in the framework of ssGBLUP, and validated the idea that properly handling the genomic annotation information and WGS data increased the predictive ability of ssGBLUP.  Moreover, while using WGS data, the genotyping strategy of maximizing the expected genetic relationship between the reference and candidate population could further improve the predictive ability of ssGBLUP.  The results from this study shed light on the comprehensive usage of genomic annotation information in WGS-based single-step genomic prediction.

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    miR-99a-5p inhibits target gene FZD5 expression and steroid hormone secretion from goat ovarian granulosa cells
    ZHU Lu, JING Jing, QIN Shuai-qi, LU Jia-ni, ZHU Cui-yun, ZHENG Qi, LIU Ya, FANG Fu-gui, LI Yun-sheng, ZHANG Yun-hai, LING Ying-hui
    2022, 21 (4): 1137-1145.   DOI: 10.1016/S2095-3119(21)63766-8
    Abstract130)      PDF in ScienceDirect      
    MicroRNA (miRNA) has vital regulatory effects on the proliferation, differentiation and secretion of ovarian granulosa cells, but the role of miR-99a-5p in goat ovarian granulosa cells (GCs) is unclear.  Both miR-99a-5p and Frizzled-5 (FZD5) were found to be expressed in GCs in goat ovaries via fluorescence in situ hybridization and immunohistochemistry, respectively, and FZD5 was verified (P<0.001) as a target gene of miR-99a-5p by double luciferase reporter gene experiments.  Furthermore, FZD5 mRNA and protein expression were both found to be regulated (P<0.05) by miR-99a-5p in GCs.  Moreover, the overexpression of miR-99a-5p or knockdown of FZD5 suppressed (P<0.05) estradiol and progesterone secretion from the GCs, as determined by ELISA.  In summary, miR-99a-5p inhibits target gene FZD5 expression and estradiol and progesterone synthesis in GCs.  Our study thus provides seminal data and new insights into the regulatory mechanisms of follicular development in the goat and other animals.
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    NH4Cl promotes apoptosis and inflammation in bovine mammary epithelial cells via the circ02771/miR-194b/TGIF1 axis
    CHEN Zhi, LIANG Yu-sheng, ZONG Wei-cheng, GUO Jia-he, ZHOU Jing-peng, MAO Yong-jiang, JI De-jun, JIAO Pei-xin, Juan J LOOR, YANG Zhang-ping
    2022, 21 (4): 1161-1176.   DOI: 10.1016/S2095-3119(21)63812-1
    Abstract138)      PDF in ScienceDirect      
    Excess ammonia (NH3) in the circulation of dairy animals can reduce animal health and the quality of products for human consumption.  To develop effective prevention and treatment methods, it is essential to examine the molecular mechanisms through which excess NH3 may affect the mammary gland.  The present study used bovine mammary epithelial cells (BMECs) to evaluate the effects of exogenous NH4Cl on the abundance of circular RNAs (circRNAs) using high-throughput sequencing.  Among the identified circRNAs, circ02771 was the most significantly upregulated by exogenous NH4Cl (P<0.05), with a fold change of 4.12.  The results of the apoptosis and proliferation assays, transmission electron microscopy, H&E staining, and immunohistochemistry revealed that circ02771 increased apoptosis and inflammation.  A double luciferase reporter assay revealed that circ02771 targeted miR-194b, and the overexpression of circ02771 (pcDNA-circ02771) reduced (P<0.05) the expression of miR-194b and led to apoptosis and inflammation.  Circ02771 also enhanced the expression of transforming growth factor beta-induced factor homeobox 1 (TGIF1), which is a target gene of miR-194b.  Overall, this study suggests that the circ02771/miR-194b/TGIF1 axis plays a role in mediating the effects of NH4Cl on BMECs.  Therefore, this axis provides a novel target to help control hazards within the mammary gland from high circulating NH4Cl levels.
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    Transcriptomic analysis elucidates the enhanced skeletal muscle mass, reduced fat accumulation, and metabolically benign liver in human follistatin-344 transgenic pigs
    LONG Ke-ren, LI Xiao-kai, ZHANG Ruo-wei, GU Yi-ren, DU Min-jie, XING Xiang-yang, DU Jia-xiang, MAI Miao-miao, WANG Jing, JIN Long, TANG Qian-zi, HU Si-lu, MA Ji-deng, WANG Xun, PAN Deng-ke, LI Ming-zhou
    2022, 21 (9): 2675-2690.   DOI: 10.1016/j.jia.2022.07.014
    Abstract216)      PDF in ScienceDirect      

