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
|
|
|
