Global landscape of protein phosphorylation during plant regeneration initiation in cotton (Gossypium hirsutum L.)
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Q Exactive HF
ORGANISM(S): Gossypium Hirsutum
TISSUE(S): Hypocotyl
SUBMITTER: 
Huihui Guo
LAB HEAD: Huihui Guo
PROVIDER: PXD061812 | Pride | 2025-03-18
REPOSITORIES: Pride
Dataset's files
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| S8148TPST_1.raw | Raw | |||
| S8148TPST_2.raw | Raw | |||
| S8148TPST_3.raw | Raw | |||
| S8148TPST_4.raw | Raw | |||
| S8148TPST_5.raw | Raw |
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Project description:Somatic embryogenesis (SE) is a biotechnological tool used to generate new individuals and is the preferred method for rapid plant regeneration. However, the molecular basis underlying somatic cell regeneration through SE is not yet fully understood, particularly regarding interactions between the proteome and post-translational modifications. Here, we performed association analysis of high-throughput proteomics and phosphoproteomics in three representative samples (non-embryogenic calli, NEC; primary embryogenic calli, PEC; globular embryos, GE) during the initiation of plant regeneration in cotton, a pioneer crop for genetic biotechnology applications. Our results showed that protein accumulation is positively regulated by phosphorylation during SE, as revealed by correlation analyses. Of the 1418 proteins that were differentially accumulated in the proteome and the 1106 phosphoproteins that were differentially regulated in the phosphoproteome, 115 proteins with 229 phosphorylation sites overlapped (co-differential). Furthermore, seven dynamic trajectory patterns of differentially accumulated proteins (DAPs) and the correlated differentially regulated phosphoproteins (DRPPs) pairs with enrichment features were observed. During the initiation of plant regeneration, functional enrichment analysis revealed that the overlapping proteins (DAPs-DRPPs) were considerably enriched in cellular nitrogen metabolism, spliceosome formation, and reproductive structure development. Moreover, 198 DRPPs (387 phosphorylation sites) were specifically regulated at the phosphorylation level and showed four patterns of stage-enriched phosphorylation susceptibility. Furthermore, enrichment annotation analysis revealed that these phosphoproteins were significantly enriched in endosomal transport and nucleus organization processes. During embryogenic differentiation, we identified five DAPs-DRPPs with significantly enriched characteristic patterns. These proteins may play essential roles in transcriptional regulation and signaling events that initiate plant regeneration through protein accumulation and/or phosphorylation modification. This study enriched the understanding of key proteins and their correlated phosphorylation patterns during plant regeneration, and also provided a reference for improving plant regeneration efficiency.
2025-03-21 | PXD062095 | Pride
Project description:In somatic embryogenesis, embryos are formed from a single or small group of somatic cells in response to stress and hormonal stimuli. In this study, a high-throughput phosphoproteomic analysis was used to identify and quantify phosphoproteins related to the acquisition of embryogenic competence in sugarcane. Embryogenic (EC) and nonembryogenic (NEC) calli were compared at the multiplication phase, resulting in the identification of 1279 phosphoproteins. After the differential accumulation analysis, 163 phosphoproteins were identified as unique to EC, and nine were unique to NEC, while 51 were upaccumulated, and 40 were downaccumulated in EC compared to NEC. These phosphopeptides were mainly single phosphorylated peptides, with serine residues representing the major site of phosphorylation. The motif-x analysis revealed the enrichment of [xxxpSPxxx], [RxxpSxxx] and [xxxpSDxxx] motifs, which are predicted phosphorylation sites for several kinases related to stress responses. Gene ontology enrichment revealed an overrepresentation of biological processes involved in embryonic development, showing that EC-related phosphoproteins (unique to EC and upaccumulated) were associated with stress responses, regulation of gene expression and epigenetic modifications. The NEC-related phosphoproteins (unique to NEC and downaccumulated) were associated with the translation process. In this sense, EC-related phosphoproteins described as potential regulators of abiotic stress tolerance in response to ABA-induced signals such as OSK3 (a catalytic subunit of SnRK1), ABF1, LEAs, REM4.1 and RD29Bs were identified. On the other hand, the NEC-related phosphoproteins EDR1 and PP2Ac-2 were negative regulators of ABA, suggesting a role of ABA in the acquisition of embryogenic competence linked to stress tolerance. Moreover, EC-related phosphoproteins associated with epigenetic modifications, such as HDA6, HDA19, and TPL, as well as those involved in embryo development, including ASIL1, M3KE1, MCM2, and GRV2, were identified as putative potential regulators of embryogenic competence. Our results provide novel data that may help elucidate the phosphorylation mechanisms controlling the activity, affinity and localization of specific proteins during somatic embryogenesis and suggest a role for these proteins in the hormonal signalling cascade leading to stress adaptation by reprogramming genetic expression.