    Follistatin (FST) is an important regulator of skeletal muscle growth and adipose deposition through its ability to bind to several members of the transforming growth factor-β (TGF-β) superfamily, and thus may be a good candidate for future animal breeding programs.  However, the molecular mechanisms underlying the phenotypic changes have yet to be clarified in pig.  We generated transgenic (TG) pigs that express human FST specifically in skeletal muscle tissues and characterized the phenotypic changes compared with the same tissues in wild-type pigs.  The TG pigs showed increased skeletal muscle growth, decreased adipose deposition, and improved metabolism status (P<0.05).  Transcriptome analysis detected important roles of the PIK3–AKT signaling pathway, calcium-mediated signaling pathway, and amino acid metabolism pathway in FST-induced skeletal muscle hypertrophy, and depot-specific oxidative metabolism changes in psoas major muscle.  Furthermore, the lipid metabolism-related process was changed in adipose tissue in the TG pigs.  Gene set enrichment analysis revealed that genes related to lipid synthesis, lipid catabolism, and lipid storage were down-regulated (P<0.01) in the TG pigs for subcutaneous fat, whereas genes related to lipid catabolism were significantly up-regulated (P<0.05) in the TG pigs for retroperitoneal fat compared with their expression levels in wild-type pigs.  In liver, genes related to the TGF-β signaling pathway were over-represented in the TG pigs, which is consistent with the inhibitory role of FST in regulating TGF-β signaling.  Together, these results provide new insights into the molecular mechanisms underlying the phenotypic changes in pig.

