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| Status |
Public on Jun 11, 2020 |
| Title |
Acetylation of cytidines by NAT10 enhances HIV-1 replication through stabilization of viral RNA |
| Organism |
Homo sapiens |
| Experiment type |
Other
|
| Summary |
As obligate parasites, viruses need to navigate a variety of cellular regulatory systems while infecting and replicating in the host cell. Post-transcriptional modifications have recently emerged as an important layer of regulation of viral RNA function. For example, our lab and others have shown that the RNA modification N6-methyladenosine (m6A) can enhance the replication of multiple viruses in cis, including Human Immunodeficiency virus 1 (HIV-1), Influenza A virus, SV40 and Kaposi's sarcoma-associated herpesvirus (KSHV). Recent reports have revealed the presence of another RNA modification, N4-acetylcytidine (ac4C) on cellular mRNAs and have shown that ac4C can enhance mRNA stability and translation. Here, we demonstrate that ac4C is present at multiple sites on HIV-1 mRNAs and on the viral genomic RNA. Through ac4C RNA immunoprecipitation (RNA-IP) and RNA-seq, we found ac4C on HIV-1 mRNAs as well as the virion genomic RNA, with ac4C sites in the coding regions of the pol, env, nef genes, and the trans-activation response (TAR) hairpin. Phenotypically, we observe that increasing the expression level of the ac4C acetyltransferase NAT10 leads to an increase in viral replication that is dependent on the RNA binding and enzymatic domains of NAT10. Moreover, both CRISPR-depletion of NAT10 (Ξ”NAT10) and treatment with the small molecule NAT10 inhibitor Remodelin, diminishes HIV-1 replication in T cells. Lastly, silent mutations introduced to prevent ac4C deposition on the viral genome indeed diminished HIV-1 replication. Our data suggest that HIV-1 has evolved to incorporate ac4C in essential viral gene coding regions and regulatory RNA structures, and that NAT10-dependent ac4C addition enhances HIV-1 replication.
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| Overall design |
PA-ac4C-seq on poly(A)+ RNA from mock-infected or HIV-1-infected CEM-SS and CEM cells; PA-ac4C-seq on RNA extracted from HIV-1 infected CEM-SS, SupT1, and CEM cells; PA-ac4C-seq validation of ac4C depletion in virions produced from control or NAT10-depleted CEM cells; GFP (control) or NAT10 PAR-CLIP on HIV-1-infected CEM cells.
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| Contributor(s) |
Tsai K, Cullen BR |
| Citation(s) |
32533923 |
| NIH grant(s) |
| Grant ID |
Grant title |
Affiliation |
Name |
| R01 DA046111 |
Epitranscriptomic modification of HIV-1 transcripts: Effects of drugs of abuse |
DUKE UNIVERSITY |
BRYAN R. CULLEN |
| U54 AI150470 |
The Center for HIV RNA Studies (CRNA) |
REGENTS OF THE UNIVERSITY OF MICHIGAN - ANN ARBOR |
ALICE TELESNITSKY |
| P30 AI064518 |
Centers for AIDS Research (CFAR) |
DUKE UNIVERSITY |
Kent J. Weinhold |
|
| Submission date |
Dec 21, 2019 |
| Last update date |
Sep 10, 2020 |
| Contact name |
Bryan R. Cullen |
| E-mail(s) |
bryan.cullen@duke.edu
|
| Organization name |
Duke University Medical Center
|
| Department |
Molecular Genetics and Microbiology
|
| Street address |
213 Research Dr. 420 CARL Bldg.
|
| City |
Durham |
| State/province |
NC |
| ZIP/Postal code |
27710 |
| Country |
USA |
| |
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| Platforms (3) |
| GPL18573 |
Illumina NextSeq 500 (Homo sapiens) |
| GPL20301 |
Illumina HiSeq 4000 (Homo sapiens) |
| GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
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| Samples (12)
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| Relations |
| BioProject |
PRJNA597154 |
| SRA |
SRP238484 |
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