PKR-Like Endoplasmic Reticulum Kinase Is Necessary for Lipogenic Activation during HCMV Infection

Yongjun Yu,Francis J Pierciey,James C Alwine,Tobi G Maguire,Blossom Damania

doi:10.1371/journal.ppat.1003266

Abstract

PKR-like endoplasmic reticulum (ER) kinase (PERK) is an ER-associated stress sensor protein which phosphorylates eukaryotic initiation factor 2α (eIF2α) to induce translation attenuation in response to ER stress. PERK is also a regulator of lipogenesis during adipocyte differentiation through activation of the cleavage of sterol regulatory element binding protein 1 (SREBP1), resulting in the upregulation of lipogenic enzymes. Our recent studies have shown that human cytomegalovirus (HCMV) infection in human fibroblasts (HF) induces adipocyte-like lipogenesis through the activation of SREBP1. Here, we report that PERK expression is highly increased in HCMV-infected cells and is necessary for HCMV growth. Depletion of PERK, using short hairpin RNA (shRNA), resulted in attenuation of HCMV growth, inhibition of lipid synthesis and reduction of lipogenic gene expression. Examination of the cleavage of SREBP proteins showed PERK depletion inhibited the cleavage of SREBP1, but not SREBP2, in HCMV-infected cells, suggesting different cleavage regulatory mechanisms for SREBP1 and 2. Further studies showed that the depletion of SREBP1, but not SREBP2, reduced lipid synthesis in HCMV infection, suggesting that activation of SREBP1 is sufficient to induce lipogenesis in HCMV infection. The reduction of lipid synthesis by PERK depletion can be partially restored by expressing a Flag-tagged nuclear form of SREBP1a. Our studies also suggest that the induction of PERK in HCMV-infected cells stimulates SREBP1 cleavage by reducing levels of Insig1 (Insulin inducible gene 1) protein; this occurs independent of the phosphorylation of eIF2α. Introduction of an exogenous Insig1-Myc into HCMV infected cells significantly reduced HCMV growth and lipid synthesis. Our data demonstrate that the induction of PERK during HCMV infection is necessary for full activation of lipogenesis; this effect appears to be mediated by limiting the levels of Insig1 thus freeing SREBP1-SCAP complexes for SREBP1 processing.

Highlights

Viruses rely on the host cells to make viral proteins, replicate viral genomes and produce infectious virions
We provide evidence that the induction of the unfolded protein response (UPR) is connected to lipogenic activation during human cytomegalovirus (HCMV) infection
We show that the endoplasmic reticulum (ER) stress sensor protein, PKR-like endoplasmic reticulum (ER) kinase (PERK), is critical for lipogenic activation induced during HCMV infection

Summary

Introduction

Viruses rely on the host cells to make viral proteins, replicate viral genomes and produce infectious virions. Studies in the last few years have shown that infection of HCMV can cause dramatic alterations of glucose and glutamine metabolism in the host cells [1,2,3]. Induction of the adipocyte specific glucose transporter 4 (GLUT4), to replace the less efficient GLUT1, allows HCMV infected cells to significantly increase glucose uptake [4]. Instead of producing energy in the tricarboxylic acid (TCA) cycle, a large amount of the glucose-derived carbon exits the mitochondria in the form of citrate to be converted to cytosolic acetyl-CoA to support fatty acid synthesis, which is necessary to support the viral infection [2]. Our recent studies have shown that HCMV infection is able to induce adipocyte-like lipogenesis through the activation of SREBP1 [5]

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Discussion

Conclusion