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    The expression, function, and coding potential of circular RNA circEDC3 in chicken skeletal muscle development
    WEI Yuan-hang, ZHAO Xi-yu, SHEN Xiao-xu, YE Lin, ZHANG Yao, WANG Yan, LI Di-yan, ZHU Qing, YIN Hua-dong
    2022, 21 (5): 1444-1456.   DOI: 10.1016/S2095-3119(21)63826-1
    Abstract93)      PDF in ScienceDirect      
    As an emerging class of non-coding transcripts, circular RNAs (circRNAs) are proved to participate in the complex process of myogenesis in diverse species.  A previous study has identified circular RNA EDC3 (circEDC3) as a typical covalently closed circular RNA abundant in chicken skeletal muscle.  This study found that circEDC3 is a conservative circular RNA and performed functional analysis to investigate the role of circEDC3 in chicken muscle growth.  The results indicated that circEDC3 could inhibit (P<0.05) chicken skeletal muscle satellite cells (SMSCs) proliferation and differentiation but had no significant influence on SMSCs apoptosis.  Additionally, bioinformatics analysis showed that circEDC3 had promising coding potential.  The open reading frames (ORF) were found in circEDC3 in this study.  Furthermore, this study predicted that circEDC3 had internal ribosome entry sites (IRES) and N6-methyladenosine (m6A) motifs in different species, implying that circEDC3 might be translatable.  This study revealed that circEDC3 might be a negative regulator in chicken muscle development and suggested it has protein-coding potential in different species.
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    Integrative analysis of hypothalamic transcriptome and genetic association study reveals key genes involved in the regulation of egg production in indigenous chickens
    WANG Dan-dan, ZHANG Yan-yan, TENG Meng-lin, WANG Zhang, XU Chun-lin, JIANG Ke-ren, MA Zheng, LI Zhuan-jian, TIAN Ya-dong, Kang Xiang-tao, LI Hong, LIU Xiao-jun
    2022, 21 (5): 1457-1474.   DOI: 10.1016/S2095-3119(21)63842-X
    Abstract192)      PDF in ScienceDirect      
    Indigenous chicken products are increasingly favored by consumers due to their unique meat and egg quality.  However, the relatively poor egg-laying performance largely impacts the economic benefits and hinders sustainable development of the local chicken industry.  Thus, excavating key genes and effective molecular markers associated with egg-laying performance is necessary to improve egg production via genetic selection in indigenous breeds.  In the present study, comparative hypothalamic transcriptome between pre-laying (15 weeks old) and peak-laying (30 weeks old) Lushi blue-shelled-egg (LBS) chicken was performed.  A total of 518 differentially expressed genes (DEGs) were identified.  Among the DEGs, 64 genes were enriched in 10 Gene Ontology (GO) terms associated with reproductive regulation via GO analysis and considered as potential candidate genes regulating egg-laying performance.  Of the 64 genes, 16 showed high connectivity (degree≥12) by protein–protein interaction (PPI) network analysis and were considered as potential core candidate genes (PCCGs).  To further look for key candidate genes from the PCCGs, firstly, the expression patterns of the 16 genes were examined in the hypothalamus of two indigenous breeds (LBS and Gushi (GS) chickens) between the pre-laying and peak-laying stages using quantitative real-time PCR (qRT-PCR).  Eleven out of the 16 genes showed significantly differential expression (P<0.05) with the same changing trends in the two breeds.  Then, correlations between the expression levels of the above 11 genes and egg numbers and reproductive hormone concentrations in serum were investigated in high-yielding and low-yielding GS chickens.  Of the 11 genes, eight showed significant correlations (P<0.05) between their expression levels and egg numbers, and between expression levels and reproductive hormone concentration in serum.  Furthermore, an association study on single nucleotide polymorphisms (SNPs) identified in these eight genes and egg production traits was carried out in 640 GS hens, and a significant association (P<0.05) between the SNPs and egg numbers was confirmed.  In conclusion, the eight genes, including CNR1, AP2M1, NRXN1, ANXA5, PENK, SLC1A2, SNAP25 and TRH, were demonstrated as key genes regulating egg production in indigenous chickens, and the SNPs sites within the genes might be served as markers to provide a guide for indigenous chicken breeding.  These findings provide a novel insight for further understanding the regulatory mechanisms of egg-laying performance and developing molecular markers to improve egg production of indigenous breeds.
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    MiR-140 downregulates fatty acid synthesis by targeting transforming growth factor alpha (TGFA) in bovine mammary epithelial cells
    CHU Shuang-feng, ZHAO Tian-qi, Abdelaziz Adam Idriss ARBAB, YANG Yi, CHEN Zhi, YANG Zhang-ping
    2022, 21 (10): 3004-3016.   DOI: 10.1016/j.jia.2022.07.039
    Abstract237)      PDF in ScienceDirect      

    Fat is an indispensable nutrient and basic metabolite for sustaining life, and milk is particularly rich in fatty acids, including a variety of saturated and unsaturated fatty acids.  MicroRNA (miRNA) and mRNA play an important role in the regulation of milk fat metabolism in mammary gland tissue.  It has been shown that lipid metabolism has a complex transcriptional regulation, but the mechanism by which milk fat synthesis is regulated through miRNA–mRNA interactions is poorly understood.  In this study, we performed transcriptome sequencing with bovine mammary gland tissue in the late lactation (270 and 315 days after parturition) to identify the key gene that regulating milk fat metabolism.  A total of 1 207 differentially coexpressed genes were selected, 828 upregulated genes and 379 downregulated genes were identified.  The transforming growth factor alpha (TGFA) gene was selected as the target gene, and luciferase reporter assay, Western blotting and qRT-PCR were used for further study.  The results demonstrated that miR-140 was an upstream regulator of TGFA, and miR-140 could inhibit (P<0.01) unsaturated fatty acid and triglyceride (TAGs) production in bovine mammary epithelial cells (BMECs).  In contrast, TGFA promoted (P<0.01) unsaturated fatty acid and TAG production.  Rescue experiments further indicated the miR-140/TGFA regulatory mechanism.  Taken together, these results suggest that the miR-140/TGFA pathway can inhibit (P<0.01) milk fat metabolism and improve milk quality by genetic means.

