Nicholas K. Conrad, Ph.D.
Assistant Professor of Microbiology

Office: 214-648-7437
Fax: 214-648-5907
Email:  nicholas.conrad@utsouthwestern.edu

When a virus invades a host cell, it usurps critical components of the cellular machinery to ensure its propagation and, in the case of pathogenic viruses, cause disease.  Therefore, an understanding of the interactions between a virus and its host cell is essential in developing strategies to combat viral disease.  Yet this is not the only reason to study virology.  The history of molecular biology demonstrates that we gain considerable insight into cellular gene function by examining virus-host interactions.  The overarching goals of our lab are to better understand the mechanisms of gene expression and regulation in both Kaposi's sarcoma-associated herpesvirus (KSHV; also known as HHV-8) and in its human host cell.   More specifically, we compare and contrast KSHV and host gene expression at the level of RNA synthesis, stability, and processing.

KSHV is a large (~165 kb) double-stranded DNA gammaherpesvirus whose life cycle has both latent and lytic phases.  During viral latency, the genome is maintained indefinitely as a stable circular episome and few KSHV genes are expressed.  Upon induction of the lytic phase of the virus, a carefully orchestrated cascade of viral gene expression ensues, ultimately resulting in the generation of progeny virus and host cell death.  Importantly, KSHV is the cause of several human diseases including Kaposi's sarcoma (KS), the most common AIDS-associated cancer.  The lymphoproliferative disorders primary effusion lymphoma (PEL) and some subsets of multicentric Castleman's disease (MCD) are also caused by KSHV.  These diseases primarily affect immunocompromised individuals, but KS is also endemic in certain parts of Africa and the Mediterranean. 

The polyadenylated nuclear RNA (PAN; also known as nut-1 and T1.1) is the most abundant KSHV lytic transcript.  In fact, as much as 70% of the polyadenylated transcripts in the lytic phase are PAN!  PAN is a non-coding RNA of unknown function that is transcribed by RNA polymerase II, has a 3´ polyadenylate (polyA) tail, but is not exported from the nucleus.  The impressive nuclear abundance of PAN is due to its high levels of transcription and to a cis-acting RNA element, called the ENE, which increases transcript stability.  Our studies use this unique RNA to probe differences between cellular and viral gene expression.

One difference between KSHV and host genes is that, while the majority of mammalian genes contain introns, most KSHV genes (including PAN) do not.  This is important because, in many cases, introns and/or their removal by splicing are critical for high levels of gene expression.  Indeed, the KSHV ORF57 protein post-transcriptionally increases mRNA levels from intronless genes.  ORF57 is essential for viral lytic replication as are its homologs in other herpesviruses.  These homologous proteins are thought to enhance mRNA export and ORF57 may, at least in part, also increase export efficiency.  However, other laboratories have shown that ORF57 increases PAN RNA levels.  Since PAN RNA is not exported, we seek to determine the pathway by which ORF57 increases PAN RNA levels and to uncover the mechanistic details of ORF57-mediated up-regulation. 

Other areas of interest include examination of the interrelationships between polyadenylation, polyA tail length and nuclear RNA stability.  Extensive work in yeast has shown that polyadenylated nuclear transcripts are degraded by specific mechanisms, but these decay pathways remain elusive in mammalian nuclei.  Since PAN RNA is exclusively localized to the nucleus and its stability is controlled by the presence of the ENE, we can use it as a model transcript to examine the decay of nuclear polyadenylated RNA.  In addition, we are interested in transcription termination in KSHV genes.  While KSHV utilizes the mammalian transcription apparatus, KSHV genes are relatively close to one another.  If a polymerase does not properly terminate at an upstream gene, this could lead to interference with downstream gene expression.  Therefore, we are asking whether KSHV genes use a different mechanism(s) to regulate elongation/termination efficiency of RNA pol II.

The work in our lab is focused on obtaining a better understanding of gene expression mechanisms in both KSHV and host genes.  Understanding the molecular mechanisms that underlie the gene expression of KSHV may, in the long term, lead to insights into how to inhibit viral pathogenicity.  Moreover, we hope these studies will elucidate human cell biology by exploring the fundamental mechanisms of gene expression. 

Selected Publications:

Conrad, N.K., Shu M.D., Uyhazi K.E., and Steitz, J.A. (2007)  Mutational analysis of a viral RNA element that counteracts rapid RNA decay by interaction with the polyadenylate tail  Proc. Natl. Acad. Sci USA.  104: 10412-10417.

Conrad, N. K., Fok, V., Cazalla, D., Borah, S., and Steitz, J. A. (2006) The challenge of viral snRNPs Cold Spr. Harb. Symp. Quant. Biol. 71:377-84.

Conrad, N. K., Mili, S., Marshall, E. L., Shu, M. D., and Steitz, J. A. (2006) Identification of a rapid mammalian deadenylation-dependent decay pathway and its inhibition by a viral RNA element Mol Cell 24, 943-953.

Conrad, N. K., and Steitz, J. A. (2005) A Kaposi's sarcoma virus RNA element that increases the nuclear abundance of intronless transcripts EMBO J 24, 1831-1841.