Validation And Analysis Of Differentially Accumulated Fertility-related Protein In Photoperiod Induced Male Sterile Rice

Fertility regulation mechanism of Photoperiod/Thermo-sensitive genic male sterile lines was the core content of two-line hybrid rice research foundation. Our previous work has received differentially expressed proteins about flavonoid metabolism, carbohydrate synthesis and lipid synthesis, which were found through by proteomic analysis in photoperiod-sensitive genic male sterile rice nongken 58S(NK58S) and short photoperiod-sensitive genic male sterile rice D52 S under long-day length and short-day length.In order to better understand the relationship between these differences proteins and fertility transition and provide a basis for further analytical mechanism of fertility transition induction by photoperiodic, this study has received a significant results of induction fertility conversion, which were found by handling the NK58 S and D52 S make use of 14 hours long-day length(LD) and 10 hours short-day length(SD) at the same temperature conditions. We did validation under the transcriptional level for differentially expressed proteins, such as, 4-coumarate: coenzyme A ligase 9(4CLL9), chalcone synthase(CHS), caffeic acid-O-methyl transferase(COMT), UDP-glucose pyrophosphorylase(UGPase), sucrose synthase 1(SUS1), glucose-1-phosphate acyltransferase adenosine(AGPase), granule-bound starch synthase II(GBSSII). We also did validation under the protein level for these differentially expressed proteins. At the same time, we measured the content of flavonoids, soluble sugar, starch, soluble protein, proline, malondialdehyde(MDA) and the activity of antioxidant enzymes. We hope that these results will contribute to a more comprehensive understanding of the mechanisms of fertility regulations in photoperiod-sensitive genic male sterile rice and short photoperiod-sensitive genic male sterile rice. The main obtained results are as follows:1. At the transcriptional level: Flavonoid biosynthesis-related genes 4CLL9, CHS and COMT relative expression levels were gradually increased with the advance of panicle development process under different photoperiodic conditions in D52 S and NK58 S, and observations from m RNA levels of 4CLL9, CHS and COMT in our study showed that their expression patterns were different between sterile and fertile plants both in D52 S and NK58 S at stage 7, suggesting that flavonoid biosynthesis are intimately associated with fertility transformation. Carbohydrate metabolism- related genes UGPase, SUS1, AGPase and GBSSII relative expression levels were not regular pattern with fertility conversion under different photoperiodic conditions in D52 S and NK58 S.2. On the protein level: The expression patterns of CHS and 4CLL9 were distinctive under different day-lengths in D52 S and NK58 S, the results revealed that originally expression patterns and background buildup of CHS and 4CLL9 were difference in different varieties, but these enzymes exhibited the same change at the transcriptional level during the plant fertility conversion.3. Physiological indexes: The content of flavonoids were decreased with panicle development process in D52 S and NK58 S, the results revealed that the content of flavonoids under short-day conditions were significantly higher than those levels under long-day conditions at stage 5, the content of flavonoids in sterile panicles were significantly higher than those levels in fertile panicles at stage 6, the content of flavonoids in fertile panicles were significantly higher than those levels in sterile panicles at stage 7. Presumably infloresence flavonoid metabolism was affected by photoperiod. Flavonoids may remove oxygen free radicals with antioxidant enzyme system, thereby reducing peroxidation in young spike of fertile plants and finally produce normal pollen.The content of infloresence soluble sugar and starch were respectively increased and decreased with the advance of panicle development process in D52 S and NK58 S, and the content of soluble sugar and starch under short-day conditions were significantly higher than those levels under long-day conditions. The content of soluble protein and proline were the same change trend in D52 S and NK58 S, but the content of soluble protein in fertile panicles were significantly higher than those levels in sterile panicles at stage 5, and the content of proline were not significantly different between fertile plants and sterile plants. Our study provide the basic data for related gene expression, protease leve and dynamic changes of end products in the course of male sterility by photoperiodic induction, however, we need to do further research about the relationship between these results and metabolic regulation of fertility induction or fertility expression.