Chemical Biology of HIV-1 Nef

HIV-1 Nef 的化学生物学

基本信息

项目摘要

Summary. Existing antiretroviral drugs do not clear HIV-1 latent reservoirs, underscoring the urgent need for new therapeutic strategies. The HIV-1 Nef accessory factor is an attractive target for drug development because of its critical roles in the HIV-1 life cycle and immune system escape. Our group has discovered novel small molecules that bind directly to Nef and block many of its functions, including enhancement of viral infectivity and replication in donor PBMCs. Importantly, our Nef inhibitors rescue cell-surface MHC-I expression in latently infected, patient-derived CD4+ T-cells, enabling recognition and killing by autologous CTLs. Thus, Nef inhibitors represent an innovative approach to antiretroviral therapy that may provide a path to eradication of viral reservoirs. Our most promising class of inhibitors (hydroxypyrazoles) bind tightly to their Nef protein target in vitro and are active against multiple Nef functions in cell-based systems without cytotoxicity. Experiments proposed here will leverage these compounds as chemical probes to shed new light on Nef functions while unraveling their mechanism of action with the following Specific Aims: Aim 1. Map the binding site for hydroxy- pyrazole Nef inhibitors by X-ray crystallography. Preliminary and published data strongly suggest that hydroxy- pyrazole Nef inhibitors, which disrupt multiple Nef functions, may perturb the structure of functional Nef-effector complexes. X-ray crystallography of inhibitors with Nef alone and in complexes with host cell effector proteins will be used to test this idea and identify inhibitor binding sites. Aim 2. Identification of Nef residues essential for inhibitor action through in vitro selection. Using PCR-based saturation mutagenesis, we have replaced every codon in the Nef core region with each of the 64 nucleotide triplets in the context of HIV-1. CD4 T cells will be infected with the Nef mutant viral ‘library’ in the presence or absence of Nef inhibitors, and viral supernatants analyzed by deep sequencing to identify mutations enriched by inhibitor treatment. This method has the potential to identify Nef regions that allosterically influence inhibitor action in addition to residues directly involved in ligand binding. Aim 3. Explore the mechanisms by which Nef inhibitors suppress HIV-1 infectivity. Hydroxypyrazole Nef inhibitors reduce HIV-1 infectivity in TZM-bl reporter cells to the same extent as Nef-deleted viruses. This Aim will explore the whether Nef inhibitors restore virion incorporation of SERINC proteins and Ezrin, two host cell restriction factors linked to Nef. We will also pursue Nef inhibitor effects on overall HIV-1 protein composition by whole-virus proteomics, which has the potential to identify host cell factors that are uniquely incorporated (or excluded) by Nef inhibition. Aim 4. Investigate the mechanism of Nef inhibitor action on MHC-I downregulation. This Aim will explore the effect of Nef inhibitors on crystal structures of Nef in complexes with the MHC-I cyto- plasmic tail and the AP-1 µ1 subunit, interactions essential for immune escape. Inhibitor effects on Nef interac- tions with MHC-I and AP-1 will also be explored in cells using bimolecular fluorescence complementation (BiFC). These studies will clarify the mechanisms by which Nef inhibitors restore CTL responses to HIV infection.
总结。现有的抗逆转录病毒药物不能清除HIV-1潜伏库,因此迫切需要

项目成果

期刊论文数量(0)
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Thomas E. Smithgall其他文献