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    Genome-wide detection for runs of homozygosity analysis in three pig breeds from Chinese Taihu Basin and Landrace pigs by SLAF-seq data

    TONG Shi-feng, ZHU Mo , XIE Rui , LI Dong-feng , ZHANG Li-fan , LIU Yang
    2022, 21 (11): 3293-3301.   DOI: 10.1016/j.jia.2022.08.061
    Abstract175)      PDF in ScienceDirect      

    Erhualian (E), Meishan (MS) and Mi (MI) pigs are excellent indigenous pig breeds in Chinese Taihu Basin, which have made great contributions to the genetic improvement of commercial pigs.  Investigation of the genetic structure and inbreeding level of the 3 pig breeds is of great significance for the sustainable breeding of commercial pigs.  The length and number of runs of homozygosity (ROH) as well as the frequency of genomes covered by ROH can be used as indicators to evaluate the level of inbreeding and the origin of the population.  In this study, the ROH characteristics of E, MS, MI and Landrace (L) pigs were analyzed by SLAF-seq data, and the inbreeding coefficient based on ROH (FROH) was calculated.  In addition, we have identified candidate genes in the genomic regions associated with ROH.  A total of 10 568 ROH were detected in 116 individuals of 4 pig breeds.  The analysis showed that there were significant differences in genetic structure between 3 Taihu Basin pig breeds and L, and the genetic structure of E and MI was similar.  The results of FROH showed that the inbreeding level of MS was the highest (0.25±0.07), while E and MI were lower than L.  Compared with the other 3 pig populations, MS showed a higher frequency of long ROH (>5 Mb), indicating higher inbreeding in MS in recent generations.  A large number of candidate genes related to reproductive traits are located in the genomic regions with a high frequency of ROH, and these genes are expected to be used as candidate genes in marker-assisted selection (MAS) breeding programs.  Our findings can provide theoretical support for genetic conservation and genetic improvement of 3 pig breeds in Chinese Taihu Basin.

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    PPAR gamma2: The main isoform of PPARγ that positively regulates the expression of the chicken Plin1 gene
    SUN Yu-hang, ZHAI Gui-ying, PANG Yong-jia, LI Rui, LI Yu-mao, CAO Zhi-ping, WANG Ning, LI Hui, WANG Yu-xiang
    2022, 21 (8): 2357-2371.   DOI: 10.1016/S2095-3119(21)63896-0
    Abstract101)      PDF in ScienceDirect      

    Perilipin1 (PLIN1) is a major phosphorylated protein that specifically coats the surface of neutral lipid droplets (LDs) in adipocytes and plays a crucial role in regulating the accumulation and hydrolysis of triacylglycerol (TG).  Mammalian studies have shown that Plin1 gene transcription is mainly regulated by peroxisome proliferator-activated receptor-gamma (PPARγ), the master regulator of adipogenesis.  However, the regulatory mechanism of the chicken Plin1 (cPlin1) gene is poorly understood.  The present study aimed to investigate whether Plin1 is regulated by PPARγ in chickens and identify its exact molecular mechanism.  Reporter gene and expression assays showed that PPARγ2, but not PPARγ1, activated (P<0.01) the cPlin1 gene promoter.  An electrophoretic mobility shift assay and mutational analysis revealed that PPARγ2 bound to a special site in the cPlin1 gene promoter to enhance its expression.  In summary, our results show that PPARγ promotes the expression of the cPlin1 gene and that PPARγ2 is the main regulatory isoform.

