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Identification and Functional Characterization of a Novel Fc Gamma-Binding Glycoprotein in Rhesus Cytomegalovirus.

ABSTRACT: Receptors recognizing the Fc part of immunoglobulin G (Fc?Rs) are key determinants in antibody-mediated immune responses. Members of the Herpesviridae interfere with this immune regulatory network by expressing viral Fc?Rs (vFc?Rs). Human cytomegalovirus (HCMV) encodes four distinct vFc?Rs that differ with respect to their IgG subtype specificity and their impact on antibody-mediated immune function in vitro The impact of vFc?Rs on HCMV pathogenesis and immunomodulation in vivo is not known. The closest evolutionary animal model of HCMV is rhesus CMV (RhCMV) infection of rhesus macaques. To enable the characterization of vFc?R function in this model, we studied IgG binding by RhCMV. We show that lysates of RhCMV-infected cells contain an IgG-binding protein of 30?kDa encoded by the gene Rh05 that is a predicted type I glycoprotein belonging to the RL11 gene family. Upon deletion of Rh05, IgG-Fc binding by RhCMV strain 68-1 is lost, whereas ectopic expression of Rh05 results in IgG binding to transfected cells consistent with Rh05 being a vFc?R. Using a set of reporter cell lines stably expressing human and rhesus Fc?Rs, we further demonstrate that Rh05 antagonizes host Fc?R activation. Compared to Rh05-intact RhCMV, RhCMV?Rh05 showed an increased activation of host Fc?R upon exposure of infected cells to IgG from RhCMV-seropositive animals, suggesting that Rh05 protects infected cells from opsonization and IgG-dependent activation of host Fc?Rs. However, antagonizing host Fc?R activation by Rh05 was not required for the establishment and maintenance of infection of RhCMV, even in a seropositive host, as shown by the induction of T cell responses to heterologous antigens expressed by RhCMV lacking the gene region encoding Rh05. In contrast to viral evasion of natural killer cells or T cell recognition, the evasion of antibody-mediated effects does not seem to be absolutely required for infection or reinfection. The identification of the first vFc?R that efficiently antagonizes host Fc?R activation in the RhCMV genome will thus permit more detailed studies of this immunomodulatory mechanism in promoting viral dissemination in the presence of natural or vaccine-induced humoral immunity.IMPORTANCE Rhesus cytomegalovirus (RhCMV) offers a unique model for studying human cytomegalovirus (HCMV) pathogenesis and vaccine development. RhCMV infection of nonhuman primates greatly broadened the understanding of mechanisms by which CMVs evade or reprogram T cell and natural killer cell responses in vivo However, the role of humoral immunity and viral modulation of anti-CMV antibodies has not been studied in this model. There is evidence from in vitro studies that HCMVs can evade humoral immunity. By gene mapping and with the help of a novel cell-based reporter assay system we characterized the first RhCMV encoded IgG-Fc? binding glycoprotein as a potent antagonist of rhesus Fc?R activation. We further demonstrate that, unlike evasion of T cell immunity, this viral Fc? receptor is not required to overcome anti-CMV immunity to establish secondary infections. These findings enable more detailed studies of the in vivo consequences of CMV evasion from IgG responses in nonhuman primate models.

SUBMITTER: Kolb P

PROVIDER: S-EPMC6364020 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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Identification and Functional Characterization of a Novel Fc Gamma-Binding Glycoprotein in Rhesus Cytomegalovirus.

Kolb Philipp P Sijmons Steven S McArdle Matthew R MR Taher Husam H Womack Jennie J Hughes Colette C Ventura Abigail A Jarvis Michael A MA Stahl-Hennig Christiane C Hansen Scott S Picker Louis J LJ Malouli Daniel D Hengel Hartmut H Früh Klaus K

Journal of virology 20190205 4

Receptors recognizing the Fc part of immunoglobulin G (FcγRs) are key determinants in antibody-mediated immune responses. Members of the Herpesviridae interfere with this immune regulatory network by expressing viral FcγRs (vFcγRs). Human cytomegalovirus (HCMV) encodes four distinct vFcγRs that differ with respect to their IgG subtype specificity and their impact on antibody-mediated immune function in vitro The impact of vFcγRs on HCMV pathogenesis and immunomodulation in vivo< ...[more]

