Characterization And Gene Cloning Of White Green Leaf 2(WGL2) In Rice

Plastid ribosomal proteins(PRPs)play important roles in the translation of key proteins involved in chloroplast development and photosynthesis.PRPs have been extensively studied in many plants,but the related molecular mechanisms are still unclear due to the numerous homologous genes of PRP existed and the wide range of fields involved in plants.In the present study,we used ethyl methane sulfonate mutagenesis and obtained a novel rice mutant called white green leaf 2(wgl2).The wgl2 mutants exhibited an albino phenotype from germination through the three-leaf stage,and then gradually transitioned to green through the later developmental stages.Consistent with this albino phenotype,wgl2 mutants had lower Chl a,Chl b,and Car contents relative to the wild type(NPB).Moreover,the total chlorophyll content of wgl2 and the ratio of Chl a/b were much lower than in the wild type.The wgl2 albino mutant had chloroplasts with disrupted architecture,as well as no stromal thylakoids or stacked grana thylakoids,instead displaying oval-shaped vesicles.Genetic analysis showed that wgl2 was controlled by a single recessive nuclear gene,Map-based cloning and DNA sequencing analyses of wgl2 revealed a single-nucleotide substitution(G to T)in the first exon of LOC_Os03g55930,which resulted in a substitution of glycine 92 to valine,but it did not change the three-dimensional structure of the protein.Complementation and targeted deletion experiments confirmed that the point mutation in WGL2 was responsible for the wgl2 mutant phenotype.BLAST-P analysis of the NCBI database showed that the WGL2 C-terminal belongs to the ribosomal S9 super family,and was highly conserved in higher plants.The phylogenetic analysis of WGL2 homologs showed that WGL2 homologues could be clearly divided into two groups: monocots and dicots.The subcellular localization of WGL2 showed that WGL2 was mainly located in chloroplast and performed biological function in chloroplast.Through qRT-PCR,we found WGL2 was ubiquitously expressed in all tissues examined,which was the highest in flag leaf,the second in leaf sheath,the lowest in other parts such as young root,uppermost internode and young panicle.The mutation of WGL2 led to significant changes in the expression of genes related to chlorophyll biosynthesis,photosynthesis,chloroplast development,and ribosome development compared with WT.In conclusion,our study had proved through a series of experiments that WGL2 was essential for early chloroplast development in rice.Further molecular study of wgl2 mutant will reveal its function in early plant development.