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    Transcriptome analysis of the spleen of heterophils to lymphocytes ratio-selected chickens revealed their mechanism of differential resistance to Salmonella
    WANG Jie, ZHANG Qi, Astrid Lissette BARRETO SÁNCHEZ, ZHU Bo, WANG Qiao, ZHENG Mai-qing, LI Qing-he, CUI Huan-xian, WEN Jie, ZHAO Gui-ping
    2022, 21 (8): 2372-2383.   DOI: 10.1016/S2095-3119(21)63770-X
    Abstract94)      PDF in ScienceDirect      

    Salmonella is one of the most common food-borne pathogens and its resistance in chicken can be improved through genetic selection.  The heterophils/lymphocytes (H/L) ratio in the blood reflects the immune system status of chicken.  We compared the genome data and spleen transcriptomes between the H/L ratio-selected and non-selected chickens, after Salmonella infection, aiming to identify the key genes participating in the antibacterial activity in the spleen.  The results revealed that, the selected population had stronger (P<0.05) liver resistance to Salmonella typhimurium (ST) than the non-selected population.  In the selected and non-selected lines, the identified differentiation genes encode proteins involved in biological processes or metabolic pathways that included the TGF-beta signaling pathway, FoxO signaling pathway, and Salmonella infection pathway.  The results of the analysis of all identified differentially expressed genes (DEGs) of spleen revealed that the G protein-coupled receptor (GPCR) and insulin-like growth factor (IGF-I) signaling pathways were involved in the Salmonella infection pathway.  Integrated analysis of DEGs and FST (fixation index), identified candidate genes involved in Salmonella infection pathway, such as GPR39, NTRK2, and ANXA1.  The extensive genomic changes highlight the polygenic genetic of the immune response in these chicken populations.  Numerous genes related to the immune performance are differentially expressed in the selected and non-selected lines and the selected lines has a higher resistance to Salmonella. 

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    Weighted gene co-expression network analysis identifies potential regulators in response to Salmonella Enteritidis challenge in the reproductive tract of laying ducks
    ZHANG Yu, LUO Shu-wen, HOU Li-e, GU Tian-tian, ZHU Guo-qiang, Wanwipa VONGSANGNAK, XU Qi, CHEN Guo-hong
    2022, 21 (8): 2384-2398.   DOI: 10.1016/S2095-3119(21)63888-1
    Abstract129)      PDF in ScienceDirect      

    Salmonella Enteritidis (SE) is a zoonotic and vertically transmitted pathogen, often colonized in the reproductive tract of adult poultry, which can result in direct contamination of eggs and threaten human health.  Previous studies have revealed that some pattern recognition receptors and resistance genes were involved in regulating immune responses to SE invasion in birds.  However, the role of these immune response genes was not independent, and the interactions among the genes remained to be further investigated.  In this study, SE burden and colonization were determined in reproductive tissue after the ducks were SE-infected, and RNA-sequencing was performed to construct co-expression networks by weighted gene co-expression network analysis (WGCNA).  The result showed that SE could be isolated from 22% of infected-birds in any segment of the reproductive tract and the SE was readily colonized in the stroma, small follicle, isthmus, and vagina of the reproductive tracts in morbid ducks.  The top central, highly connected genes were subsequently identified three specific modules in the above four tissues at the defined cut-offs (P<0.01), including 60 new candidate regulators and 125 transcription factors.  Moreover, those 185 differentially expressed genes (DEGs) in these modules were co-expressed.  Moreover, the hub genes (TRAF3, CXCR4 and IL13RA1) were identified to act with many other genes through immune response pathways including NF-kappaB, Toll-like receptor, steroid biosynthesis, and p53 signaling pathways.  These data provide references that will understand the immune regulatory relationships during SE infection, but also assist in the breeding of SE-resistant lines through potential biomarkers.

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