The HIV-1 Gag protein orchestrates co-factor packaging, assembly and release of the immature virion. As the virion emerges, Gag is then processed by the viral protease to produce the mature CA protein, which reassembles into the conical capsid that organizes the viral core for uncoating and replication in a new host cell. Research during the previous funding period has revealed the structure of the viral capsid and begun to define its roles in viral replication. We now propose to expand our studies of particle assembly to include: 1) structural studies of CA proteins aimed at defining the molecular details of CA-ligand and CA-CA interactions, 2) structural studies of larger Gag protein fragments aimed at understanding how Gag organizes the immature capsid and changes conformation during viral assembly, and 3) development of assays that will be used to study viral capsid functions in vitro. Preliminary results include the growth of high quality crystals of a protein spanning the first 278 residues of HIV-1 Gag, the demonstration that the amino- terminal end of CA can adopt at least three distinct conformations, and the implementation of procedures for viral core preparation and endogenous reverse transcription. We will also study how the assembling Gag lattice interacts with cytoskeletal proteins during viral assembly. Although roles for cytoskeletal proteins in HIV-1 assembly have not been characterized previously, we have identified a series of protein-protein interactions that link HIV-1 Gag and its cellular binding partner, TSG101, to proteins that normally control actomyosin organization and activation. Both overexpression and inhibition of these regulatory proteins block virus assembly and release without altering Gag expression levels, indicating that they play a direct role in particle assembly, possibly by helping to remodel the cortical actin cytoskeleton and/or to drive particle envelopment. Proposed studies will define this protein interaction network in detail, and define its role in HIV-1 particle assembly.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Method to Extend Research in Time (MERIT) Award (R37)
Project #
5R37AI045405-14
Application #
8284351
Study Section
Special Emphasis Panel (NSS)
Program Officer
Salzwedel, Karl D
Project Start
1999-06-01
Project End
2014-05-31
Budget Start
2012-06-01
Budget End
2013-05-31
Support Year
14
Fiscal Year
2012
Total Cost
$295,010
Indirect Cost
$98,990
Name
University of Utah
Department
Biochemistry
Type
Schools of Medicine
DUNS #
009095365
City
Salt Lake City
State
UT
Country
United States
Zip Code
84112
Lamorte, Louie; Titolo, Steve; Lemke, Christopher T et al. (2013) Discovery of novel small-molecule HIV-1 replication inhibitors that stabilize capsid complexes. Antimicrob Agents Chemother 57:4622-31
Sundquist, Wesley I; Kräusslich, Hans-Georg (2012) HIV-1 assembly, budding, and maturation. Cold Spring Harb Perspect Med 2:a006924
Ganser-Pornillos, Barbie K; Chandrasekaran, Viswanathan; Pornillos, Owen et al. (2011) Hexagonal assembly of a restricting TRIM5alpha protein. Proc Natl Acad Sci U S A 108:534-9
Ganser-Pornillos, Barbie K; Yeager, Mark; Sundquist, Wesley I (2008) The structural biology of HIV assembly. Curr Opin Struct Biol 18:203-17
Langelier, Charles R; Sandrin, Virginie; Eckert, Debra M et al. (2008) Biochemical characterization of a recombinant TRIM5alpha protein that restricts human immunodeficiency virus type 1 replication. J Virol 82:11682-94
Kelly, Brian N; Kyere, Sampson; Kinde, Isaac et al. (2007) Structure of the antiviral assembly inhibitor CAP-1 complex with the HIV-1 CA protein. J Mol Biol 373:355-66
Morita, Eiji; Sandrin, Virginie; Chung, Hyo-Young et al. (2007) Human ESCRT and ALIX proteins interact with proteins of the midbody and function in cytokinesis. EMBO J 26:4215-27
Javanbakht, Hassan; Yuan, Wen; Yeung, Darwin F et al. (2006) Characterization of TRIM5alpha trimerization and its contribution to human immunodeficiency virus capsid binding. Virology 353:234-46
Diaz-Griffero, Felipe; Vandegraaff, Nick; Li, Yuan et al. (2006) Requirements for capsid-binding and an effector function in TRIMCyp-mediated restriction of HIV-1. Virology 351:404-19
Stremlau, Matthew; Perron, Michel; Lee, Mark et al. (2006) Specific recognition and accelerated uncoating of retroviral capsids by the TRIM5alpha restriction factor. Proc Natl Acad Sci U S A 103:5514-9

Showing the most recent 10 out of 14 publications