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Use of transcriptome sequencing to explore the effect of CSRP3 on chicken myoblasts

SHAN Yan-ju, JI Gai-ge, ZHANG Ming, LIU Yi-fan, TU Yun-jie, JU Xiao-jun, SHU Jing-ting, ZOU Jian-min

2023, 22 (4): 1159-1171. DOI: 10.1016/j.jia.2022.08.067

Abstract （343） PDF in ScienceDirect

The mechanisms that regulate the specificity and maintenance of chicken muscle fiber types remain largely unknown. In mammals, CSRP3 has been shown to play a vital role in the maintenance of typical muscle structure and function. This study investigated the role that CSRP3 plays in chicken skeletal muscle. First, the antibody against chicken CSRP3 protein was prepared, and the expression levels of the mRNA and protein of the CSRP3 gene in four chicken skeletal muscles with different myofiber compositions were compared. Then the effects of CSRP3 silencing on the expression profile of chicken myoblast transcriptomes were analyzed. The results showed that the expression levels of the mRNA and protein of the CSRP3 gene were both associated with the composition of fiber types in chicken skeletal muscles. A total of 650 genes with at least 1.5-fold differences (Q<0.05) were identified, of which 255 genes were upregulated and 395 genes were downregulated by CSRP3 silencing. Functional enrichment showed that several pathways, including adrenergic signaling in cardiomyocytes, adipocytokine signaling pathway and apelin signaling pathway, were significantly (P<0.05) enriched both in differentially expressed genes and all expressed genes. The co-expressed gene network suggested that CSRP3 silencing caused a compensatory upregulation (Q<0.05) of genes related to the assembly of myofibrils, muscle differentiation, and contraction. Meanwhile, two fast myosin heavy chain genes (MyH1B and MyH1E) were upregulated (Q<0.05) upon CSRP3 silencing. These results suggested that CSRP3 plays a crucial role in chicken myofiber composition, and affects the distribution of chicken myofiber types, probably by regulating the expression of MyH1B and MyH1E.

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PGC-1α differentially regulates the mRNA expression profiles of genes related to myofiber type specificity in chicken

SHAN Yan-ju, JI Gai-ge, ZOU Jian-min, ZHANG Ming, TU Yun-jie, LIU Yi-fan, JU Xiao-jun, SHU Jing-ting

2020, 19 (8): 2083-2094. DOI: 10.1016/S2095-3119(20)63177-X

Abstract （122） PDF in ScienceDirect

Previous studies on mammals showed that peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α) played a prominent role in regulating muscle fiber type transition and composition. However, the role of PGC-1α in chicken muscle has seldom been explored. To investigate the effect of PGC-1α on chicken skeletal muscles in this study, the PGC-1α gene was overexpressed or silenced in chicken primary myoblasts by using lentivirus, and then the effects of the PGC-1α gene overexpression and knockdown on the mRNA expression profile of genes related to myofiber type specificity were examined during fiber formation. The results showed that overexpression of PGC-1α from proliferation to differentiation was accompanied by the up-regulated expression of Pax7, MyoD, and CnAα, which was significantly (P<0.01) increased after one day of transfection (1I). The enhancement of MyoG, MEF2c, and MyHC SM expression lagged, which was improved significantly (P<0.01) after four days of transfection (1I3D). Overexpression of PGC-1α decreased (P<0.01) the MyHC FWM expression after four days of transfection (1I3D), and it had no significant impact (P>0.05) on the expression of CnB1, NFATc3, and MyHC FRM during myofiber formation. The effective silence (P<0.01) of PGC-1α by lentivirus mediating short hairpin RNA (shRNA) was detected after four days of transfection (1I3D) in cultures, and the lack of its function in chicken primary myoblasts significantly (P<0.01) down-regulated the expression of Pax7, MyoD, CnAα, MyoG, MEF2c, and MyHC SM, significantly (P<0.01) up-regulated the expression of MyHC FWM, and had no significant impact (P>0.05) on the expression of CnB1, NFATc3, and MyHC FRM. These results indicated that the role of PGC-1α in regulating the fiber type specificity of chicken skeletal muscles might be similar to that in mammals, which interplayed with key genes related to myocyte differentiation and calcineurin signaling pathway.

