Functional Analysis Of Two RNA Recognition Motifs Encoded By RFCA In Rice (Orysa Sativa L.)

In eukaryotic cells, regulation of gene expression at the post-transcriptional level is mainly achieved by proteins, which containing well defined sequence motifs involved in RNA binding. Most RNA-binding proteins contain one or several conserved domains, such as the RNA recognition motif (RRM), the K-homology (KH) motif, RGG (Arg-Gly-Gly) boxes, and double-stranded RNA-binding domains (dsRBDs)RNA recognition motifs as important regulators of gene expression are highly conserved in animals and plants. The FCA floral promotion gene in Arabidopsis encodes a protein, containing two RNA recognition motifs (RRM) and a WW protein interaction domain. Here we isolated rFCA cDNA from rice. FCA in rice (rFCA) was homologous to FCA-gamma of Arabidopsis and contained conserved two RRM domains. To investigate the function of RRM domain, fragment RRM1 and RRM2 of rFCA were introduced into rice subspecies Oryza sativa L. subsp. Indica var. 9311 and another rice subspecies Oryza sativa L. subsp. Japonica var. zhonghua11 transformation. We examined whether RNA recognition motif domains, rFCA-RRM1 and rFCA-RRM2 would cause any similar effect on the development of the transgenic plants by regulating the rice gene expression. In transgenic lines, pCAM-RRM1 and pBY-RRM2, the vegetative stage and growth of transgenic rice plants were retarded about two weeks compared with the wild type rice. Transgenic plants showed relatively dark green and thicker leaves. It was indicated that the two transgenic lines plants had larger seeds and pollen. Two transgenic lines exhibited similar phenotypes, flowering time delay, seed size and cell volume of transgenic plants was increased. These results showed that constitutive overexpression of RRM domains could regulate transgenic rice cellular size. The patterns of overexpression of two RRM domains and their similar morphologies indicate they may play a similar role. To test the function of RRM domain in increasing the transgenic rice seed mass, RRM1: GFP was introduced into another rice subspecies Nanjing Dalidao, and the similar phenotypes were obtained. We also found that the weight of seed increased in the transgenic rice. Thus, the overexpression of RRM1 may have involved into regulate seed development.These results indicated that the same phenotypic characteristics of late flowering and increased rice seed mass, were associated with the overexpression of the rFCA RRMs.