A Bunyavirus-Inducible Ubiquitin Ligase Targets RNA Polymerase IV for Degradation during Viral Pathogenesis in Rice.

The ubiquitin-proteasome system (UPS) is an important post-translational regulatory mechanism that controls many cellular functions in eukaryotes. Here, we show that stable expression of P3 protein encoded by Rice grassy stunt virus (RGSV), a negative-strand RNA virus in the Bunyavirales, causes developmental abnormities similar to the disease symptoms caused by RGSV, such as dwarfing and excess tillering, in transgenic rice plants. We found that both transgenic expression of P3 and RGSV infection induce ubiquitination and UPS-dependent degradation of rice NUCLEAR RNA POLYMERASE D1a (OsNRPD1a), one of two orthologs of the largest subunit of plant-specific RNA polymerase IV (Pol IV), which is required for RNA-directed DNA methylation (RdDM). Furthermore, we identified a P3-inducible U-box type E3 ubiquitin ligase, designated as P3-inducible protein 1 (P3IP1), which interacts with OsNRPD1a and mediates its ubiquitination and UPS-dependent degradation in vitro and in vivo. Notably, both knockdown of OsNRPD1 and overexpression of P3IP1 in rice plants induced developmental phenotypes similar to RGSV disease symptomss. Taken together, our findings reveal a novel virulence mechanism whereby plant pathogens target host RNA Pol IV for UPS-dependent degradation to induce disease symptoms. Our study also identified an E3 ubiquitin ligase, which targets the RdDM compotent NRPD1 for UPS-mediated degradation in rice.

The complete chloroplast genome sequence of Anoectochilus roxburghii.

The complete chloroplast sequence of the Anoectochilus roxburghii, a popular traditional Chinese medicine for the treatment of cancer, was determined in this study. The chloroplast genome (cpDNA)^ was 152,802 bp in length, containing a pair of inverted repeats of 52,728 bp separated by a large single-copy region and a small single-copy region of 82,641 bp and 17,433 bp, respectively. The chloroplast genome encodes 116 predicted functional genes, including 81 protein-coding genes, four ribosomal RNA genes, and 31 transfer RNA genes, 25 of which are duplicated in the inverted repeat regions. The cpDNA is GC-rich (36.9%).

Rice grassy stunt virus nonstructural protein p5 serves as a viral suppressor of RNA silencing and interacts with nonstructural protein p3.

Rice grassy stunt virus (RGSV), a member of the genus Tenuivirus, causes serious rice disease in Southeast Asian countries. In this study, a green fluorescent protein (GFP)-based transient expression assay was conducted to show that p5, encoded on RNA5 in the viral sense, is a viral suppressor of RNA silencing (VSR). Protein-protein interactions (PPIs) between p5 and all RGSV proteins except pC1 and pC2 were investigated using Gal4-based yeast two-hybrid (Y2H) experiments. The results demonstrated that p5 interacts with itself and with p3 encoded on RNA3 in the viral sense. p5-p5 and p5-p3 interactions were detected by bimolecular fluorescence complementation (BiFC) assay, and the p5-p3 interaction was confirmed by subcellular co-localization and co-immunoprecipitation (Co-IP) assays. Using the Y2H system, we demonstrated that the p5-p3 interaction requires both the N-terminal (amino acid residues 1 to 99) and C-terminal (amino acid residues 94 to 191) domains of p5. In addition, either p5 or p3 could enhance the pathogenicity of potato virus X (PVX) in Nicotiana benthamiana plants. A much more significant enhancement of PVX pathogenicity and accumulation was observed when p5 and p3 were expressed together. Our data also showed that RGSV p3 does not function as a VSR, and it had no effect on the VSR activity of p5 or the subcellular localization pattern of p5 in plant cells from Nicotiana benthamiana.