园艺作物基因功能与分子调控机制Horticulture — Gene function · Molecular mechanism
The OVATE family proteins (OFPs) are plant-specific proteins that modulate diverse aspects of plant growth and development. In tomato, OFP20 has been shown to interact with TONNEAU1 Recruiting Motif (TRM) proteins to regulate fruit shape. In this study, we demonstrated that the mutation of StOFP20 caused a shift from round to oval shaped tubers in a diploid accession C151, supporting the role of StOFP20 in controlling tuber shape. Its expression reached a maximum in the tuber initiation stage and then decreased as the tuber develops. To help elucidate the mechanism of tuber shape regulation by StOFP20, 27 TONNEAU1 Recruiting Motif (TRM) proteins were identified and 23 of them were successfully amplified in C151. A yeast two-hybrid assay identified three TRM proteins that interacted with StOFP20, which was confirmed by firefly luciferase complementation in tobacco leaves. The OVATE domain was indispensable for the interactions, while the necessity of the M10 motif in TRM proteins varied among the interactions between StOFP20 and the three TRMs. In summary, both StOFP20 and SlOFP20 directed interactions with TRM proteins, but the corresponding interactants were not completely consistent, implying that they exert regulatory roles through mechanisms that are only partially overlapping.
Early defoliation, which usually occurs during summer in pear trees, is gradually becoming a major problem that poses a serious threat to the pear industry in southern China. However, there is no system for evaluating the responses of different cultivars to early defoliation, and our knowledge of the potential molecular regulation of the genes underlying this phenomenon is still limited. In this study, we conducted field investigations of 155 pear accessions to assess their resistance or susceptibility to early defoliation. A total of 126 accessions were found to be susceptible to early defoliation, and only 29 accessions were resistant. Among them, 19 resistant accessions belong to the sand pear species (Pyrus pyrifolia). To identify the resistance genes related to early defoliation, the healthy and diseased samples of two sand pear accessions, namely, the resistant early defoliation accession ‘Whasan’ and the susceptible early defoliation accession ‘Cuiguan’, were used to perform RNA sequencing. Compared with ‘Cuiguan’, a total of 444 genes were uniquely differentially expressed in ‘Whasan’. Combined with GO and KEGG enrichment analyses, we found that early defoliation was closely related to the stress response. Furthermore, a weighted gene co-expression network analysis revealed a high correlation of WRKY and ethylene responsive factor (ERF) transcription factors with early defoliation resistance. This study provides useful resistant germplasm resources and new insights into potentially essential genes that respond to early defoliation in pears, which may facilitate a better understanding of the resistance mechanism and molecular breeding of resistant pear cultivars
PpMAPK6 regulates peach bud endodormancy release through interactions with PpDAM6
The MADS-box (DAM) gene PpDAM6, which is related to dormancy, plays a key role in bud endodormancy release, and the expression of PpDAM6 decreases during endodormancy release. However, the interaction network that governs its regulation of the endodormancy release of flower buds in peach remains unclear. In this study, we used yeast two-hybrid (Y2H) assays to identify a mitogen-activated protein kinase, PpMAPK6, that interacts with PpDAM6 in a peach dormancy-associated SSHcDNA library. PpMAPK6 is primarily located in the nucleus, and Y2H and bimolecular fluorescence complementation (BiFC) assays verified that PpMAPK6 interacts with PpDAM6 by binding to the MADS-box domain of PpDAM6. Quantitative real-time PCR (qRT-PCR) analysis showed that the expression of PpMAPK6 was opposite that of PpDAM6 in the endodormancy release of three cultivars with different chilling requirements (Prunus persica ‘Chunjie’, Prunus persica var. nectarina ‘Zhongyou 5’, Prunus persica ‘Qingzhou peach’). In addition, abscisic acid (ABA) inhibited the expression of PpMAPK6 and promoted the expression of PpDAM6 in flower buds. The results indicated that PpMAPK6 might phosphorylate PpDAM6 to accelerate its degradation by interacting with PpDAM6. The expression of PpMAPK6 increased with decreasing ABA content during endodormancy release in peach flower buds, which in turn decreased the expression of PpDAM6 and promoted endodormancy release.
