Restriction of Retrotransposition and Retroviral Infection by RNase L and APOBEC3G

The OAS/RNase L system is one of the major restriction pathways activated during the type I interferon response to viral infections. In this thesis I primarily explored its non-canonical functions beyond blocking viruses through degrading viral and cellular single-stranded RNA. Using genetically-engineered retrotransposon constructs, I demonstrated that wild type and constitutively-active truncated human RNase L, but not a nuclease-dead RNase L mutant (R667A), impaired retrotransposition of both human LINE-1 and mouse IAP elements (Chapter 2). Depletion of endogenous RNase L with siRNA increased levels of LINE-1 retrotransposition. Wild type RNase L, but not RNase L R667A, dramatically reduced L1 RNA accumulation, expression of both L1 open reading frame proteins and formation of LINE-1 ribonucleoprotein particles. By controlling dysregulated LINE-1 retrotransposition, RNase L could contribute to the maintenance of genomic stability and fulfill its tumor suppressor function. The interaction between RNase L and APOBEC3G during HIV-1 delta Vif infection was investigated in Chapter 3. I found both A3G and RNase L were packaged into HIV-1 delta Vif virions. Virions containing A3G in the absence of RNase L exhibited higher infectivity compared to virons containing both proteins, which correlated with decreased numbers of A3G-mediated mutations of viral cDNA recovered from infected cells. Activation of RNase L by 2'-5' oligoadenylate caused redistribution of A3G from enzymatically-inactive high molecular mass complex to enzymatically-active low molecular mass complex. Virion-associated A3G exhibited elevated cytidine deaminase activity in the presence of RNase L. These results supported a regulatory mechanism for A3G during HIV-1 infections whereby RNase L plays an activating role. Finally in Chapter 5, I determined the in vivo hypermutation characteristic of non-human primate APOBEC3 proteins with XMRV as a model retrovirus. Editing signatures of A3G, as well as A3F and A3DE, were observed in the XMRV proviral sequences from rhesus macaques inoculated intravenously. Mutation and inhibition of XMRV by rhesus A3 proteins were further validated in cultured human cells. Together, results from this thesis shed light on the intricate yet complex control of retrotransposition and retroviral Infection by host restriction factors RNase L and APOBEC3G.

Name thesis:degree_name

Doctor of Philosophy

Level thesis:degree_level

doctoral

Discipline thesis:degree_discipline

Molecular Medicine

Grantor dc:publisher

Case Western Reserve University School of Graduate Studies

Year dc:date

2013

Author dc:creator

• Zhang, Ao

Contributors dc:contributor

• Silverman, Robert
• Stark, George

• Biomedical Research

Statement dc:rights

• unrestricted
• This thesis or dissertation is protected by copyright: all rights reserved. It may not be copied or redistributed beyond the terms of applicable copyright laws.

Language dc:language

English

Repository record dc:identifier

http://rave.ohiolink.edu/etdc/view?acc_num=case1364979902

OAI identifier oai:identifier

oai:etd.ohiolink.edu:case1364979902