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Project description:To identify transcription factor (TF) genes exhibiting preferential expression during SE, expression profiles of 1,880 TFs were compared in two genotypes with largely different capacities for SE induction in IZE culture on auxin medium, namely the highly embryonic Col-0 accession and the tanmei mutant unable to form somatic embryos. Our study revealed 729 TFs whose expression changes during the 10-day incubation period of SE; 141 TFs displayed distinct differences in expression patterns in embryogenic versus non-embryogenic cultures. This study provides comprehensive data focused on the expression of TF genes during SE and provides guidelines for further research on functional genomics of SE. The experiment was designed to monitor the expression of 1,880 TF genes at three distinctive stages of IZE-derived embryogenic culture, which refer to: (i) freshly isolated explants competent to undergo embryogenic transition (0 d), (ii) explant tissue subjected to SE induction (5 d), and (iii) the advanced phase of embryogenesis related to somatic embryo formation (10 d).
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Project description:To identify transcription factor (TF) genes exhibiting preferential expression during SE, expression profiles of 1,880 TFs were compared in two genotypes with largely different capacities for SE induction in IZE culture on auxin medium, namely the highly embryonic Col-0 accession and the tanmei mutant unable to form somatic embryos. Our study revealed 729 TFs whose expression changes during the 10-day incubation period of SE; 141 TFs displayed distinct differences in expression patterns in embryogenic versus non-embryogenic cultures. This study provides comprehensive data focused on the expression of TF genes during SE and provides guidelines for further research on functional genomics of SE.
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Project description:Somatic embryogenesis (SE), a morphogenic process that takes advantage of the regenerative potential of plants to replicate whole plants starting from somatic explants, can be a source of variation with potential applications in plant breeding. It is one of the most suitable tools to apply functional genomics studies and genetic improvement in plants. However, beyond a few pioneering works mainly focused on model plants, the molecular characterization of SE mechanisms is still elusive, especially for woody species. In grapevine, this process is affected by many factors such as explant type, culture conditions and, most importantly, genotype. Many cultivars, in fact, have shown recalcitrance to tissue culture and transformation, and very low SE competence. Thus, the understanding of SE competence behind the regenerative aptitude is fundamental to the widespread application of the so-called “next-generation breeding techniques”, such as cisgenesis and genome editing, in grapevine. Here, we explored genetic and epigenetic features of the SE process in grapevine by investigating the behavior of two genotypes showing opposite SE competence. Embryogenic tissues were induced from immature stamens excised from field-collected flower clusters of Sangiovese (highly competent for embryogenesis) and Cabernet Sauvignon (poorly competent for embryogenesis). A multilayered approach was used to profile mRNA, smallRNAs and methylated DNA with high-throughput sequencing technologies in the initial explants, on undifferentiated calli induced after 40 days of culture, and in embryogenic and non-embryogenic calli after 3 months of culture. A comprehensive comparison of transcriptomes of the different types of calli with the grapevine gene expression atlas revealed that, in grapevine, the dedifferentiation to the callus formation during the embryogenesis process occurs via a ‘berry’ developmental pathway. This is dissimilar from that shown in Arabidopsis, in which dedifferentiated calli are more similar to the tip of a root meristem. Interestingly, a Gene Ontology (GO) analysis revealed that secondary metabolism and gene expression regulation/epigenetics are the enriched functional categories of genes differentially down- and up-regulated in embryogenic vs non-embryogenic calli, respectively. These results prompted us to define the epigenetic landscape dynamics during SE in grapevine, revealing a significant increase in DNA methylation, especially in intergenic regions, in the embryogenic calli tissues. Finally, we proposed potential key regulators of SE in different genotypes that could represent putative targets of next-generation breeding techniques in grapevine.