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Myofiber development during embryonic to neonatal development in duck breeds differing in muscle growth rates

LI Hui-fang, SHU Jing-ting, SHAN Yan-ju, CHEN Wen-feng, SONG Chi, XU Wen-juan

2016, 15 (2): 403-413. DOI: 10.1016/S2095-3119(14)60949-7

Abstract （1747） PDF in ScienceDirect

Little is known about the muscle developmental patterns during embryonic to neonatal development in ducks. We investigated the developmental patterns in the lateral gastrocnemius muscles of Gaoyou and Jinding ducks differing in their muscle growth rates during the final stages of egg incubation and the first week after hatching. Expression of the MyoD gene was quantified by quantitative real-time PCR (qRT-PCR). The average cross-sectional area and diameter of the fibers increased from embryonic day 21 (E21), peaking at E27, and then declining slightly 7 d after hatching. The density of the fibers decreased initially but increased after hatching in both breeds and sexes. The within-breed variation in muscle fiber-type composition was greater than the average variation between the breeds. Overall, the percentage of type I fibers increased and that of type IIb fibers decreased consistently. However, the percentage of type IIa fibers was almost constant as development proceeded in both duck breeds. The profiles of MyoD mRNA expression were similar in both breeds, and a significantly positive relationship was observed between the expression of MyoD and the percentage of type IIb fibers. This study firstly revealed the characteristics of duck muscle development and differences between the two breeds differing in growth rates. Moreover, type IIb fibers might convert to type I fibers in the lateral gastrocnemius, while MyoD may potentially function in controlling the muscle fiber phenotype during the secondary myogenesis of muscle development.

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Differentiation of expression profiles of two calcineurin subunit genes in chicken skeletal muscles during early postnatal growth depending on anatomical location of muscles and breed

SHAN Yan-ju, XU Wen-juan, SHU Jing-ting, ZHANG Ming, SONG Wei-tao, TAO Zhi-yun, ZHU Chunhong, LI Hui-fang

2016, 15 (05): 1085-1094. DOI: 10.1016/S2095-3119(15)61162-5

Abstract （1780） PDF in ScienceDirect

Calcineurin (Cn or CaN) is implicated in the control of skeletal muscle fiber phenotype and hypertrophy. However, little information is available concerning the expression of Cn in chickens. In the present study, the expression of two Cn subunit genes (CnAα and CnB1) was quantified by qPCR in the lateral gastrocnemius (LG, mainly composing of red fast-twitch myofibers), the soleus (mainly composing of red slow-twitch myofibers) and the extensor digitorum longus (EDL, mainly composing of white fast-twitch myofibers) from Qingyuan partridge chickens (QY, slow-growing chicken breed) and Recessive White chickens (RW, fast-growing chicken breed) on different days (1, 8, 22, 36, 50 and 64 days post-hatching). Although CnAα and CnB1 gene expressions were variable with different trends in different skeletal muscles in the two chicken breeds during postnatal growth, it is highly muscle phenotype and breed specific. In general, the levels of CnAα and CnB1 gene expressions of the soleus were lower than those of EDL and LG in both chicken breeds at the same stages. Compared between the two chicken breeds, the levels of CnAα gene expression of the three skeletal muscles in QY chickens were higher than those in RW chickens on days 1 and 22. However, on day 64, the levels of both CnAα and CnB1 gene expressions of the three skeletal muscles were lower in QY chickens than those in RW chickens. Correlation analysis of the levels of CnAα and CnB1 gene expressions of the same skeletal muscle showed that there were positive correlations for all three skeletal muscle tissues in two chicken breeds. These results provide some valuable clues to understand the role of Cn in the development of chicken skeletal muscles, with a function that may be related to meat quality.

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