Sugar content is a determinant of apple (Malus×domestica Borkh.) sweetness. However, the molecular mechanism underlying sucrose accumulation in apple fruit remains elusive. Herein, this study reported the role of the sucrose transporter MdSUT2.1 in the regulation of sucrose accumulation in apples. The MdSUT2.1 gene encoded a protein with 612 amino acid residues that could be localized at the plasma membrane when expressed in tobacco leaf protoplasts. MdSUT2.1 was highly expressed in fruit and was positively correlated with sucrose accumulation during apple fruit development. Moreover, complementary growth assays in a yeast mutant validated the sucrose transport activity of MdSUT2.1. MdSUT2.1 overexpression in apples and tomatoes resulted in significant increases in sucrose, fructose, and glucose contents compared to the wild type (WT). Further analysis revealed that the expression levels of sugar metabolism- and transport-related genes SUSYs, NINVs, FRKs, HXKs, and TSTs increased in apples and tomatoes with MdSUT2.1 overexpression compared to WT. Finally, unlike the tonoplast sugar transporters MdTST1 and MdTST2, the promoter of MdSUT2.1 was not induced by exogenous sugars. These findings provide valuable insights into the molecular mechanism underlying sugar accumulation in apples.
Arabinogalactan proteins (AGPs) are widely distributed in the plant kingdom and play a vital role during the process of plant sexual reproduction. In this study, we performed a comprehensive identification of the PbrAGPs expressed in pear pollen and further explored their influences on pollen tube growth. Among the 187 PbrAGPs that were found to be expressed in pear pollen tubes, 38 PbrAGPs were specifically expressed in pollen according to the RNA-seq data. The PbrAGPs were divided into two groups of highly expressed and specifically expressed in pear pollen. We further tested their expression patterns using RT-PCR and RT-qPCR. Most of the PbrAGPs were expressed in multiple tissues and their expression levels were consistent with reads per kilobase per million map reads (RPKM) values during pollen tube growth, implying that PbrAGPs might be involved in the regulation of pear pollen tube growth. We also constructed phylogenetic trees to identify the functional genes in pear pollen tube growth. Therefore, 19 PbrAGPs (PbrAGP1 to PbrAGP19) were selected to test their influences on pollen tube growth. Recombinant proteins of the 19 PbrAGP-His were purified and used to treat pear pollen, and 11 of the PbrAGP-His recombinant proteins could promote pear pollen tube growth. Additionally, pollen tube growth was inhibited when the expression levels of PbrAGP1 and PbrAGP5 were knocked down using an antisense oligonucleotide assay. PbrAGP1 and PbrAGP5 were localized in the plasma membrane and might not alter the distribution of pectin in the pollen tube. In summary, this study identified the PbrAGPs expressed in pear pollen and lays the foundation for further exploring their functions in pollen tube growth.
Small auxin up RNA (SAUR) is a large gene family that is widely distributed among land plants. In this study, a comprehensive analysis of the SAUR family was performed in sweet cherry, and the potential biological functions of PavSAUR55 were identified using the method of genetic transformation. The sweet cherry genome encodes 86 SAUR members, the majority of which are intron-less. These genes appear to be divided into seven subfamilies through evolution. Gene duplication events indicate that fragment duplication and tandem duplication events occurred in the sweet cherry. Most of the members mainly underwent purification selection pressure during evolution. During fruit development, the expression levels of PavSAUR16/45/56/63 were up-regulated, and conversely, those of PavSAUR12/61 were down-regulated. Due to the significantly differential expressions of PavSAUR13/16/55/61 during the fruitlet abscission process, they might be the candidate genes involved in the regulation of physiological fruit abscission in sweet cherry. Overexpression of PavSAUR55 in Arabidopsis produced earlier reproductive growth, root elongation, and delayed petal abscission. In addition, this gene did not cause any change in the germination time of seeds and was able to increase the number of lateral roots under abscisic acid (ABA) treatment. The identified SAURs of sweet cherry play a crucial role in fruitlet abscission and will facilitate future insights into the mechanism underlying the heavy fruitlet abscission that can occur in this fruit crop.
