Identification and Characterisation of Human Cytomegalovirus-Mediated Degradation of Helicase-Like Transcription Factor

Viruses are known to degrade host factors that are important in innate antiviral immunity in order to infect successfully. To systematically identify host proteins targeted for early degradation by human cytomegalovirus (HCMV), the lab developed orthogonal screens using high resolution multiplexed mass spectrometry. Taking advantage of broad and selective proteasome and lysosome inhibitors, proteasomal degradation was found to be heavily exploited by HCMV. Several known antiviral restriction factors, including components of cellular promyelocytic leukemia (PML) were enriched in a shortlist of proteasomally degraded proteins during infection. A particularly robust novel ‘hit’ was helicase-like transcription factor (HLTF), a DNA repair protein that participates in error-free repair of stalled replication forks. HLTF was found degraded very early during infection, and its expression remained low throughout the course of HCMV lytic cycle. De novo expression of UL145, a previously uncharacterized viral protein, was found necessary and sufficient to degrade HLTF via recruitment of the cullin 4/DDB1 E3 ligase complex. HLTF degradation was reported in human immunodeficiency virus (HIV) infection, however the interaction between HLTF and viruses remain largely elusive. The roles of UL145 were explored in hopes of understanding functions of HLTF in HCMV infection. UL145 was identified as a non-essential immediate early protein, however had a possible role in type I interferon (IFN) induction regulation in later stages of HCMV lytic progression. As the key host protein to be rescued by UL145 deletion, depletion of HLTF was found to transiently impair IFNβ transcription and HCMV infection. I hypothesise that HLTF is an undiscovered nuclear viral DNA sensor that triggers an antiviral interferon response during viral DNA replication. Additionally, work presented here expands the range of powerful screening technologies to identify HCMV restriction factor candidates by identifying virally degraded host proteins. Further investigation of these candidates will contribute to our understanding of how HCMV modulates host protein expression to evade antiviral factors.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2021

Author dc:creator

• Lin, Kai-Min

Advisor dc:contributor.advisor

• Weekes, Michael

• human cytomegalovirus
• UL145
• helicase-like transcription factor
• protein degradation
• TMT proteomics
• type I interferon
• antiviral restriction factor

Licence

• https://apollo8-f-pro.lib.cam.ac.uk/bitstreams/5925b8cf-b793-4347-bfe5-536f31e48f8c/download
• https://creativecommons.org/licenses/by/4.0/

Language dc:language

eng

Author Identifier

0000-0003-2109-1530
0000-0003-3196-5545

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/318048