Contents - College of Medical and Dental Sciences - University of ...
Contents - College of Medical and Dental Sciences - University of ...
Contents - College of Medical and Dental Sciences - University of ...
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The 11 th International Workshop on KSHV & Related Agents, Birmingham, UK<br />
Virus-Cell Interactions I Abstract 27<br />
KSHV AND CELLULAR MIRNA EXPRESSION DURING LATENCY AND LYTIC<br />
REACTIVATION IN PEL CELLS<br />
*1Soo-Jin Han * 1 Jianhong Hu, * 1 Karlie Plaisance * 2 Wendell Miley 2 Rachel Bagni, Chang<br />
Hee Kim 3 , 2 Denise Whitby <strong>and</strong> 2 Rolf Renne<br />
1. Department <strong>of</strong> Molecular Genetics <strong>and</strong> Microbiology, <strong>University</strong> <strong>of</strong> Florida, Gainesville,<br />
Florida 32610, 2.Viral Oncology Section, AIDS <strong>and</strong> Cancer Virus Program, SAIC-<br />
Frederick, NCI-Frederick, Frederick MD 21702 3. Laboratory <strong>of</strong> Molecular Technology,<br />
Advanced Technology Program, SAIC-Frederick, NCI-Frederick, Frederick, MD 21702<br />
(*equal contribution).<br />
Abstract<br />
MicroRNAs are small non-coding RNA molecules which post-transcriptionally regulate<br />
gene expression by binding to 3’UTRs <strong>of</strong> target genes thereby contributing to important<br />
biological processes. Kaposi’s sarcoma associated herpesvirus (KSHV) encodes 12<br />
miRNAs within the KSHV latency associated region (KLAR), some <strong>of</strong> which target host<br />
cellular genes thereby affecting angiogenesis, apoptosis, <strong>and</strong> B cell development.<br />
Originally, KSHV miRNAs were cloned from BCBL-1 cells in the presence <strong>and</strong> absence <strong>of</strong><br />
TPA induction, suggesting that miRNAs may contribute to both phases <strong>of</strong> infection.<br />
The KLAR region is expressed from at least three different promoters giving rise to singly<br />
<strong>and</strong> multiply spliced transcripts. To further elucidate the complexity <strong>of</strong> this locus with<br />
emphasis on transcripts that could serve as pri-miRNAs, we performed a detailed RT-PCR<br />
<strong>and</strong> RNase protection analysis <strong>and</strong> identified several new transcripts that could give rise<br />
to miRNA expression. We identified novel multiply-spliced transcripts originating from the<br />
LANAp <strong>and</strong> LTI promoters upstream <strong>of</strong> LANA, which contained 3’ exons within the<br />
Kaposin locus. Furthermore, the observed splicing patterns from these long KLAR<br />
encompassing transcripts changed during lytic replication, suggesting that both LANAp<br />
<strong>and</strong> LTI could drive miRNA expression.<br />
Next, we utilized a custom miRNA array containing both viral <strong>and</strong> human miRNAs to<br />
investigate miRNA output in PEL cells during latency <strong>and</strong> reactivated by Ad-ORF50.<br />
Surprisingly, while we identified marked changes in the splicing pattern in this region,<br />
viral miRNA expression was not significantly altered in PEL cells 48 hours post<br />
reactivation. Cellular miRNA expression signatures during both conditions will also be<br />
discussed.<br />
Presenting author Email: rrenne@ufscc.ufl.edu<br />
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