Human Cytomegalovirus Degrades DMXL1 to Inhibit Autophagy, Lysosomal Acidification and Viral Assembly

Abstract

Human cytomegalovirus (HCMV) is an important human pathogen and a master regulator of host intrinsic, innate, and adaptive immunity. HCMV hijacks host intracellular compartments to assemble new virions, and lysosomes are essential for this process. We combined a comprehensive proteomic analysis of host proteins targeted for degradation by HCMV with a database of proteins involved in vacuolar acidification. Dmx-like protein 1 (DMXL1) was the only protein that acidifies vacuoles yet is degraded by HCMV.Systematic comparison of viral deletion mutants revealed that the uncharacterised 7kDa US33A protein is necessary and sufficient for DMXL1 degradation. Functional experiments using cells stably expressing US33A, or infected with adenovirus vectors expressing US33A, or infected with recombinant HCMV deleted for US33A, demonstrated that HCMV degrades DMXL1 to inhibit lysosomal acidification and autophagic cargo degradation. Formation of the viral assembly compartment, which is known to require lysosomes, occurred significantly later in cells infected with US33A-expressing virus, with reduced viral replication. These data thus identify an entirely new viral strategy for cellular remodelling. US33A recruits the E3 ubiquitin ligase Kip1 ubiquitination-promoting complex (KPC) and acts akin to a proteolysis-targeting chimera (PROTAC) to degrade DMXL1. The potential thus exists to employ US33A in novel therapies for viral infection or rheumatic conditions, in which inhibition of lysosome acidification can attenuate disease.

Note:
Funding Information: This work was supported by a Medical Research Council Project Grant (MR/X000516/1) to MPW, a MRC grant (MR/S00971X/1) to RJS, the UK Dementia Research Institute (funded by the MRC, Alzheimer’s Research UK and the Alzheimer’s Society), The Cambridge Commonwealth, European & International Trust (SK), the Nehru Trust for Cambridge University and the Trinity-Henry Barlow Scholarship (SK) and the NIHR Cambridge Biomedical Research Centre (NIHR203312).

Declaration of Interests: DCR is a consultant for Aladdin Healthcare Technologies Ltd., Mindrank AI, Nido Biosciences, Drishti Discoveries, Retro Biosciences and PAQ Therapeutics. None of the other authors have any interests to declare.

Keywords: proteomics, systems virology, human cytomegalovirus, lysosome, pH, viral replication, viral assembly, autophagy