Structure and Dynamics of Membrane-associated ICP47, a Viral Inhibitor of the MHC I Antigen-processing Machinery

Abstract

To evade the host's immune response, herpes simplex virus employs the immediate early gene product ICP47 (IE12) to suppress antigen presentation to cytotoxic T-lymphocytes by inhibition of the ATP-binding cassette transporter associated with antigen processing (TAP). ICP47 is a membrane-associated protein adopting an ␣-helical conformation. Its active domain was mapped to residues 3-34 and shown to encode all functional properties of the full-length protein. The active domain of ICP47 was reconstituted into oriented phospholipid bilayers and studied by proton-decoupled 15 N and 2 H solid-state NMR spectroscopy. In phospholipid bilayers, the protein adopts a helix-loop-helix structure, where the average tilt angle of the helices relative to the membrane surface is ϳ15°(؎7°). The alignment of both structured domains exhibits a mosaic spread of ϳ10°. A flexible dynamic loop encompassing residues 17 and 18 separates the two helices. Refinement of the experimental data indicates that helix 1 inserts more deeply into the membrane. These novel insights into the structure of ICP47 represent an important step toward a molecular understanding of the immune evasion mechanism of herpes simplex virus and are instrumental for the design of new therapeutics.Survival of vertebrates is strongly dependent on the adaptive immune system, which confers protection against pathogens or cancer. On the cellular level, the major histocompatibility complex (MHC) 4 class I-dependent pathway of antigen processing allows for presentation of antigenic peptides at the cell surface and can trigger the elimination of virus-infected or malignantly transformed cells by cytotoxic T lymphocytes (1-3). The efficiency of antigen presentation depends on the transporter associated with antigen processing (TAP), a member of the ATPbinding cassette protein family that translocates peptides generated by proteasomal protein degradation into the endoplasmic reticulum for loading onto MHC class I molecules (4). This process requires a macromolecular peptide-loading complex of ϳ1 MDa comprising the transporter subunits TAP1 and TAP2, the MHC class I heavy chain, the non-covalently associated ␤ 2 -microglobulin, the adaptor protein tapasin, and several auxiliary factors (5-9).Herpes simplex virus type 1 is a highly abundant human pathogen that achieves lifelong persistence in the ganglia of the nervous system. Upon exogenous stimuli, it can be repeatedly reactivated and infect related mucosal tissues leading to clinical symptoms (10). To escape immune surveillance, herpes simplex virus compromises the host's cytotoxic T lymphocyte response via ICP47, an 88-amino-acid immediate early gene product (IE12) that blocks TAP function (11-15). In the absence of a functional TAP transporter (within 3 h of infection with herpes simplex virus), peptide loading onto MHC class I molecules is inhibited, and as a consequence, empty MHC I molecules are retained in the endoplasmic reticulum and ultimately directed to proteasomal degradation. By binding with nanomolar affinity t...

The Varicellovirus UL49.5 Protein Blocks the Transporter Associated with Antigen Processing (TAP) by Inhibiting Essential Conformational Transitions in the 6+6 Transmembrane TAP Core Complex

“…TAP appears to represent a favorite target for herpesvirus immune evasion strategies. The ␣-herpesviruses HSV-1 and HSV-2 encode the ICP47 protein, which blocks peptide binding to the cytosolic side of TAP (27)(28)(29)(30)(31). The US6 protein encoded by human CMV inhibits TAP by preventing ATP binding to TAP (32)(33)(34)(35)(36).…”

Preventing vaccinia virus class‐I epitopes presentation by HSV‐ICP47 enhances the immunogenicity of a TAP‐independent cancer vaccine epitope

“…These data are in agreement with published reports 29,30 showing that ICP47 binds to TAP and blocks peptide transport from the cytosol to ER thereby preventing HLA class I loading and translocation to the cell surface. 3,31 The maximum downregulating effect, related to ICP47 expression and HLA turnover is detectable after 36 h of infection with a nonreplicating virus. The extent of HLA ''downregulation'' could reach 80% of expression levels observed in noninfected cells.…”

“…2 In the absence of a functional TAP, peptide loading onto MHC class-I molecules is inhibited and, as a consequence, empty MHC class-I molecules are retained in the ER. 3 VV was first used for global smallpox eradication in the early 1980s, but still serves, at present time, as useful recombinant viral vector. Several unique features of VV including safety, wide host range, efficient infection and gene expression, stability, accommodation of large DNA sequence, cytoplasmic replication and ease of administration make it an excellent choice as a vaccine vehicle in vivo.…”

“…Our group has previously generated recombinant vaccinia virus expressing antigenic epitopes derived from three different melanoma differentiation antigens, Melan-A/MART-1 [27][28][29][30][31][32][33][34][35] , GP100 280-288 and tyrosinase [1][2][3][4][5][6][7][8][9] . The antigenic epitopes are encoded together with a polypeptide from adenovirus 19K derived leader sequence (MRYMILGLLALAAVCSA) driving the resulting recombinant products directly into the ER and thereby bypassing antigen processing steps.…”

Manipulation of MHC-I/TCR Interaction for Immune Therapy

“…The MHC-I of human cells expressed US6 dramatically reduced (11)(12)(13). To evade the host's immune response, herpes simplex virus employs the immediate early gene product ICP47 (IE12) to suppress antigen presentation to CTLs by inhibition of TAP (14). The ability of TAP to translocate antigenic peptides from the cytosol to the lumen of the ER for presentation on MHC-I in HIV-1-infected cells is abolished (15).…”