分类: 生物学 >> 生物物理学 >> 生物物理、生物化学与分子生物学 提交时间: 2016-05-12
摘要: Replication and transcription of influenza virus genome mainly depend on its RNA-dependent RNA polymerase (RdRP), composed of the PA, PB1, and PB2 subunits. Although extensively studied, the underlying mechanism of the RdRP complex is still unclear. Here we report the biochemical characterization of influenza RdRP subcomplex comprising PA, PB1, and N terminus of PB2, which exist as dimer in solution and can assemble into a tetramer state, regulated by vRNA promoter. Using single-particle cryo-electron microscopy, we have reconstructed the RdRP tetramer complex at 4.3 angstrom, highlighting the assembly and interfaces between monomers within the tetrameric structure. The individual RdRP subcomplex contains all the characterized motifs and appears as a cage-like structure. High-throughput mutagenesis profiling revealed that residues involved in the oligomer state formation are critical for viral life cycle. Our results lay a solid base for understanding the mechanism of replication of influenza and other negative-stranded RNA viruses.
分类: 生物学 >> 生物物理学 提交时间: 2016-05-12
摘要: Non-homologous end joining (NHEJ) is a major pathway to repair DNA double-strand breaks (DSBs), which can display different types of broken ends. However, it is unclear how NHEJ factors organize to repair diverse types of DNA breaks. Here, through systematic analysis of the human NHEJ factor interactome, we identify PAXX as a direct interactor of Ku. The crystal structure of PAXX is similar to those of XRCC4 and XLF. Importantly, PAXX-deficient cells are sensitive to DSB-causing agents. Moreover, epistasis analysis demonstrates that PAXX functions together with XLF in response to ionizing radiation-induced complex DSBs, whereas they function redundantly in response to Topo2 inhibitor-induced simple DSBs. Consistently, PAXX and XLF coordinately promote the ligation of complex but not simple DNA ends in vitro. Altogether, our data identify PAXX as a new NHEJ factor and provide insight regarding the organization of NHEJ factors responding to diverse types of DSB ends.