Identification of host proteins differentially associated with HIV-1 RNA splice variants

HIV-1 generates unspliced (US), partially spliced (PS), and completely spliced (CS) classes of RNAs, each playing distinct roles in viral replication. Elucidating their host protein ‘interactomes’ is crucial to understanding virus-host interplay. Here, we present HyPR-MSSV for isolation of US, PS, and CS transcripts from a single population of infected CD4+ T-cells and mass spectrometric identification of their in vivo protein interactomes. Analysis revealed 212 proteins differentially associated with the unique RNA classes, including preferential association of regulators of RNA stability with US and PS transcripts and, unexpectedly, mitochondria-linked proteins with US transcripts. Remarkably, >80 of these factors screened by siRNA knockdown impacted HIV-1 gene expression. Fluorescence microscopy confirmed several to co-localize with HIV-1 US RNA and exhibit changes in abundance and/or localization over the course of infection. This study validates HyPR-MSSV for discovery of viral splice variant protein interactomes and provides an unprecedented resource of factors and pathways likely important to HIV-1 replication.

Download Full-text

Identifying HIV-1 RNA splice variant protein interactomes using HyPR-MSSV

10.1101/2020.09.15.298190 ◽

2020 ◽

Author(s):

Rachel A. Knoener ◽

Edward L. Evans ◽

Jordan T. Becker ◽

Mark Scalf ◽

Bayleigh E. Benner ◽

...

Keyword(s):

Gene Expression ◽

Splice Variant ◽

Rna Stability ◽

Mass Spectrometric ◽

Host Protein ◽

Single Population ◽

Mass Spectrometric Identification ◽

Variant Protein ◽

Hiv 1

ABSTRACTHIV-1 generates unspliced (US), partially spliced (PS), and completely spliced (CS) classes of RNAs; each playing distinct roles in viral replication. Elucidating their host protein “interactomes” is crucial to understanding virus-host interplay. Here, we present HyPR-MSSV for isolation of US, PS, and CS transcripts from a single population of infected CD4+ T-cells and mass spectrometric identification of their in vivo protein interactomes. Analysis revealed 212 proteins differentially associated with the unique RNA classes; including, preferential association of regulators of RNA stability with US- and PS-transcripts and, unexpectedly, mitochondria-linked proteins with US-transcripts. Remarkably, >80 of these factors screened by siRNA knock-down impacted HIV-1 gene expression. Fluorescence microscopy confirmed several to co-localize with HIV-1 US RNA and exhibit changes in abundance and/or localization over the course of infection. This study validates HyPR-MSSV for discovery of viral splice variant protein interactomes and provides an unprecedented resource of factors and pathways likely important to HIV-1 replication.

Download Full-text

Identification of a Novel Human Immunodeficiency Virus Type 1 Integrase Interactor, Gemin2, That Facilitates Efficient Viral cDNA Synthesis In Vivo

Journal of Virology ◽

10.1128/jvi.02471-05 ◽

2006 ◽

Vol 80 (12) ◽

pp. 5670-5677 ◽

Cited By ~ 42

Author(s):

Seiji Hamamoto ◽

Hironori Nishitsuji ◽

Teruo Amagasa ◽

Mari Kannagi ◽

Takao Masuda

Keyword(s):

