To identify potential protein markers associated with tillering in rice, a d10 allelic mutant (JHCA) with a high-tillering dwarf phenotype and a wild-type cultivar (GLA4) with normal numbers of tillers were used to produce a hybrid (F 1 ), and 2-DE (two-dimensional electrophoresis) analysis was conducted in seedling leaf sheaths and roots of the rice hybrid and its parents. A total of 61 protein spots representing qualitative or quantitative polymorphisms were detected. Among them, 38 spots were successfully identified as 23 different proteins by MALDI-TOF/MS. The identified proteins were further divided into P/As (presence/absence variations), PSs (position shift variations) and qVs (quantitative variations). Most of the identified polymorphic proteins exhibited tissue-specific expression. Two spot families, identified as a putative vitamin B12-independent methionine synthase and a putative phosphoglycerate mutase were found. The profiles of expression abundance of three iso-spots in each spot family showed a reciprocal relationship between the two parents.
Background
The normal metabolism of transitory starch in leaves plays an important role in ensuring photosynthesis, delaying senescence and maintaining high yield in crops. OsCKI1 (casein kinase I1) plays crucial regulatory roles in multiple important physiological processes, including root development, hormonal signaling and low temperature-treatment adaptive growth in rice; however, its potential role in regulating temporary starch metabolism or premature leaf senescence remains unclear. To reveal the molecular regulatory mechanism of OsCKI1 in rice leaves, physiological, transcriptomic and proteomic analyses of leaves of osckI1 allele mutant lses1 (leaf starch excess and senescence 1) and its wild-type varieties (WT) were performed.
Results
Phenotypic identification and physiological measurements showed that the lses1 mutant exhibited starch excess in the leaves and an obvious leaf tip withering phenotype as well as high ROS and MDA contents, low chlorophyll content and protective enzyme activities compared to WT. The correlation analyses between protein and mRNA abundance are weak or limited. However, the changes of several important genes related to carbohydrate metabolism and apoptosis at the mRNA and protein levels were consistent. The protein-protein interaction (PPI) network might play accessory roles in promoting premature senescence of lses1 leaves. Comprehensive transcriptomic and proteomic analysis indicated that multiple key genes/proteins related to starch and sugar metabolism, apoptosis and ABA signaling exhibited significant differential expression. Abnormal increase in temporary starch was highly correlated with the expression of starch biosynthesis-related genes, which might be the main factor that causes premature leaf senescence and changes in multiple metabolic levels in leaves of lses1. In addition, four proteins associated with ABA accumulation and signaling, and three CKI potential target proteins related to starch biosynthesis were up-regulated in the lses1 mutant, suggesting that LSES1 may affect temporary starch accumulation and premature leaf senescence through phosphorylation crosstalk ABA signaling and starch anabolic pathways.
Conclusion
The current study established the high correlation between the changes in physiological characteristics and mRNA and protein expression profiles in lses1 leaves, and emphasized the positive effect of excessive starch on accelerating premature leaf senescence. The expression patterns of genes/proteins related to starch biosynthesis and ABA signaling were analyzed via transcriptomes and proteomes, which provided a novel direction and research basis for the subsequent exploration of the regulation mechanism of temporary starch and apoptosis via LSES1/OsCKI1 in rice.
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