ID: 36: MCPIP1/Regnase-1 is a negative feedback inhibitor regulating IL-17 signaling and inflammation
  • DOI:
    10.1016/j.cyto.2015.08.066
  • 发表时间:
    2015-11-01
  • 期刊:
  • 影响因子:
  • 作者:
    Abhishek Garg;Nilesh Amatya;Kong Chen;J. Agustin Cruz;Prerna Grover;Natasha Whibley;Heather R. Conti;Gerard Hernandez Mir;Tatiana Sirakova;Erin C. Childs;Thomas E. Smithgall;Partha S. Biswas;Jay K. Kolls;Mandy J. McGeachy;Pappachan E. Kolattukudy;Sarah L. Gaffen
  • 通讯作者:
    Sarah L. Gaffen
Allosteric restriction enhances sensitivity of the AML-associated Src-family kinase Fgr to ATP-site inhibitors
  • DOI:
    10.1016/j.bpj.2023.11.2076
  • 发表时间:
    2024-02-08
  • 期刊:
  • 影响因子:
  • 作者:
    Giancarlo Gonzalez-Areizaga;John J. Alvarado;Du Shoucheng;Thomas E. Smithgall
  • 通讯作者:
    Thomas E. Smithgall
Mutagenesis of the HIV-1 Nef homodimerization interface suppresses multiple functions without altering the core fold
  • DOI:
    10.1016/j.bpj.2023.11.1237
  • 发表时间:
    2024-02-08
  • 期刊:
  • 影响因子:
  • 作者:
    Catherine E. Thomas;Frank Heinrich;John J. Alvarado;Thomas E. Smithgall
  • 通讯作者:
    Thomas E. Smithgall
Membrane Bound Structure of the HIV-1 Accessory Protein Nef
  • DOI:
    10.1016/j.bpj.2017.11.226
  • 发表时间:
    2018-02-02
  • 期刊:
  • 影响因子:
  • 作者:
    Rebecca Eells;Kindra Whitlatch;Bradley Treece;Frank Heinrich;John Jeff Alvarado;Thomas E. Smithgall;Mathias Lösche
  • 通讯作者:
    Mathias Lösche
Membrane Binding of HIV-1 Accessory Protein Nef on Sparsely-Tethered Bilayer Lipid Membranes: An Spr Study
  • DOI:
    10.1016/j.bpj.2018.11.350
  • 发表时间:
    2019-02-15
  • 期刊:
  • 影响因子:
  • 作者:
    Christopher Kervick;Manish Aryal;Frank Heinrich;Thomas E. Smithgall;Mathias Lösche
  • 通讯作者:
    Mathias Lösche

Thomas E. Smithgall的其他文献

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{{ truncateString('Thomas E. Smithgall', 18)}}的其他基金

Precision Targeting of Myeloid Src-family Kinases in Acute Myelogenous Leukemia
急性髓系白血病中髓系 Src 家族激酶的精确靶向
  • 批准号:
    10308327
  • 财政年份:
    2021
  • 资助金额:
    $ 61.93万
  • 项目类别:
Chemical Biology of HIV-1 Nef
HIV-1 Nef 的化学生物学
  • 批准号:
    10684695
  • 财政年份:
    2020
  • 资助金额:
    $ 61.93万
  • 项目类别:
PROTACS Against Nef as a Functional Cure for HIV Infection
PROTACS 针对 Nef 作为 HIV 感染的功能性治疗
  • 批准号:
    10200007
  • 财政年份:
    2020
  • 资助金额:
    $ 61.93万
  • 项目类别:
Chemical Biology of HIV-1 Nef
HIV-1 Nef 的化学生物学
  • 批准号:
    10471355
  • 财政年份:
    2020
  • 资助金额:
    $ 61.93万
  • 项目类别:
PROTACS Against Nef as a Functional Cure for HIV Infection
PROTACS 针对 Nef 作为 HIV 感染的功能性治疗
  • 批准号:
    10079715
  • 财政年份:
    2020
  • 资助金额:
    $ 61.93万
  • 项目类别:
Precision Targeting of Myeloid Src-family Kinases in Acute Myelogenous Leukemia
急性髓系白血病中髓系 Src 家族激酶的精确靶向
  • 批准号:
    10687861
  • 财政年份:
    2019
  • 资助金额:
    $ 61.93万
  • 项目类别:
Precision Targeting of Myeloid Src-family Kinases in Acute Myelogenous Leukemia
急性髓系白血病中髓系 Src 家族激酶的精确靶向
  • 批准号:
    10388497
  • 财政年份:
    2019
  • 资助金额:
    $ 61.93万
  • 项目类别:
Precision Targeting of Myeloid Src-family Kinases in Acute Myelogenous Leukemia
急性髓系白血病中髓系 Src 家族激酶的精确靶向
  • 批准号:
    9814793
  • 财政年份:
    2019
  • 资助金额:
    $ 61.93万
  • 项目类别:
Precision Targeting of Myeloid Src-family Kinases in Acute Myelogenous Leukemia
急性髓系白血病中髓系 Src 家族激酶的精确靶向
  • 批准号:
    10740923
  • 财政年份:
    2019
  • 资助金额:
    $ 61.93万
  • 项目类别:
Precision Targeting of Myeloid Src-family Kinases in Acute Myelogenous Leukemia
急性髓系白血病中髓系 Src 家族激酶的精确靶向
  • 批准号:
    10524124
  • 财政年份:
    2019
  • 资助金额:
    $ 61.93万
  • 项目类别:
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