Human Immunodeficiency Virus ◽

Human Immunodeficiency Virus Type ◽

Virus Type ◽

Host Protein ◽

Cdna Synthesis ◽

Immunodeficiency Virus ◽

Viral Cdna ◽

Hiv 1

ABSTRACT Retroviral integrase (IN) catalyzes the integration of viral cDNA into a host chromosome. Additional roles have been suggested for IN, including uncoating, reverse transcription, and nuclear import of the human immunodeficiency virus type 1 (HIV-1) genome. However, the underlying mechanism is largely unknown. Here, using a yeast two-hybrid system, we identified a survival motor neuron (SMN)-interacting protein 1 (Gemin2) that binds to HIV-1 IN. Reduction of Gemin2 with small interfering RNA duplexes (siGemin2) dramatically reduced HIV-1 infection in human primary monocyte-derived macrophages and also reduced viral cDNA synthesis. In contrast, siGemin2 did not affect HIV-1 expression from the integrated proviral DNA. Although Gemin2 was undetectable in cell-free viral particles, coimmunoprecipitation experiments using FLAG-tagged Gemin2 strongly suggested that Gemin2 interacts with the incoming viral genome through IN. Further experiments reducing SMN or other SMN-interacting proteins suggested that Gemin2 might act on HIV-1 either alone or with unknown proteins to facilitate efficient viral cDNA synthesis soon after infection. Thus, we provide the evidence for a novel host protein that binds to HIV-1 IN and facilitates viral cDNA synthesis and subsequent steps that precede integration in vivo.

Download Full-text

Derivation of simian tropic HIV-1 infectious clone reveals virus adaptation to a new host

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1818059116 ◽

2019 ◽

Vol 116 (21) ◽

pp. 10504-10509 ◽

Cited By ~ 4

Author(s):

Fabian Schmidt ◽

Brandon F. Keele ◽

Gregory Q. Del Prete ◽

Dennis Voronin ◽

Christine M. Fennessey ◽

...

Keyword(s):

Mononuclear Cells ◽

Infectious Clone ◽

Host Protein ◽

Peripheral Blood Mononuclear ◽

New Host ◽

Species Specific ◽

Hiv 1 ◽

Pigtail Macaques

To replicate in a new host, lentiviruses must adapt to exploit required host factors and evade species-specific antiviral proteins. Understanding how host protein variation drives lentivirus adaptation allowed us to expand the host range of HIV-1 to pigtail macaques. We have previously derived a viral swarm (in the blood of infected animals) that can cause AIDS in this new host. To further exploit this reagent, we generated infectious molecular clones (IMCs) from the viral swarm. We identified clones with high replicative capacity in pigtail peripheral blood mononuclear cells (PBMC) in vitro and used in vivo replication to select an individual IMC, named stHIV-A19 (for simian tropic HIV-1 clone A19), which recapitulated the phenotype obtained with the viral swarm. Adaptation of HIV-1 in macaques led to the acquisition of amino acid changes in viral proteins, such as capsid (CA), that are rarely seen in HIV-1–infected humans. Using stHIV-A19, we show that these CA changes confer a partial resistance to the host cell inhibitor Mx2 from pigtail macaques, but that complete resistance is associated with a fitness defect. Adaptation of HIV-1 to a new host will lead to a more accurate animal model and a better understanding of virus–host interactions.

Download Full-text

The Incorporation of Host Proteins into the External HIV-1 Envelope

Viruses ◽

10.3390/v11010085 ◽

2019 ◽

Vol 11 (1) ◽

pp. 85 ◽

Cited By ~ 10

Author(s):

Jonathan Burnie ◽

Christina Guzzo

Keyword(s):

Lipid Membrane ◽

Hiv Treatment ◽

Biologically Active ◽

Host Protein ◽

Host Proteins ◽

Novel Treatments ◽

Treatment And Prevention ◽

Viral Egress ◽

Protein Incorporation ◽

Hiv 1

The incorporation of biologically active host proteins into HIV-1 is a well-established phenomenon, particularly due to the budding mechanism of viral egress in which viruses acquire their external lipid membrane directly from the host cell. While this mechanism might seemingly imply that host protein incorporation is a passive uptake of all cellular antigens associated with the plasma membrane at the site of budding, this is not the case. Herein, we review the evidence indicating that host protein incorporation can be a selective and conserved process. We discuss how HIV-1 virions displaying host proteins on their surface can exhibit a myriad of altered phenotypes, with notable impacts on infectivity, homing, neutralization, and pathogenesis. This review describes the canonical and emerging methods to detect host protein incorporation, highlights the well-established host proteins that have been identified on HIV-1 virions, and reflects on the role of these incorporated proteins in viral pathogenesis and therapeutic targeting. Despite many advances in HIV treatment and prevention, there remains a global effort to develop increasingly effective anti-HIV therapies. Given the broad range of biologically active host proteins acquired on the surface of HIV-1, additional studies on the mechanisms and impacts of these incorporated host proteins may inform the development of novel treatments and vaccine designs.

