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  • Nuclear export mediated by CRM

    2019-07-11

    Nuclear export mediated by CRM1 is initiated by an interaction between RanGTP and the HEAT9 loop of CRM1, triggering a conformational change in the NES binding site that permits NES–cargo protein binding (Dong et al., 2009, Fung and Chook, 2014). Our docking simulation results predicted that DP2392-E10 binds at the region around the HEAT9 and HEAT10 repeats of CRM1. This may directly cause a conformational change of the NES binding site of CRM1, or indirectly affect the folding of the HEAT9 loop that is critical for RanGTP binding. Thus, these unusual conformational changes of CRM1 decrease the affinity of binding with the NP-NES3 domain, interfere with the nuclear export function of NP, and inhibit viral replication. This proposed mechanism of inhibition by DP2392-E10 differs from that of LMB or Verdinixor, which directly bind the C258 of CRM1 NES binding site to prevent the CRM1/NES–cargo protein interaction (Kudo et al., 1999, Perwitasari et al., 2014). Our CELAVIEW analysis indicated that DP2392-E10 also inhibits nuclear export of HIV-1 Rev-NES, which plays a well-defined role in nuclear export of HIV-1 unspliced and singly spliced RNAs for viral protein production and viral particle assembly (Fornerod et al., 1997). Antiviral activity against intact HIV-1 S-Adenosylhomocysteine is further worthily verification that will provide a new class of anti-HIV-1 inhibitor based on the Rev nuclear export function for the highly active antiretroviral therapy. In addition, DP2392-E10 is expected to exert an antiviral effect against a broad range of viruses that exploit the cellular CRM1 nuclear export machinery, including severe acute respiratory syndrome coronavirus (Freundt et al., 2009), hepatitis viruses (Cerutti et al., 2011, Forgues et al., 2001), and herpes simplex virus (Williams et al., 2008).
    Materials and methods
    Acknowledgments We thank the Platform for Drug Discovery, Informatics, and Structural Life Science of the Ministry of Education, Culture, Sports, Science and Technology, Japan, for providing DP2392-E10. We are also grateful to Dr. Shinji Watanabe (Center for Influenza Virus Research, National Institute of Infectious Diseases, Japan), Dr. Hiroshi Kida, and Dr. Yoshihiro Sakoda (Graduate School of Veterinary Medicine, Hokkaido University, Japan) for giving influenza A subtype viruses, Dr. Yoshihiro Kawaoka (Institute of Medical Science, University of Tokyo, Japan) for providing the plasmids used for reverse genetics, and Ms. Kaori Honda (Bio-Active Compounds Discovery Research Unit, RIKEN CSRS, Japan) for technical support with DP2392-E10 photo-crosslinked Sepharose beads. This work was supported in part by a research project of the Ministry of Agriculture, Forestry and Fisheries of Japan aimed at improving food safety and animal health (grant number 13406421), and by the Japan Society for the Promotion of Science (JSPS) Postdoctoral Fellowship (grant number 15F15416).
    Introduction Dendritic cells (DC) are the most potent antigen-presenting cells (APC) of the immune system and are specialized to trigger primary immune responses. Because of their efficiency to stimulate naïve T helper cells and cytotoxic T cells DC have been designated as “nature\'s adjuvants” (Banchereau and Steinman 1998; Banchereau et al. 2000; Mellman and Steinman 2001). Immature DC, dedicated to the myeloid line, are located at the peripheral organs and the interstitium (or interstitial fluid). They are specialized to capture and to process antigens into immunogenic peptides that are subsequently presented by major histocompatibility complex (MHC) complexes. DC maturation is induced by inflammatory signals like microbiological and viral structures or inflammatory stimuli. The maturation process of DC is accompanied by migration of the DC to the draining lymph nodes. During maturation the DC are significantly changing their morphology, a process that is highlighted by the development of motile veils. Moreover, the development of immature DC into potent T cell stimulators is accompanied by an enhanced expression of costimulatory surface proteins, like CD80 and CD86, as well as adhesion molecules (Dieu et al. 1998).