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Alan Rein, Ph.D.

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HIV DRP Retroviral Replication Laboratory
Head, Retrovirus Assembly Section
Senior Investigator
NCI-Frederick
P.O. Box B
Building 535, Room 211
Frederick, MD 21702-1201
Phone:  
 301-846-1361
Fax:  
 301-846-6013
E-Mail:  
 rein@ncifcrf.gov
Link:
 Other Homepage

Biography

Dr. Alan Rein obtained his Ph.D. with Dr. Harry Rubin at the University of California at Berkeley and did postdoctoral research as an American Cancer Society Fellow under the direction of Dr. Sheldon Penman at the Massachusetts Institute of Technology. Dr. Rein has been associated with the National Cancer Institute (NCI) since 1976 and served as Head of the Retroviral Genetics Section in the ABL-Basic Research Program from 1984 to 1999. In 1999, he joined the HIV Drug Resistance Program as Head of the Retrovirus Assembly Section and was appointed to the NCI Senior Biomedical Research Service. Dr. Rein was an Organizer of the 1995 Cold Spring Harbor Laboratory Meeting on Retroviruses and the 2001 International Retroviral Nucleocapsid Symposium. He currently serves on the Editorial Boards of the Journal of Biological Chemistry, Virology, and Journal of Virology. He is also a member of the Center of Excellence in HIV/AIDS & Cancer Virology and the Chemistry and Structural Biology Faculty in the Center for Cancer Research, NCI. His research has dealt with a number of aspects of the biology and molecular biology of murine and human retroviruses, including virus assembly and maturation, viral envelope function, translational suppression, and pathogenesis.

Research

Mechanisms in Retroviral Replication and Pathogenesis

The goal of the research efforts in the Retrovirus Assembly Section is to extend our understanding of basic mechanisms in retroviral replication and pathogenesis. This understanding may lead to new methods of combatting retrovirus-induced disease, including AIDS.

There appear to be several different modes of interaction between retroviral proteins and nucleic acids, each with important functional consequences for viral replication. First, an exquisitely specific recognition by the Gag polyprotein (the structural protein of the virus particle) selects the viral RNA for packaging during virus assembly. This recognition involves zinc fingers in the protein. We are studying the mechanism by which the Gag protein recognizes and packages the genomic RNA of the virus during assembly in vivo. Our research strongly suggests that the recognition signal involves the three-dimensional structure formed by a dimer of genomic RNA molecules. We are studying the structure of the dimer linkage and its possible role in packaging of viral RNA. Our mutational studies also show that the zinc fingers have other crucial functions, in addition to their role in the recognition process. These additional functions are now under investigation.

We have found that virus particles lacking genomic RNA contain cellular mRNA molecules in place of the viral RNA. While the majority of mRNA species are packaged into the virions nonselectively, there are a few species that are significantly enriched. We are currently analyzing the mechanism of this enrichment; it is possible that this will help us understand how Gag normally recognizes the viral RNA during wild-type particle assembly.

We have analyzed the properties of recombinant HIV-1 Gag protein in some detail. When nucleic acid is added to Gag, the protein assembles into virus-like particles. These particles are significantly smaller than authentic viral particles, but particles of the correct size are formed if inositol phosphates, as well as nucleic acid, are provided to the protein. In the absence of nucleic acid, the protein is in monomer-dimer equilibrium, using the previously described interface within the C-terminal domain of its capsid moiety. When inositol hexakisphosphate is added, the protein switches from monomer-dimer to monomer-trimer equilibrium. We are currently investigating the mechanism of this effect, and trying to identify the Gag-Gag interfaces in the trimers. It is possible that the formation of trimers in solution is related to the size of particles assembled in the presence of nucleic acid.

The association constant for dimerization of Gag with a mutation at the dimer-interface within capsid is ~100-fold lower than that for wild-type Gag. Thus, this protein remains monomeric at relatively high concentrations. We have used the mutant protein as a model for monomeric Gag, and have subjected it to a series of hydrodynamic and biophysical analyses. We have also constructed molecular models of Gag from the existing structures of Gag cleavage products. Comparison of the predicted properties of these models with the experimentally determined hydrodynamic and other properties of the monomer indicates that the protein is folded over in solution, with its N and C termini relatively close together in three-dimensional space. Since Gag molecules are extended rods in immature virus particles, they must undergo a drastic conformational change during particle assembly.

Current work is devoted to the identification of Gag-Gag and Gag-nucleic acid interfaces used during particle assembly and to the understanding of the mechanism by which nucleic acid induces assembly. We are also attempting to detect the extension of Gag by direct biophysical measurements and are investigating the conformational changes accompanying Gag cleavage during virus maturation. Other experiments are dedicated to analysis of the specific incorporation of viral RNA during normal assembly, despite the ability of cellular mRNAs to support efficient assembly in the absence of viral RNA.

Our collaborators include Robert Fisher, Ph.D., Andrew Stephen, Ph.D., SAIC Frederick; Robert Shoemaker, NCI; Judith Levin, Ph.D., National Institute of Child Health and Human Development; Mark Yeager, M.D., Ph.D., and Kelly Dryden, Ph.D., The Scripps Research Institute; Karin Musier-Forsyth, Ph.D., University of Minnesota; Itay Rousso, Ph.D., The Weizmann Institute; Massimo Palmarini, Ph.D., University of Glasgow Veterinary School; Mark Williams, Ph.D., Northeastern University; Larry Kleiman, Ph.D., McGill AIDS Centre; Kevin Weeks, Ph.D., University of North Carolina; Daniele Fabris, Ph.D., University of Maryland-Baltimore County; Jacob Leibowitz, Ph.D., National Institutes of Health; Susan Krueger, Ph.D., and Joseph Curtis, Ph.D., National Institute of Standards and Technology; Mamuka Kvaratskhelia, Ph.D., Ohio State University; Shyam Biswal, Ph.D., Johns Hopkins University School of Public Health; and Thoru Pederson, Ph.D., University of Massachusetts.

This page was last updated on 2/9/2009.