hnRNPs Relocalize to the cytoplasm following infection with vesicular stomatitis virus.
Vesicular stomatitis virus (VSV) matrix protein inhibits nuclear-cytoplasmic mRNA transport. The goal of this work is to determine whether VSV inhibits the nuclear-cytoplasmic transport of heterogeneous ribonucleoproteins (hnRNPs), which are thought to serve as mRNA export factors. Confocal microscopy experiments showed that hnRNPA1, hnRNPK, and hnRNPC1/C2, but not hnRNPB1 or lamin A/C, are relocalized to the cytoplasm during VSV infection. We determined whether protein import is inhibited by VSV by transfecting cells with a plasmid encoding enhanced green fluorescent protein (EGFP) tagged with either the M9 nuclear localization sequence (NLS) or the classical NLS. These experiments revealed that both the M9 NLS and the classical NLS are functional during VSV infection. These data suggest that the inhibition of protein import is not responsible for hnRNP relocalization during VSV infection but that hnRNP export is enhanced. We found that hnRNPA1 relocalization was significantly reduced following the silencing of the mRNA export factor Rae1, indicating that Rae1 is necessary for hnRNP export. In order to determine the role of hnRNPA1 in VSV infection, we silenced hnRNPA1 in HeLa cells and assayed three aspects of the viral life cycle: host protein synthesis shutoff concurrent with the onset of viral protein synthesis, replication by plaque assay, and cell killing. We observed that host shutoff and replication are unaffected by the reduction in hnRNPA1 but that the rate of VSV-induced apoptosis is slower in cells that have reduced hnRNPA1. These data suggest that VSV promotes hnRNPA1 relocalization in a Rae1-dependent manner for apoptotic signaling.
Pettit Kneller
Elizabeth L
EL
Department of Biochemistry, Wake Forest University School of Medicine, Medical Center Boulevard, Winston-Salem, North Carolina 27157, USA.
Connor
John H
JH
Lyles
Douglas S
DS
D017209:Apoptosis; D003593:Cytoplasm / Q000737:chemistry*; D020868:Gene Silencing; D006367:HeLa Cells; D034441:Heterogeneous-Nuclear Ribonucleoproteins / Q000378:metabolism*; D006801:Humans; D018613:Microscopy, Confocal; D034521:Nuclear Matrix-Associated Proteins / Q000378:metabolism; D029543:Nucleocytoplasmic Transport Proteins / Q000378:metabolism; D018116:Vesiculovirus / Q000502:physiology*; D010948:Viral Plaque Assay; D014779:Virus Replication
D034441:Heterogeneous-Nuclear Ribonucleoproteins; D034521:Nuclear Matrix-Associated Proteins; D029543:Nucleocytoplasmic Transport Proteins; C490522:RAE1 protein, human