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| Status |
Public on Feb 01, 2015 |
| Title |
HITS-CLIP analysis uncovers a link between the Kaposi's sarcoma associated herpesvirus ORF57 protein and host pre-mRNA metabolism |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
The Kaposi's sarcoma associated herpesvirus (KSHV) is an oncogenic virus that causes Kaposi's sarcoma, primary effusion lymphoma (PEL), and some forms of multicentric Castleman's disease. The KSHV ORF57 protein is a conserved posttranscriptional regulator of gene expression that is essential for virus replication. ORF57 is multifunctional, but most of its activities are directly linked to its ability to bind RNA. We globally identified virus and host RNAs bound by ORF57 during lytic reactivation in PEL cells using high-throughput sequencing of RNA isolated by crosslinking immunoprecipitation (HITS-CLIP). As expected, ORF57-bound RNA fragments mapped throughout the KSHV genome, including the known ORF57 ligand PAN RNA. In agreement with previously published ChIP results, we observed that ORF57 bound RNAs near the oriLyt regions of the genome. Examination of the host RNA fragments revealed that a subset of the ORF57-bound RNAs was derived from transcript 5´ ends. The position of these 5´-bound fragments correlated closely with the 5´-most exonintron junction of the pre-mRNA. We selected four candidates (BTG1, EGR1, ZFP36, and TNFSF9) and analyzed their pre-mRNA and mRNA levels during lytic phase. Analysis of both steady-state and newly made RNAs revealed that these candidate ORF57-bound pre-mRNAs persisted for longer periods of time throughout infection than control RNAs, consistent with a role for ORF57 in pre-mRNA metabolism. In addition, exogenous expression of ORF57 was sufficient to increase the pre-mRNA levels and, in one case, the mRNA levels of the putative ORF57 targets. These results demonstrate that ORF57 interacts with specific host pre-mRNAs during lytic reactivation and alters their processing, likely by stabilizing pre-mRNAs. These data suggest that ORF57 is involved in modulating host gene expression in addition to KSHV gene expression during lytic reactivation.
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| Overall design |
HITS-CLIP was performed on TREx BCBL-Rta cells 20 hpi using antibodies against ORF57. Three biological replicates were performed.
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| Contributor(s) |
Sei E, Wang T, Volovnik Hunter O, Xie Y, Conrad NK |
| Citation(s) |
25710169 |
| Submission date |
Dec 22, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Tao Wang |
| E-mail(s) |
tao.wang@utsouthwestern.edu
|
| Organization name |
UT Southwestern Medical Center
|
| Street address |
5323 Harry Hines Boulevard
|
| City |
Dallas |
| State/province |
TX |
| ZIP/Postal code |
75390 |
| Country |
USA |
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| Platforms (1) |
| GPL16791 |
Illumina HiSeq 2500 (Homo sapiens) |
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| Samples (6)
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| Relations |
| BioProject |
PRJNA270976 |
| SRA |
SRP051468 |
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