The human cytomegalovirus ER resident glycoprotein UL148 activates the unfolded protein response

Abstract

17Eukaryotic cells are equipped with three sensors that respond to the accumulation of 18 misfolded proteins within the lumen of the endoplasmic reticulum (ER) by activating the 19 unfolded protein response (UPR), which functions to resolve proteotoxic stresses 20 involving the secretory pathway. Here, we identify UL148, a viral ER resident 21 glycoprotein from human cytomegalovirus (HCMV) as an inducer of the UPR. Metabolic 22labeling results indicate that global mRNA translation is markedly decreased when 23 UL148 expression is induced in uninfected cells. Further, we find evidence suggesting 24 that ectopic expression of UL148 is sufficient to activate at least two UPR sensors: the 25 inositol requiring enzyme-1 (IRE1), as indicated by splicing of Xbp1 mRNA, and the 26 PKR-like ER kinase (PERK), as indicated by phosphorylation of eIF2 and 27 accumulation of ATF4 protein. During WT HCMV infection, Xbp-1 splicing, eIF2 28 phosphorylation and ATF4 accumulation neatly accompanied the onset of UL148 29 expression. However, the appearance of these UPR indicators was either markedly 30 delayed or absent during UL148-null infections. siRNA depletion of PERK dampened 31 the extent of eIF2 phosphorylation and ATF4 induction observed during WT infection, 32 further implicating PERK as opposed to other eIF2 kinases. Finally, UL148-null 33 infections showed statistically significant 2-to 4-fold decreases in the levels of 34 transcripts canonically regulated by PERK/ATF4 and by the ATF6 pathway, when 35 compared to WT virus infection. Taken together, our results argue that UL148 is 36 sufficient to activate the UPR when expressed ectopically and that UL148 is an 37 important cause of UPR activation in the context of the HCMV infected cell. 38 39 3 IMPORTANCE. 40

Human Cytomegalovirus Tropism Modulator UL148 Interacts with SEL1L, a Cellular Factor That Governs Endoplasmic Reticulum-Associated Degradation of the Viral Envelope Glycoprotein gO

“…The second mechanism, i.e., the possibility that UL148 generally inhibits ERAD, is buttressed to a degree by other recent findings from our laboratory: UL148 appears to cause ER stress and to strongly contribute to the induction of the unfolded protein response (UPR), both during HCMV infection and when expressed ectopically outside the context of infection (33). Indeed, the negative effects of siRNA depletion of Hrd1 or SEL1L that we observed on viral gene expression for viral proteins other than gO, e.g., pp150 and gB, appeared to be more severe during WT infection than during UL148-null virus infections (Fig.…”

“…Notably, the UPR observed during depletion of core ERAD components such as SEL1L may be exacerbated by UL148. UL148 itself appears to be a substantial source of ER stress and UPR induction during HCMV infection (33), which may explain why pp150 expression is reduced during WT but not UL148-null infection of cells depleted for SEL1L ( Fig. 4A).…”