期刊论文详细信息
PLoS Pathogens
Phosphoproteomic Analysis of KSHV-Infected Cells Reveals Roles of ORF45-Activated RSK during Lytic Replication
Zachary Turpin1  Denis Avey1  Wenwei Li1  Fanxiu Zhu1  Sarah Tepper1 
[1] Department of Biological Science, Florida State University, Tallahassee, Florida, United States of America
关键词: Phosphorylation;    Kaposi's sarcoma-associated herpesvirus;    Protein translation;    Virions;    Viral replication;    Transcriptional control;    DNA-binding proteins;    Gene expression;   
DOI  :  10.1371/journal.ppat.1004993
学科分类:生物科学(综合)
来源: Public Library of Science
PDF
【 摘 要 】

Kaposi’s Sarcoma-Associated Herpesvirus (KSHV) is an oncogenic virus which has adapted unique mechanisms to modulate the cellular microenvironment of its human host. The pathogenesis of KSHV is intimately linked to its manipulation of cellular signaling pathways, including the extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase (MAPK) pathway. We have previously shown that KSHV ORF45 contributes to the sustained activation of both ERK and p90 ribosomal S6 kinase (RSK, a major functional mediator of ERK/MAPK signaling) during KSHV lytic replication. ORF45-activated RSK is required for optimal KSHV lytic gene expression and progeny virion production, though the underlying mechanisms downstream of this activation are still unclear. We hypothesized that the activation of RSK by ORF45 causes differential phosphorylation of cellular and viral substrates, affecting biological processes essential for efficient KSHV lytic replication. Accordingly, we observed widespread and significant differences in protein phosphorylation upon induction of lytic replication. Mass-spectrometry-based phosphoproteomic screening identified putative substrates of ORF45-activated RSK in KSHV-infected cells. Bioinformatic analyses revealed that nuclear proteins, including several transcriptional regulators, were overrepresented among these candidates. We validated the ORF45/RSK-dependent phosphorylation of several putative substrates by employing KSHV BAC mutagenesis, kinase inhibitor treatments, and/or CRISPR-mediated knockout of RSK in KSHV-infected cells. Furthermore, we assessed the consequences of knocking out these substrates on ORF45/RSK-dependent regulation of gene expression and KSHV progeny virion production. Finally, we show data to support that ORF45 regulates the translational efficiency of a subset of viral/cellular genes with complex secondary structure in their 5’ UTR. Altogether, these data shed light on the mechanisms by which KSHV ORF45 manipulates components of the host cell machinery via modulation of RSK activity. Thus, this study has important implications for the pathobiology of KSHV and other diseases in which RSK activity is dysregulated.

【 授权许可】

CC BY   

【 预 览 】
附件列表
Files Size Format View
RO201902015503129ZK.pdf 6220KB PDF download
  文献评价指标  
  下载次数:14次 浏览次数:7次