Functional Analysis Of OsLPL2 And Identification Of Topp4-1 Revertant Mutant In Arabidopsis

Actin cytoskeleton plays a critical role in plant epidermal cells during morphogenesis.Abnormal actin will result in a series of deficient phenotype.In Arabidopsis,current researches of actin nucleation are focusing on ROP-SCAR/WAVE-ARP2/3 signaling pathway.The SCAR/WAVE complex(PIR/NAP125/HSPC300/ABI/SCAR)is a effector that convert small GTPase signals into defined ARP2/3 activation responses.In rice,the regulatory mechanism of actin is still unknown.In previous study,lpl2-1(less pronounced lobe epidermal cell 2-1),a mutant which is controlled by the recessive heredity of single gene of autosomes,have been identified.lpl2-1 shows epidermal pavement cells with smooth marginal lobes.The excalation of Os03g05020(OsLPL2)leads to premature transcription termination.We run BLAST and find that OsLPL2 encodes a putative protein homologous to AtPIR.OsLPL2 shares high degree of amino acid similarity with its homologous proteins in higher plants which indicates that it may play a conservative role in evolution.Since AtPIR can directly interact with AtNAP125,we then order a rice mutant lpl3-1,whose mutant gene(OsLPL3)homologues to AtNAP125.lpl3-1 exhibits a phenotype similar to lpl2-1.To further investigate the possible function of OsLPL2 in actin regulation,yeast two-hybrid has been used to explore whether a similar mechanism of actin exists in rice.The results show that OsLPL2 can directly interact with OsLPL3 and the PH domain of OsLPL2 in the N-terminal,which plays a role in interaction.In contrast,neither OsLPL2 nor OsLPL3 can interact with OsLPL1(homologues to AtHSPC300).We also demonstrate that OsLPL2 fails to interact with proteins(such as OsRAC5,OsRAC6 and OsRAC2)homologues to AtROP2.Together with previous results,we prove that regulatory mechanism of actin exists in rice which similar to Arabidopsis.In addition,we identify a series of revertant mutants by screening an activation tagging mutagenized topp4-1 population.In this study,we characterize the phenotypes of revertant mutant 5067-8#.It can rescue phenotype defects of topp4-1 in height,leaf shape,silique,pavement cell and so on.The T-DNA insertion site is located on chromosome 1.Relative expression of candidate gene has been detected to determine the true gene which can function in 5067-8#.This study provides basic materials for the important role of type-one phosphatase(TOPP4)during plant growth and development.