The Active Site of ICP47, a Herpes Simplex Virus–encoded Inhibitor of the Major Histocompatibility Complex (MHC)-encoded Peptide Transporter Associated with Antigen Processing (TAP), Maps to the NH2-terminal 35 Residues

Abstract

The herpes simplex virus (HSV) immediate early protein ICP47 inhibits the transporter associated with antigen processing (TAP)-dependent peptide translocation. As a consequence, empty major histocompatibility complex (MHC) class I molecules are retained in the endoplasmic reticulum and recognition of HSV-infected cells by cytotoxic T lymphocytes is abolished. We chemically synthesized full-length ICP47 (sICP47) and show that sICP47 inhibits TAP-dependent peptide translocation in human cells. Its biological activity is indistinguishable from that of recombinant ICP47 (rICP47). By using synthetic peptides, we mapped the core sequence of ICP47 minimally required for TAP inhibition to residues 2–35. This segment is located within the region of the molecule conserved between ICP47 from HSV-1 and HSV-2. Through alanine scanning substitution we identified three segments within this region that are critical for the ability to inhibit TAP function. The interaction of ICP47 with TAP is unlikely to mimic precisely that of the transported peptides, as deduced from differential labeling of the TAP1 and TAP2 subunits using sICP47 fragments with chemical cross-linkers.

Varicella-Zoster Virus Retains Major Histocompatibility Complex Class I Proteins in the Golgi Compartment of Infected Cells

“…These observations indicate that the pathway by which VZV infection reduces MHC I cell surface expression is novel, differing from the effects of the other human herpesviruses, HSV and HCMV (24). HSV has been shown to induce the retention of MHC I molecules in the ER through a specific interaction of a viral protein (ICP47) with the TAP complex, resulting in the prevention of TAP-mediated antigen transport (4,19,21,25,48,50). In contrast, HCMV has been demonstrated to use several viral immunomodulatory proteins that affect various stages of the MHC I assembly pathway, including prevention of TAPmediated antigen transport, retention of MHC I molecules in the ER, and dislocation of retained complexes to the cytosol for degradation (2,3,22,23,29,30,36,49).…”

“…In this respect, other herpesviruses, including HSV, HCMV, and MCMV, have been shown to encode immunomodulatory proteins that directly associate with components of the MHC I biosynthesis pathway (2,19,21,25,29,33,36,45,48,49,50). Immunoprecipitation of radiolabeled MHC I molecules in VZV-and mock-infected cells did not reveal binding of a viral protein, but a modified, ϳ40-kDa MHC I molecule was detected consistently in VZV-infected cells, which may reflect accelerated degradation of MHC I molecules retained in the Golgi complex.…”

Equine Herpesvirus Type 4 UL56 and UL49.5 Proteins Downregulate Cell Surface Major Histocompatibility Complex Class I Expression Independently of Each Other

“…Several other herpesviruses have evolved mechanisms to interfere with TAP function through different strategies exerted by unique gene products. Herpes simplex virus 1 (HSV-1) and HSV-2 encode the ICP47 protein that acts as a high-affinity competitor for peptide binding to TAP and thus prevents peptide translocation (3,4,25,29,38,63,79). The bovine herpesvirus 1 (BHV-1) UL49.5 protein induces a conformational arrest of the peptide transporter and degradation of TAP by the proteasome (47,82).…”

Varicelloviruses avoid T cell recognition by UL49.5-mediated inactivation of the transporter associated with antigen processing

“…Three different proteins specifically inhibiting peptide translocation by TAP have been described within the herpesvirus family. Herpes simplex virus 1 (HSV-1) and HSV-2 encode a soluble cytoplasmic protein, ICP47, that associates with the peptide binding site formed by the C-terminal cytosolic domains of TAP1 and TAP2, thereby acting as a high-affinity competitor for peptide binding (14)(15)(16)(17)(18)(19). Human cytomegalovirus (HCMV) uses an ERresident type I membrane protein, US6, to inhibit TAP (20)(21)(22)(23)(24)(25).…”

“…The HSV-encoded ICP47 blocks the association of peptides with the cytosolic peptide-binding domain of TAP (14)(15)(16)(17)(18)(19). To test whether UL49.5 acts in a similar fashion, peptide binding to TAP was evaluated by using microsomes isolated from UL49.5-expressing and control cells (Fig.…”