Sucrose phosphate synthase (SPS) is a rate-limiting enzyme that works in conjunction with sucrose-6-phosphate phosphatase (SPP) for sucrose synthesis, and it plays an essential role in energy provisioning during growth and development in plants as well as improving fruit quality. However, studies on the systematic analysis and evolutionary pattern of the SPS gene family in apple are still lacking. In the present study, a total of seven MdSPS and four MdSPP genes were identified from the Malus domestica genome GDDH13 v1.1. The gene structures and their promoter cis-elements, protein conserved motifs, subcellular localizations, physiological functions and biochemical properties were analyzed. A chromosomal location and gene-duplication analysis demonstrated that whole-genome duplication (WGD) and segmental duplication played vital roles in MdSPS gene family expansion. The Ka/Ks ratio of pairwise MdSPS genes indicated that the members of this family have undergone strong purifying selection during domestication. Furthermore, three SPS gene subfamilies were classified based on phylogenetic relationships, and old gene duplications and significantly divergent evolutionary rates were observed among the SPS gene subfamilies. In addition, a major gene related to sucrose accumulation (MdSPSA2.3) was identified according to the highly consistent trends in the changes of its expression in four apple varieties (‘Golden Delicious’, ‘Fuji’, ‘Qinguan’ and ‘Honeycrisp’) and the correlation between gene expression and soluble sugar content during fruit development. Furthermore, the virus-induced silencing of MdSPSA2.3 confirmed its function in sucrose accumulation in apple fruit. The present study lays a theoretical foundation for better clarifying the biological functions of the MdSPS genes during apple fruit development.
As there is a strong interest in red-skinned pears, the molecular mechanism of anthocyanin regulation in red-skinned pears has been widely investigated; however, little is known about the molecular mechanism of anthocyanin regulation in red-fleshed pears due to limited availability of such germplasm, primarily found in European pears (Pyrus communis). In this study, based on transcriptomic analysis in red-fleshed and white-fleshed pears, we identified an ethylene response factor (ERF) from P. communis, PcERF5, of which expression level in fruit flesh was significantly correlated with anthocyanin content. We then verified the function of PcERF5 in regulating anthocyanin accumulation by genetic transformation in both pear skin and apple calli. PcERF5 regulated anthocyanin biosynthesis by different regulatory pathways. On the one hand, PcERF5 can activate the transcription of flavonoid biosynthetic genes (PcDFR, PcANS and PcUFGT) and two key transcription factors encoding genes PcMYB10 and PcMYB114. On the other hand, PcERF5 interacted with PcMYB10 to form the ERF5-MYB10 protein complex that enhanced the transcriptional activation of PcERF5 on its target genes. Our results suggested that PcERF5 functioned as a transcriptional activator in regulating anthocyanin biosynthesis, which provides new insights into the regulatory mechanism of anthocyanin biosynthesis. This new knowledge will provide guidance for molecular breeding of red-fleshed pear.
PLATZ is a novel zinc finger DNA-binding protein that plays an important role in regulating plant growth and development and resisting abiotic stress. However, there has been very little research on the function of this family gene in tomatoes, which limits its application in germplasm resource improvement. Therefore, the PLATZ gene family was identified and analyzed in tomato, and its roles were predicted and verified to provide a basis for in-depth research on SlPLATZ gene function. In this study, the PLATZ family members of tomato were identified in the whole genome, and 19 SlPLATZ genes were obtained. Functional prediction was conducted based on gene and promoter structure analysis and RNA-seq-based expression pattern analysis. SlPLATZ genes that responded significantly under different abiotic stresses or were significantly differentially expressed among multiple tissues were screened as functional gene resources. SlPLATZ17 was selected for functional verification by experiment-based analysis. The results showed that the downregulation of SlPLATZ17 gene expression reduced the drought and salt tolerance of tomato plants. Tomato plants overexpressing SlPLATZ17 had larger flower sizes and long, thin petals, adjacent petals were not connected at the base, and the stamen circumference was smaller. This study contributes to understanding the functions of the SlPLATZ family in tomato and provides a reference for functional gene screening.
Under appropriate culture conditions, plant cells can regenerate new organs or even whole plants. De novo organ regeneration is an excellent biological system, which usually requires additional growth regulators, including auxin and cytokinin. Nitrate is an essential nutrient element for plant vegetative and reproductive development. It has been reported that nitrate is involved in auxin biosynthesis and transport throughout the growth and development of plants. In this study, we demonstrated that the ectopic expression of the MdNLP7 transcription factor in Arabidopsis could regulate the regeneration of root explants. MdNLP7 mainly participated in the regulation of callus formation, starting with pericycle cell division, and mainly affected auxin distribution and accumulation in the regulation process. Moreover, MdNLP7 upregulated the expression of genes related to auxin biosynthesis and transport in the callus formation stage. The results demonstrated that MdNLP7 may play a role in the nitrate-modulated regeneration of root explants. Moreover, the results revealed that nitrate–auxin crosstalk is required for de novo callus initiation and clarified the mechanisms of organogenesis.
