Cytomegalovirus Encodes a Positive Regulator of Antigen Presentation

Abstract

Murine cytomegalovirus encodes three regulators of antigen presentation to antiviral CD8 T cells. According to current paradigms, all three regulators are committed to the inhibition of the presentation of antigenic peptides. Whereas m152/gp40 catalyzes the retention of peptide-loaded major histocompatibility complex (MHC) class I molecules in a cis-Golgi compartment, m06/gp48 binds stably to class I molecules and directs them into the cellular cargo-sorting pathway of lysosomal degradation. Regulator m04/gp34 also binds stably to class I molecules, but unlike m152 and m06, it does not downmodulate MHC class I cell surface expression. It has entered the literature as a direct inhibitor of T-cell recognition of the MHC-peptide complex at the cell surface. In this work, we have studied the presentation of antigenic viral peptides in cells infected with a comprehensive set of mutant viruses expressing the three regulators separately as well as in all possible combinations. The results redefine m04 as a positive regulator dedicated to the facilitation of antigen presentation. When expressed alone, it did not inhibit T-cell recognition, and when expressed in the presence of m152, it restored antigen presentation by antagonizing the inhibitory function of m152. Its intrinsic positive function, however, was antagonized and even slightly overcompensated for by the negative regulator m06. In an adoptive cell transfer model, the opposing forces of the three regulators were found to govern immune surveillance in the infected host. While negative regulators, also known as immunoevasins, are common, the existence of a positive regulator is without precedent and indicates an intriguing genetic potential of this virus to influence antigen presentation.

Epitope-specific in vivo protection against cytomegalovirus disease by CD8 T cells in the murine model of preemptive immunotherapy

“…Unlike the lysis of infected target cells, the ELISpot assay gives an estimate of the numbers of CD8 T cells that are capable of detecting presented peptide and that receive sufficient signaling required for IFN-c synthesis and secretion. In the ELISpot assay the inhibition by the concerted action of the three immunoevasins was less complete compared to the cytolysis assay in that a small proportion of CD8 T cells, most likely those with the highest functional avidity and TCR affinity, were sensitized by cells infected with mCMV-WT [7]. Nevertheless, there is consent between our group and Ann Hill's group (see the contribution by Doom and Hill in this issue of MMI) that the inhibition exerted by the concerted action of all three immunoevasins is stronger than for any of the immunoevasins expressed alone or for any combination of two of them.…”

“…2. As in our experience cultures of infected MEF always contain cells that are refractory to infection [7], a phenomenon that is independent of the multiplicity of infection (data not shown), it is essential to restrict the analysis of MHC class-I and of RAE-1 expression to infected cells. This was achieved by twocolor cytofluorometric analysis of the expression of the viral ER-resident glycoprotein m164/gp36.5 (S.A.O.-K., manuscript in preparation) and MHC class-I glycoprotein L d (Fig.…”

“…As shown for cells infected with wild-type mCMV (mCMV-WT), the concerted action of immunoevasins m04/gp34, m06/gp48, and m152/gp40 completely prevents lysis of infected cells by a large array of CTLs differing in MHCrestriction and in the epitopes recognized [6,7], although CTL specific for the D d -restricted m164-epitope represent a reported exception [8]. In principle, these findings also apply to the triggering of a second effector function of CD8 T cells, namely the secretion of interferon (IFN)-c as measured in an ELISpot assay.…”

“…(Left column) Two-color cytofluorometric analysis of the expression of the ER-resident viral glycoprotein m164/ gp36.5 (ordinate log fluorescence scale; Alexa Fluor 488), identifying infected cells, and the MHC class-I molecule L d (abscissa log fluorescence scale; PE) performed as described in greater detail previously [7]. Shown are dot plots with 2,000 cells displayed.…”

“…This was achieved by twocolor cytofluorometric analysis of the expression of the viral ER-resident glycoprotein m164/gp36.5 (S.A.O.-K., manuscript in preparation) and MHC class-I glycoprotein L d (Fig. 2, left column) [7] or of RAE-1 (Fig. 2, right column), respectively.…”