Genomic Identification Of PLL And PG Gene Families And Functional Analysis In Rice

Pectin is a type of complex polysaccharide widely present in the cell walls of higher plants.The pectin network is systematically disassembled along with the plant cells undergoing dramatic changes in shape,so pectin degradation is critical to many developmental processes in plants.Due to the complex composition and structure of pectin,various enzymes are required to participate in its degradation process,including polygalacturonases(PG),pectate lyases,and pectin methylesterases.Pectate lyases catalyze the cleavage of de-esterified pectin by(3 elimination.The PLL(pectate lyase-like)genes are widely present in various plant tissues,including pollen,mature fruit,pistil,tubular molecules,latex,and fiber.Polygalacturonases(PGs)catalyze the random hydrolysis and disassembly of the 1,4-a-D-galactosiduronic linkages of pectin.Functions of PLL genes and PG genes have been reported in many plants.They are mainly involved in fruit maturation,pollen development and pollen tube elongation,cell differentiation and isolation.However,functional studies of them in rice are still limited.In this study,a whole genome-wide identification,detailed bioinformatics and phylogenetic analysis of PLL and PG genes in rice were performed.In addition,artificial miRNA technology was adopted to silence two PLL genes,OsPLL3 and OsPLL4 for further functional elucidation.The main findings of this study are as follows:1.12 PLL genes were identified in rice genome.PLL proteins from rice and several plant species were used for phylogenetic analysis,results showed that PLL genes were conserved in terrestrial plants and can be divided into five branches.The results of tissue expression analysis showed that eight PLL genes could not be detected,suggesting that this gene family may have undergone pseudogenizations during evolution.Moreover,four genes,inculding OsPLL1,OsPLL3,OsPLL4,and OsPLL12 were abundantly expressed during seed germination and panicle development.In addition,OsPLL3 and OsPLL4 showed progressively higher expression patterns during young panicle development.2.In this study,artificial miRNAs expression vectors were constructed to silence OsPLL3 and OsPLL4 for functional elucidation.Agrobacterium tumefaciens EHA105 was used as a mediator to infect rice calli of Nipponbare.After screening,transgenic positive plants were successfully obtained.Results showed that the expression of OsPLL3 was reduced by more than 90%in RNAi-OsPLL3 lines,whereas OsPLL4 expression was between 0.33 and 0.48 in RNAi-OsPLL4 lines.I2-KI staining indicated that some pollen grains in the transgenic lines showed an irregular and atrophic shape,and cross section of anther showed that some of the pollen grains in the transgenic plants were degraded during the 13th stage of anther development.These results indicated that the silence of OsPLL3 and OsPLL4 affected the process of pollen maturation and resulted in partial sterility.3.The PG gene family has 44 members in rice,19 and 10 PG genes are related to tandem repeats and partial repeats,respectively,indicating that both of them are involved in the expansion of this gene family.By colinear analysis,three colinear gene pairs in PG genes were found between rice and Arabidopsis,indicating that they may have shared common ancestors before their divergences between monocotyledons and dicotyledons.The phylogenetic tree constructed in this study was divided into seven branches.Members in the same branch had relatively conserved gene structures and motifs.There are a large number of cis-acting elements that respond to plant hormones and stress in the promoter region of the PG genes.Based on the microarray data analysis,it was found that PG genes in branch E were constitutively expressed,and some genes were specifically expressed in young panicles.