A mutation in the promoter of the yellow stripe-like transporter gene in cucumber results in a yellow cotyledon phenotype
Alternative splicing of the PECTINESTERASE gene encoding a cell wall-degrading enzyme affects postharvest softening in grape
The firmness of table grape berries is a crucial quality parameter. Despite extensive research on postharvest fruit softening, its precise molecular mechanisms remain elusive. To enhance our comprehension of the underlying molecular factors, we initially identified differentially expressed genes (DEGs) by comparing the transcriptomes of folic acid (FA)-treated and water-treated (CK) berries at different time points. We then analyzed the sequences to detect alternatively spliced (AS) genes associated with postharvest softening. A total of 2,559 DEGs were identified and categorized into four subclusters based on their expression patterns, with subcluster-4 genes exhibiting higher expression in the CK group compared with the FA treatment group. There were 1,045 AS-associated genes specific to FA-treated berries and 1,042 in the CK-treated berries, respectively. Gene Ontology (GO) annotation indicated that the AS-associated genes in CK-treated berries were predominantly enriched in cell wall metabolic processes, particularly cell wall degradation processes. Through a comparison between treatment-associated AS genes and subcluster-4 DEGs, we identified eight genes, including Pectinesterase 2 (VvPE2, Vitvi15g00704), which encodes a cell wall-degrading enzyme and was predicted to undergo an A3SS event. The reverse transcription polymerase chain reaction further confirmed the presence of a truncated transcript variant of VvPE2 in the FA-treated berries. Our study provides a comprehensive analysis of AS events in postharvest grape berries using transcriptome sequencing and underscores the pivotal role of VvPE2 during the postharvest storage of grape berries.
Genome-wide identification of the CONSTANS-LIKE (COL) family and mechanism of fruit senescence regulation by PpCOL8 in sand pear (Pyrus pyrifolia)
Establishing VIGS and CRISPR/Cas9 techniques to verify RsPDS function in radish
Duplicated chalcone synthase (CHS) genes modulate flavonoid production in tea plants in response to light stress
EjGASA6 promotes flowering and root elongation by enhancing gibberellin biosynthesis
The Gibberellic Acid-stimulated Arabidopsis (GASA) gene family is involved in the regulation of gene expression and plant growth, development, and stress responses. To investigate the function of loquat GASA genes in the growth and developmental regulation of plants, a loquat EjGASA6 gene homologous to Arabidopsis AtGASA6 was cloned. EjGASA6 expression was induced by gibberellin, and ectopic transgenic plants containing this gene exhibited earlier bloom and longer primary roots since these phenotypic characteristics are related to higher gibberellin content. Transcriptome analysis and qRT-PCR results showed that the expression levels of GA3ox1 and GA20ox1, which encode key enzymes in gibberellin biosynthesis, were significantly increased. Furthermore, we confirmed that EjGASA6 could promote the expression of GA20ox1 via the luciferase reporter system. Overall, our results suggest that EjGASA6 promotes blooming and main-root elongation by positively regulating gibberellin biosynthesis. These findings broaden our understanding of the role of GASAs in plant development and growth, and lay the groundwork for future research into the functions of EjGASA6 in regulating loquat growth and development.
Seed size is an important agronomic trait in melons that directly affects seed germination and subsequent seedling growth. However, the genetic mechanism underlying seed size in melon remains unclear. In the present study, we employed Bulked-Segregant Analysis sequencing (BSA-seq) to identify a candidate region (~1.35 Mb) on chromosome 6 that corresponds to seed size. This interval was confirmed by QTL mapping of three seed size-related traits from an F2 population across three environments. This mapping region represented nine QTLs that shared an overlapping region on chromosome 6, collectively referred to as qSS6.1. New InDel markers were developed in the qSS6.1 region, narrowing it down to a 68.35 kb interval that contains eight annotated genes. Sequence variation analysis of the eight genes identified a SNP with a C to T transition mutation in the promoter region of MELO3C014002, a leucine-rich repeat receptor-like kinase (LRR-RLK) gene. This mutation affected the promoter activity of the MELO3C014002 gene and was successfully used to differentiate the large-seeded accessions (C-allele) from the small-seeded accessions (T-allele). qRT-PCR revealed differential expression of MELO3C014002 between the two parental lines. Its predicted protein has typical LRR-RLK family domains, and phylogenetic analyses reveled its similarity with the homologs in several plant species. Altogether, these findings suggest MELO3C014002 as the most likely candidate gene involved in melon seed size regulation. Our results will be helpful for better understanding the genetic mechanism regulating seed size in melons and for genetically improving this important trait through molecular breeding pathways.