The Molecular Basis Of Research On The Rice Lodicules Adjustment To The Floret Opening Mechanism

The rice floret opening is started by the expanding lodicule. This process is connected with respiration and regulated by many factors. In this paper, the materials were sampled from different stage lodicules in Indica riceⅡ-32B, we use the methods of Proteomic Techniques,electron microscope cytochemical method and phosphorylation Western blot analysis technology to study the change of proteome in lodicules, spatial and temporal distribution character of Ca²⁺ and the alter of situation of protein phosphorylation before and after flowered, aiming to explore the mechanism of how the rice lodicules regulared opening of the glumous flower. The results are as follows:(1) Study on proteomics of different stages of rice lodicules in the process of floret opening.After colloidal coomassie staining,the 2-DE image analysis by PDQuest software detected 192 protein spots in the lodicule one day before flowering,198 spots in the lodicule nearly anthesis,and 212 spots in the lodicule at anthesis. To contrast with the lodicule at anthesis results, the lodicule one day before flowering matches 159 protein spots and the lodicule flowering matches 182 protein spots.The MALDI-TOF-TOF MS Identification resulte of 13 protein spots with significant and stable difference is: VacuolarATP synthase catalytic subunit A, vacuolar acid invertase, aldolase C-1, sucrose synthase, triosephosphate isomerase, glyceraldehyde-3-phosphate isomerase, glyceraldehyde-3-phosphate,β-D-xylosidase, Tubulin beta-1 chain, Tubulin beta-3 chain, RGP1 protein, EF1B.Nearly anthesis, aldolase C-1, triosephosphate isomerase, glyceraldehyde-3- phosphate which are 3 key enzyme relating to glycolysis are expression higher level. the result showes that glycolysis is activity in the cell of lodicule nearly anthesis; vacuolar acid invertase is appear nearly anthesis and lodicule at anthesis. It catalyzes the hydrolysis of sucrose into glucose and fructose. glucose, fructose and intermediary metabolite which are from glycolysis resulted in high osmotic potential and water potential. The higher expression of VacuolarATP synthase catalytic subunit A were beneficial to obtain the optimal acidic pH for vesicle trafficking, hydrolase activating, and membrane transporting, and it may be also favorable for accumulating heavy metal ions into vacuoled. The activity ofβ-D-xylosidase increased nearly anthesis which were beneficial for the degradation of plant cell walls. The expressing of Tubulin was lowered nearly anthesis that showed Tubulin had on correlation with cell expansion.(2) Ca²⁺ location in the lodicule cell before and after rice floret anthesisThe result of potassium pyroantimonate precipitation analysis showed the space-time dynamic difference of Ca²⁺ deposit before and after rice floret anthesis: one day before anthesis, Ca²⁺ largely accumulated in vessel and intercellular space; nearly anthesis, most part appeared in the endovascular and on the thin membrane, small part appeared in the plasma membrane, mitochondria, endoplasmic reticulum; the time when after flowering to when the field angle getting to maximum, Ca²⁺ moved towards to intra-cellular obviously, there adhered so much Ca²⁺ deposit in endoplasmic reticulum, mitochondria, vacuolar membrane. Along with the close of floret and the shrink of the lodicule, numerous endoplasmic reticulum that took with Ca²⁺ deposit moved to mitochondria and other cellular organ, then conformed degradation of vesicles, the mitochondria which had more Ca²⁺ deposit have degraded into vesicle; three hours after anthesis, the content of lodicule cell was evacuated, and the cell crimpled, but multimembrane corpuscle that had Ca²⁺ still kept in. This result showed that there existed correlation relationship between the expanding and the shrink of lodicule and space-time dynamic change of Ca²⁺ in lodicule.(3) western blot analysis of tyrosine phosphorylation before and after rice floret anthesisThe method of tyrosine phosphorylation polyclone antibody was utilized to carry out western blot to analyze protein tyrosine phosphorylation situation before and after rice floret anthesis. The results showed that the protein of that had molecular weight at 44.2, 27.1. 26.1, 22.6, 20.9, 19.9, 18.7 and 17.0 KD were phosphorylated, when nearly anthesis, the protein of 27.1, 26.1, 18.7 KD occurred certain degree of dephosphorylation, when after anthesis, the protein of 17.0 KD also occurred certain degree of dephosphorylation, this meant that tyrosine phosphorylation existed in lodicule. In the process of rice floret anthesis, reversible phosphorylation of protein tyrosine participated the turgormovement of lodicule.