RBS1, an RNA Binding Protein, Interacts with SPIN1 and Is Involved in Flowering Time Control in Rice

The rice U-box/ARM E3 ubiquitin ligase SPL11 negatively regulates programmed cell death (PCD) and disease resistance, and controls flowering time through interacting with the novel RNA/DNA binding KH domain protein SPIN1. Overexpression of Spin1 causes late flowering in transgenic rice under short-day (SD) and long-day (LD) conditions. In this study, we characterized the function of the RNA-binding and SPIN1-interacting 1 (RBS1) protein in flowering time regulation. Rbs1was identified in a yeast-two-hybrid screen using the full-length Spin1 cDNA as a bait and encodes an RNA binding protein with three RNA recognition motifs. The protein binds RNA in vitro and interacts with SPIN1 in the nucleus. Rbs1 overexpression causes delayed flowering under SD and LD conditions in rice. Expression analyses of flowering marker genes show that Rbs1 overexpression represses the expression of Hd3a under SD and LD conditions. Rbs1 is upregulated in both Spin1 overexpression plants and in the spl11 mutant. Interestingly, Spin1 expression is increased but Spl11 expression is repressed in the Rbs1 overexpression plants. Western blot analysis revealed that the SPIN1 protein level is increased in the Rbs1 overexpression plants and that the RBS1 protein level is also up-regulated in the Spin1 overexpression plants. These results suggest that RBS1 is a new negative regulator of flowering time that itself is positively regulated by SPIN1 but negativel