The goal of this proposal is to elucidate the role of specific host proteins that interact with HIV-1 Gag in virus replication. We have identified the host proteins kindlin-3 and pacsin2 as binding partners for the NC and p6 domains of HIV-1 Gag, respectively. Knockdown and reconstitution experiments show that both proteins are critical for the spreading of HIV-1 among T cells. Based on our preliminary data, our hypothesis is that kindlin-3 and pacsin2 are involved in the cell-to- cell transmission of HIV-1. Kindlin-3 is required for the function of integrins such as LFA-1, which has been implicated in HIV transmission. However, our data demonstrate a striking requirement for kindlin-3 for HIV-1 replication even in T lymphoid cells lacking LFA-1. Our working model is that HIV-1 Gag mimics the kindlin-recruiting activity of integrins to induce donor cell polarization towards the virologial synapse and thereby promote virus transfer. We propose to investigate the mechanism by which kindlin-3 associates with HIV-1 virions, which step of the viral life cycle is promoted by kindlin-3, whether downstream partners of kindlin-3 are involved in HIV-1 replication, and whether the kindlin-3-dependent activity of integrins other than LFA-1 is important for HIV-1 replication. Pacsin2 can generate membrane curvature and nucleate filopodia formation. HIV-1 frequently buds from filopodia, and viral filopodia capped by Gag have been implicated in HIV-1 cell-cell transmission. Thus, pacsin2 may be important for HIV-1 transmission because it is involved in viral filopodia formation. We propose to investigate the role of Gag ubiquitination in the recruitment of pacsin2 into viral particles, the precise role of pacsin2 in HIV-1 replication, he role of the actin remodeling activity of pacsin2 in HIV-1 replication, and whether pacsin2 interaction partners on recycling endosomes are involved in HIV-1 replication. The transfer of HIV-1 from cell to cell is thought to be the predominant mode of transmission, but the molecular basis is poorly understood. An understanding of the roles of kindlin-3 and pacsin2 in HIV-1 replication may provide novel insights into this important mode of transmission.

Public Health Relevance

Our application proposes an in-depth investigation of the roles of specific host proteins that are taken up into virions in the spreading of HIV-1. The proposed studies are relevant to public health, because they may yield important new insights into how HIV-1 spreads from cell to cell, and may identify vulnerabilities that can be exploited to inhibit this important mode of transmission.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
2R01AI029873-26
Application #
8732327
Study Section
AIDS Molecular and Cellular Biology Study Section (AMCB)
Program Officer
Sharma, Opendra K
Project Start
1990-03-01
Project End
2019-05-31
Budget Start
2014-06-01
Budget End
2015-05-31
Support Year
26
Fiscal Year
2014
Total Cost
Indirect Cost
Name
University of Massachusetts Medical School Worcester
Department
Genetics
Type
Schools of Medicine
DUNS #
City
Worcester
State
MA
Country
United States
Zip Code
01655
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Martinelli, Nicolas; Hartlieb, Bettina; Usami, Yoshiko et al. (2012) CC2D1A is a regulator of ESCRT-III CHMP4B. J Mol Biol 419:75-88
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Usami, Yoshiko; Popov, Sergei; Popova, Elena et al. (2009) The ESCRT pathway and HIV-1 budding. Biochem Soc Trans 37:181-4
Pizzato, Massimo; Popova, Elena; Gottlinger, Heinrich G (2008) Nef can enhance the infectivity of receptor-pseudotyped human immunodeficiency virus type 1 particles. J Virol 82:10811-9
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Calistri, Arianna; Sette, Paola; Salata, Cristiano et al. (2007) Intracellular trafficking and maturation of herpes simplex virus type 1 gB and virus egress require functional biogenesis of multivesicular bodies. J Virol 81:11468-78
Usami, Yoshiko; Popov, Sergei; Gottlinger, Heinrich G (2007) Potent rescue of human immunodeficiency virus type 1 late domain mutants by ALIX/AIP1 depends on its CHMP4 binding site. J Virol 81:6614-22
Pizzato, Massimo; Helander, Anna; Popova, Elena et al. (2007) Dynamin 2 is required for the enhancement of HIV-1 infectivity by Nef. Proc Natl Acad Sci U S A 104:6812-7

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