Download Full-text

Cellular and Biochemical Mechanisms of the Retroviral Restriction Factor SAMHD1

ISRN Biochemistry ◽

10.1155/2013/728392 ◽

2013 ◽

Vol 2013 ◽

pp. 1-11 ◽

Cited By ~ 13

Author(s):

Li Wu

Keyword(s):

Host Protein ◽

Nucleic Acid Metabolism ◽

Target Cells ◽

Restriction Factors ◽

Host Proteins ◽

Retroviral Infection ◽

Deoxynucleoside Triphosphate ◽

Biochemical Mechanisms ◽

Hiv 1

Replication of HIV-1 and other retroviruses is dependent on numerous host proteins in the cells. Some of the host proteins, however, function as restriction factors to block retroviral infection of target cells. The host protein SAMHD1 has been identified as the first mammalian deoxynucleoside triphosphate triphosphohydrolase (dNTPase), which blocks the infection of HIV-1 and other retroviruses in non-cycling immune cells. SAMHD1 protein is highly expressed in human myeloid-lineage cells and CD4+ T-lymphocytes, but its retroviral restriction function is only observed in noncycling cells. Recent studies have revealed biochemical mechanisms of SAMHD1-mediated retroviral restriction. In this review, the latest progress on SAMHD1 research is summarized and the mechanisms by which SAMHD1 mediates retroviral restriction are analyzed. Although the physiological function of SAMHD1 is largely unknown, this review provides perspectives about the role of endogenous SAMHD1 protein in maintaining normal cellular function, such as nucleic acid metabolism and the proliferation of cells.

Download Full-text

Cellular Compartments of Human Immunodeficiency Virus Type 1 Replication In Vivo: Determination by Presence of Virion-Associated Host Proteins and Impact of Opportunistic Infection

Journal of Virology ◽

10.1128/jvi.74.1.139-145.2000 ◽

2000 ◽

Vol 74 (1) ◽

pp. 139-145 ◽

Cited By ~ 49

Author(s):

Stephen D. Lawn ◽

Beverly D. Roberts ◽

George E. Griffin ◽

Thomas M. Folks ◽

Salvatore T. Butera

Keyword(s):

Human Immunodeficiency Virus ◽

Opportunistic Infection ◽

Human Immunodeficiency Virus Type ◽

Host Proteins ◽

Immunodeficiency Virus ◽

Cellular Compartments ◽

Hiv 1

ABSTRACT Antigens derived from host cells are detectable in the envelope of human immunodeficiency virus type 1 (HIV-1) and result in a distinctive viral phenotype reflecting that of the host cell. An immunomagnetic capture assay targeting discriminatory host proteins was developed to differentiate between HIV-1 derived from macrophages and lymphocytes. HIV-1 propagated in macrophages or lymphocytes in vitro was selectively captured by monoclonal antibodies directed against the virally incorporated cell-type-specific host markers CD36 (macrophages) and CD26 (lymphocytes). Furthermore, by targeting these markers, virus of defined cellular origin was selectively captured from a mixed pool of in vitro-propagated viruses. This technique was further refined in order to determine the impact of opportunistic infection on HIV-1 expression from these cellular compartments in vivo. Analysis of cell-free virus purified from plasma of patients with HIV-1 infection suggested that in those with an opportunistic infection, viral replication occurred in activated lymphocytes. Interestingly, there was also significant replication in activated macrophages in those patients with untreated pulmonary tuberculosis. Thus, in addition to lymphocytes, the macrophage cellular pool may serve as an important source of cell-free HIV-1 in patients with opportunistic infections that lead to marked macrophage activation. This novel viral capture technique may allow researchers to address a wide range of important questions regarding virus-host dynamics.

