The Role of BiP in Endoplasmic Reticulum-associated Degradation of Major Histocompatibility Complex Class I Heavy Chain Induced by Cytomegalovirus Proteins

Nagendra R Hegde,Mathieu S Chevalier,Todd W Wisner,Michael C Denton,Kathy Shire,Lori Frappier,David C Johnson

doi:10.1074/jbc.m602989200

Nagendra R Hegde, Mathieu S Chevalier + Show 5 more

Open Access

https://doi.org/10.1074/jbc.m602989200

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Abstract

Human cytomegalovirus (HCMV1) US11 and US2 proteins cause rapid degradation of major histocompatibility complex (MHC) molecules, apparently by ligating cellular endoplasmic reticulum (ER)-associated degradation machinery. Here, we show that US11 and US2 bind the ER chaperone BiP. Four related HCMV proteins, US3, US7, US9, and US10, which do not promote degradation of MHC proteins, did not bind BiP. Silencing BiP reduced US11- and US2-mediated degradation of MHC class I heavy chain (HC) without altering the synthesis or translocation of HC into the ER or the stability of HC in the absence of US11 or US2. Induction of the unfolded protein response (UPR) did not affect US11-mediated HC degradation and could not explain the stabilization of HC when BiP was silenced. Unlike in yeast, BiP did not act by maintaining substrates in a retrotranslocation-competent form. Our studies go beyond previous observations in mammalian cells correlating BiP release with degradation, demonstrating that BiP is functionally required for US2- and US11-mediated HC degradation. Further, US2 and US11 bound BiP even when HC was absent and degradation of US2 depended on HC. These data were consistent with a model in which US2 and US11 bridge HC onto BiP promoting interactions with other ER-associated degradation proteins.

Highlights

major histocompatibility complex (MHC) class I heavy chain (HC) bound by US2 was proposed to be retrotranslocated across Sec61 channels [12], whereas Derlin-1 has been suggested to be the channel for US11-mediated degradation of HC [13, 15]
We and others have suggested that US2 and US11 might bridge MHC complexes onto cellular components of the ERAD machinery (58 – 60), but the identity of such luminal proteins has remained elusive
To identify cellular proteins associated with US11 and US2, the viral proteins were delivered using replication-defective Ad vectors, and radiolabeled U373 cells were extracted with 1% digitonin or 0.5% Nonidet P-40, and the US proteins were immunoprecipitated

Summary

Introduction

BiP promotes degradation of HC in US2- and US11-expressing cells by a mechanism distinct from maintaining HC solubility or “retrotranslocation competence.” US2 and US11 bound to BiP, even in the absence of HC, and bind directly to HC suggesting that the US proteins bridge BiP onto HC to promote binding onto other ERAD machinery. MHC class I or II cell-surface expression or antigen presentation [61, 69], US8 and US10 bind HC without causing degradation [70, 71]. In cells treated with a control siRNA or with transfection reagent alone (no siRNA), US11 caused extensive degradation of MHC class I HC compared with cells expressing US7 or expressing no HCMV proteins, i.e. infected with Adtet-trans alone (Fig. 2C).

Results

Conclusion