PMID: 30487278

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Project description:Fc gamma receptors (Fc?R) bind the Fc region of antibodies and therefore play a prominent role in antibody-dependent cell-based immune responses such as ADCC, CDC and ADCP. The immune effector cell activity is directly linked to a productive molecular engagement of Fc?Rs where both the protein and glycan moiety of antibody and receptor can affect the interaction and in the present study we focus on the role of the Fc?R glycans in this interaction. We provide a complete description of the glycan composition of Chinese hamster ovary (CHO) expressed human Fc? receptors RI (CD64), RIIaArg131/His131 (CD32a), RIIb (CD32b) and RIIIaPhe158/Val158 (CD16a) and analyze the role of the glycans in the binding mechanism with IgG. The interactions of the monoclonal antibody rituximab with each Fc?R were characterized and we discuss the CHO-Fc?RIIIaPhe158/Val158 and CHO-Fc?RI interactions and compare them to the equivalent interactions with human (HEK293) and murine (NS0) produced receptors. Our results reveal clear differences in the binding profiles of rituximab, which we attribute in each case to the differences in host cell-dependent Fc?R glycosylation. The glycan profiles of CHO expressed Fc?RI and Fc?RIIIaPhe158/Val158 were compared with the glycan profiles of the receptors expressed in NS0 and HEK293 cells and we show that the glycan type and abundance differs significantly between the receptors and that these glycan differences lead to the observed differences in the respective Fc?R binding patterns with rituximab. Oligomannose structures are prevalent on Fc?RI from each source and likely contribute to the high affinity rituximab interaction through a stabilization effect. On Fc?RI and Fc?RIIIa large and sialylated glycans have a negative impact on rituximab binding, likely through destabilization of the interaction. In conclusion, the data show that the IgG1-Fc?R binding kinetics differ depending on the glycosylation of the Fc?R and further support a stabilizing role of Fc?R glycans in the antibody binding interaction.

Project description:The human Fc gamma RI (CD64) is a high affinity receptor for the Fc portion of immunoglobulin (Ig), and its constitutively low expression on the cell surface of monocyte/macrophage and neutrophils is selectively upregulated by interferon gamma (IFN-gamma) treatment (Perussia, B., E. T. Dayton, R. Lazarus, V. Fanning, and G. Trinchieri. 1983. J. Exp. Med. 158:1092). Three distinct cDNAs have been cloned and code for proteins that predict three extracellular Ig-like domains (Allen, J.M., and B. Seed. 1989. Science [Wash. DC]. 243:378). Several differences in the coding region of these cDNAs suggest that in addition to polymorphic differences a second Fc gamma RI gene could possibly exist. This alternative Fc gamma RI gene (Fc gamma RIb) was defined by the lack of a genomic HindIII restriction site (van der Winkel, J. G. J., L. U. Ernst, C. L. Anderson, and I. M. Chiu. 1991. J. Biol. Chem. 266:13449). We describe the characterization a second gene (Fc gamma RIb) that has a termination codon in the third extracellular domain and therefore predicts a soluble form of a termination codon in the third extracellular domain and therefore predicts a soluble form of the receptor. We also define two distinct IFN-gamma-responsive regions in the 5' flanking sequence of Fc gamma RIb that resemble motifs that have been defined in the class II major histocompatibility complex promoter. The Fc gamma RIb promoter does not possess canonical TATA or CCAAT boxes, but does possess a palindromic motif that closely resembles the initiator sequence identified in the terminal deoxynucleotidyl transferase/human leukocyte IFN/adeno-associated virus type II P5 gene promoters (Smale, S. T., and D. Baltimore. 1989. Cell. 57:103; Seto, E., Y. Shi, and T. Shenk. 1991. Nature [Lond.]. 354:241; Roy, A. L., M. Meisterernst, P. Pognonec, and R. C. Roeder. 1991. Nature [Lond.]. 354:245) virus type II P5 gene promoters raising interesting questions as to its role in the basal and myeloid-specific transcription of this gene.

Dataset Information

Identification and Functional Characterization of a Novel Fc Gamma-Binding Glycoprotein in Rhesus Cytomegalovirus.

Publications

Identification and Functional Characterization of a Novel Fc Gamma-Binding Glycoprotein in Rhesus Cytomegalovirus.

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