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Project description:Somatic embryogenesis (SE), a morphogenic process that takes advantage of the regenerative potential of plants to replicate whole plants starting from somatic explants, can be a source of variation with potential applications in plant breeding. It is one of the most suitable tools to apply functional genomics studies and genetic improvement in plants. However, beyond a few pioneering works mainly focused on model plants, the molecular characterization of SE mechanisms is still elusive, especially for woody species. In grapevine, this process is affected by many factors such as explant type, culture conditions and, most importantly, genotype. Many cultivars, in fact, have shown recalcitrance to tissue culture and transformation, and very low SE competence. Thus, the understanding of SE competence behind the regenerative aptitude is fundamental to the widespread application of the so-called “next-generation breeding techniques”, such as cisgenesis and genome editing, in grapevine. Here, we explored genetic and epigenetic features of the SE process in grapevine by investigating the behavior of two genotypes showing opposite SE competence. Embryogenic tissues were induced from immature stamens excised from field-collected flower clusters of Sangiovese (highly competent for embryogenesis) and Cabernet Sauvignon (poorly competent for embryogenesis). A multilayered approach was used to profile mRNA, smallRNAs and methylated DNA with high-throughput sequencing technologies in the initial explants, on undifferentiated calli induced after 40 days of culture, and in embryogenic and non-embryogenic calli after 3 months of culture. A comprehensive comparison of transcriptomes of the different types of calli with the grapevine gene expression atlas revealed that, in grapevine, the dedifferentiation to the callus formation during the embryogenesis process occurs via a ‘berry’ developmental pathway. This is dissimilar from that shown in Arabidopsis, in which dedifferentiated calli are more similar to the tip of a root meristem. Interestingly, a Gene Ontology (GO) analysis revealed that secondary metabolism and gene expression regulation/epigenetics are the enriched functional categories of genes differentially down- and up-regulated in embryogenic vs non-embryogenic calli, respectively. These results prompted us to define the epigenetic landscape dynamics during SE in grapevine, revealing a significant increase in DNA methylation, especially in intergenic regions, in the embryogenic calli tissues. Finally, we proposed potential key regulators of SE in different genotypes that could represent putative targets of next-generation breeding techniques in grapevine.
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Project description:Somatic embryogenesis (SE) is a complex stress related process regulated by numerous biological factors. SE is mainly applicable to mass propagation and genetic improvement of plants through gene transfer technology and mutation breeding. In banana, SE is highly genome dependent as the efficiency varies with cultivars. To understand molecular mechanism of SE, proteomics approach was carried out to identify genes responsible for embryogenic calli (EC) induction, regeneration and germination of somatic embryos (se) in cv. Rasthali (AAB). In total, 70 spots were differentially expressed in various developmental stages of SE. Of which, 16 were uniquely expressed and 17 were highly abundant in EC than nonembryogenic calli and explant and four spots were also uniquely expressed in germinating se. Functional annotation of identified proteins revealed that calcium signaling along with stress and endogenous hormones related proteins played a vital role in EC induction and germination of se. Thus based on the outcome, callus induction media was modified and tested in five cultivars. In cv. Grand Naine (AAA), increased concentration of 3- IAA and tryptophan recorded highest EC induction of 24.28% while Red Banana with similar genome showed 18.96% in kinetin supplemented media. Similarly, in cultivars Monthan and Karpuravalli with ABB genome showed maximum EC induction in tryptophan supplemented media (8.54%) and CaCl2 enriched media (17.34%) respectively. In cv. Neypoovan (AB), higher concentration of tryptophan induced more EC. These results illustrated that EC formation is genome as well as cultivar dependent. Simultaneously, germination media was modified to induce proteins responsible for germination. In cv. Rasthali, media supplemented with 10 mM CaCl2 showed maximum increase in germination (51.79%) over control. Thus present study revealed that media modification based on proteomic studies can induce SE in recalcitrant cultivars and also enhance germination in cultivars amenable for SE.
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