Thomas Hope
Research area: Viral Posttranscriptional Regulatory Elements and Cell Biology of HIV
Degree: Ph.D.
Voice: 312.503.1360
Fax: 312.503.7912
e-mail: thope@northwestern.edu  

Detailed research description:
Introns in cellular precursor mRNAs are removed before a mRNA can be exported to the cytoplasm to prevent intron sequences from being translated. The process of splicing itself appear play multiple roles in RNA processing including 3’ end processing, stabilization of the nascent transcript, and facilitate the export of mRNA. To gain insights into these processes we study the posttranscriptional regulation of intronless viral messages. Intronless messages must be efficiently processed in the absence of splicing. Therefore, intronless messages must uncouple RNA processing and export from the splicing process making a simpler model system. We are currently focused on the posttranscriptional regulatory element (PRE) of the Hepadnaviruses, including hepatitis B virus (HBV) and woodchuck hepatitis virus (WPRE). We have found that the WPRE has the ability to posttranscriptionally stimulate the expression of heterologous cDNAs from five to ten-fold, in a variety of vector systems. This ability to stimulate heterologous gene expression has made the WPRE a popular component of vectors for gene therapy and other applications. The practical use of his element to stimulate heterologous gene expression reveals the importance of RNA processing and export in efficient gene expression. Mechanistic studies of the WPRE have found that it functions by the novel mechanism of facilitating efficient RNA processing and increasing poly(A) tail length. This is the first identification of a cis-acting RNA sequence that increases poly(A) tail length. We will identify the factors, which bind to the PREs and mediate their function. Our goal is to understand the novel mechanism of the stimulation of heterologous gene expression by the WPRE. Understanding WPRE function will allow the development of even more efficient gene expression for a variety of applications from gene therapy to large scale protein production.

Although much is known about the molecular biology of HIV, little is known about the details of interactions between the virus and cellular components such as the cytoskeleton. To gain insights into these processes we are combining the disciplines of virology and cell biology to develop the field of cellular virology. Integrating the methods of cell biology with virology will reveal exciting new details of the cell biology of HIV. Our interests range from the understanding of the nucleocytoplasmic trafficking of HIV proteins including Rev, Vpr and Matrix. We are especially excited by new methods we have developed allowing HIV to be visualized in living cells. Time-lapse analysis reveals the dynamics of the movement of individual virus particles. Further, the fluorescent tagging of complexes allows correlative electron microscopy to be used to image HIV derived complexes at the ultrastructural level. Intracellular complexes derived from a retrovirus have never been seen before! This system will allow detailed visualization of all steps of the HIV life cycle providing interesting new insights into HIV biology. Our initial studies hint that HIV is tracking down microtubules. Future studies will characterize the dynamics of virus binding and entry, the ultrastructure of cytoplasmic HIV complexes, and the dynamics of particle assembly. We will also study the affect of certain mutations, known to perturb virion infectivity, on particle behavior. For instance, mutation of the HIV Vif and Nef proteins decreases the infectivity of HIV by causing a block in the completion of reverse transcription. This observation suggests an important role for these viral proteins in a post entry step in the HIV life cycle. Visualization of the infection of cells with HIV mutated in either Nef or Vif will reveal the post entry step in the viral life cycle where these proteins act. The imaging techniques we have developed will revolutionize our understanding of the process of fusion, uncoating, nuclear localization, and other aspects of the life. 

 

Representative publications:

Henderson HI, Hope TJ. The temperature arrested intermediate of virus-cell fusion is a functional step in HIV infection. Virol J. 2006 May 25;3:36.

Wu X, Anderson JL, Campbell EM, Joseph AM, Hope TJ. Proteasome inhibitors uncouple rhesus TRIM5alpha restriction of HIV-1 reverse transcription and infection. Proc Natl Acad Sci U S A. 2006 May 9;103(19):7465-70.

Lee JH, Murphy SC, Belshan M, Sparks WO, Wannemuehler Y, Liu S, Hope TJ, Dobbs D, Carpenter S. Characterization of functional domains of equine infectious anemia virus Rev suggests a bipartite RNA-binding domain. J Virol. 2006 Apr;80(8):3844-52.

Perez O, Hope TJ. Cellular restriction factors affecting the early stages of HIV replication. Curr HIV/AIDS Rep. 2006 Feb;3(1):20-5.

Anderson JL, Hope TJ. Intracellular trafficking of retroviral vectors: obstacles and advances. Gene Ther. 2005 Dec;12(23):1667-78.

Campbell EM, Hope TJ. Gene therapy progress and prospects: viral trafficking during infection. Gene Ther. 2005 Sep;12(18):1353-9