Karyopherin Alpha 6 Is Required for Replication of Porcine Reproductive and Respiratory Syndrome Virus and Zika Virus

Abstract

Movement of macromolecules between the cytoplasm and the nucleus occurs through the nuclear pore complex (NPC). Karyopherins comprise a family of soluble transport factors facilitating the nucleocytoplasmic translocation of proteins through the NPC. In this study, we found that karyopherin α6 (KPNA6; also known as importin α7) was required for the optimal replication of porcine reproductive and respiratory syndrome virus (PRRSV) and Zika virus (ZIKV), which are positive-sense, single-stranded RNA viruses replicating in the cytoplasm. The KPNA6 protein level in virus-infected cells was much higher than that in mock-infected controls, whereas the KPNA6 transcript remains stable. Viral infection blocked the ubiquitin-proteasomal degradation of KPNA6, which led to an extension of the KPNA6 half-life and the elevation of the KPNA6 level in comparison to mock-infected cells. PRRSV nsp12 protein induced KPNA6 stabilization. KPNA6 silencing was detrimental to the replication of PRRSV, and KPNA6 knockout impaired ZIKV replication. Moreover, KPNA6 knockout blocked the nuclear translocation of PRRSV nsp1β but had a minimal effect on two other PRRSV proteins with nuclear localization. Exogenous restitution of KPNA6 expression in the KPNA6-knockout cells results in restoration of the nuclear translocation of PRRSV nsp1β and the replication of ZIKV. These results indicate that KPNA6 is an important cellular factor for the replication of PRRSV and ZIKV. Positive-sense, single-stranded RNA (+ssRNA) viruses replicate in the cytoplasm of infected cells. The roles of transport factors in the nucleocytoplasmic trafficking system for the replication of +ssRNA viruses are not known. In this study, we discovered that PRRSV and ZIKV viruses needed karyopherin α6 (KPNA6), one of the transport factors, to enhance the virus replication. Our data showed that viral infection induced an elevation of the KPNA6 protein level due to an extension of the KPNA6 half-life via viral interference of the ubiquitin-proteasomal degradation of KPNA6. Notably, KPNA6 silencing or knockout dramatically reduced the replication of PRRSV and ZIKV. PRRSV nsp1β depended on KPNA6 to translocate into the nucleus. In addition, exogenous restitution of KPNA6 expression in KPNA6-knockout cells led to the restoration of nsp1β nuclear translocation and ZIKV replication. These results reveal a new aspect in the virus-cell interaction and may facilitate the development of novel antiviral therapeutics.

Development of luciferase-linked antibody capture assay based on luciferase immunoprecipitation systems for antibody detection of porcine reproductive and respiratory syndrome virus

“…However, the two types of assay differ in reporter enzymes used (HRP for ELISA, luciferase for LACA), substrates (TMB for ELISA, luciferin for LACA), antigens (plate-coated antigen for ELISA, cell lysates for LACA) and opacity of plates (transparent for ELISA, black for LACA). Advantages of LACA over ELISA include the fact that luciferase-linked viral antigens used in LACA can be directly obtained from lysates of plasmid-transfected cells without requiring purification procedures [33][34][35][36]. Moreover, luciferase activity in LACA generates highly quantitative data spanning a relatively wider range of values (from 10 3 to 10 6 ) than for ELISA, which is more suitable for evaluating dynamic changes in antibody responses against a specific antigen over time.…”

Zika virus NS2A protein induces the degradation of KPNA2 (karyopherin subunit alpha 2) via chaperone-mediated autophagy

“…However, ZIKV-cell interactions are not well understood. Our earlier study demonstrated that ZIKV infection induces the elevation of KPNA6, another KPNA isoform, and that KPNA6 knockout dampens ZIKV replication [55]. During the study of KPNA6, we wondered if ZIKV had any effect on other KPNA isoforms.…”

The Network of Interactions Among Porcine Reproductive and Respiratory Syndrome Virus Non-structural Proteins

“…Another study have shown that nsp12 induces the phosphorylation of STAT1 at serine 727 and the expression of proinflammatory cytokines IL-1β and IL-8 ( Yu et al, 2013 ). In a most recent report, nsp12 is documented to be able to induce the stabilization of karyopherin alpha6 which is required for the nuclear translocation of nsp1β and the PRRSV replication ( Yang et al, 2018 ). These previous data have demonstrated the involvement of PRRSV nsp12 in the virus infection by modulating cellular factors.…”