Download Full-text

Proteomic interrogation of the pathogen-host interface in cholera

10.1101/2021.01.05.425471 ◽

2021 ◽

Author(s):

Abdelrahim Zoued ◽

Hailong Zhang ◽

Ting Zhang ◽

Rachel T. Giorgio ◽

Carole J. Kuehl ◽

...

Keyword(s):

Cell Surface ◽

Vibrio Cholerae ◽

Intestinal Microbiota ◽

Rabbit Model ◽

Host Protein ◽

Host Proteins ◽

Host Interactions ◽

Proteomic Investigation ◽

Critical Site

SummaryThe microbial cell surface is a critical site of microbe-host interactions that often control infection outcomes. Here, using the infant rabbit model of cholera, which provides an abundant source of in vivo Vibrio cholerae cells and diarrheal fluid, we investigated the proteomic composition of this interface. Bulk diarrheal fluid proteomes revealed that cholera toxin accounts for the vast majority of the host proteins present during infection. We developed a surface biotinylation protocol to purify and quantify both bacterial and host proteins present on the surface of diarrheal fluid-derived V. cholerae. We found that SP-D, a toxin-dependent host protein that directly binds the V. cholerae surface, is a novel intestinal defense factor. Other V. cholerae-bound host proteins also bound distinct taxa of the murine intestinal microbiota. Proteomic investigation of the microbial surface-host interface should be a valuable tool for probing microbe-host interactions and their influence on homeostasis and infection.

Download Full-text

Presence of Host ICAM-1 in Laboratory and Clinical Strains of Human Immunodeficiency Virus Type 1 Increases Virus Infectivity and CD4+-T-Cell Depletion in Human Lymphoid Tissue, a Major Site of Replication In Vivo

Journal of Virology ◽

10.1128/jvi.76.3.1004-1014.2002 ◽

2002 ◽

Vol 76 (3) ◽

pp. 1004-1014 ◽

Cited By ~ 129

Author(s):

Salim Bounou ◽

Jacques E. Leclerc ◽

Michel J. Tremblay

Keyword(s):

Human Immunodeficiency Virus ◽

Human Immunodeficiency Virus Type ◽

Lymphoid Tissue ◽

Host Proteins ◽

Major Site ◽

Immunodeficiency Virus ◽

Human Lymphoid Tissue ◽

Hiv 1

ABSTRACT Human immunodeficiency virus type 1 (HIV-1) incorporates several host proteins. Earlier studies have indicated that such foreign constituents can modulate the virus life cycle, although the potential roles that these proteins might play in the viral pathology in vivo remain unclear. In an attempt to shed light on this issue, we first exposed explants of human lymphoid tissue to isogenic viruses except for the presence or absence of host-derived ICAM-1. Incorporation of ICAM-1 alone increased HIV-1 infectivity for human tonsillar tissue cultured ex vivo. This observation was made for viruses bearing distinct coreceptor utilization profiles. Conversion of LFA-1 to a high-affinity-high-avidity state for ICAM-1 further augmented the susceptibility of human tonsillar histocultures to infection by ICAM-1-bearing virions. A more massive depletion of CD4+ T lymphocytes was seen with X4 ICAM-1/POS viruses than with isogenic ICAM-1/NEG virions. Exposure of X4 and R5 primary isolates of HIV-1 to a blocking anti-ICAM-1 antibody resulted in a decrease of virus infection. Finally, X4 and R5 virions derived from a natural human lymphoid tissue microenvironment incorporated high levels of ICAM-1. Altogether, these results indicate that the incorporation of host ICAM-1 can significantly modulate the biology of HIV-1 in a cellular milieu recognized as the major site of replication in vivo and suggest that host proteins found in HIV-1 particles may participate in the pathogenesis of this disease.

Download Full-text