Mouse cytomegalovirus m153 protein stabilizes expression of the inhibitory NKR-P1B ligand Clr-b

Oscar A. Aguilar; Isabella S. Sampaio; Mir Munir A. Rahim; Jackeline D. Samaniego; Mulualem E. Tilahun; Mithunah Krishnamoorthy; Branka Popović; Marina Babić; Astrid Krmpotić; Bebhinn Treanor; David H. Margulies; David S.J. Allan; Andrew P. Makrigiannis; Stipan Jonjić; James R. Carlyle

doi:10.1128/JVI.01220-19

Mouse cytomegalovirus m153 protein stabilizes expression of the inhibitory NKR-P1B ligand Clr-b

Oscar A. Aguilar, Isabella S. Sampaio, Mir Munir A. Rahim, Jackeline D. Samaniego, Mulualem E. Tilahun, Mithunah Krishnamoorthy, Branka Popović, Marina Babić, Astrid Krmpotić, Bebhinn Treanor, David H. Margulies, David S.J. Allan, Andrew P. Makrigiannis, Stipan Jonjić, James R. Carlyle

Medicine

Research output: Contribution to journal › Article › peer-review

8 Citations (Scopus)

Abstract

Natural killer (NK) cells are a subset of innate lymphoid cells (ILC) capable of recognizing stressed and infected cells through multiple germ line-encoded receptor-ligand interactions. Missing-self recognition involves NK cell sensing of the loss of host-encoded inhibitory ligands on target cells, including MHC class I (MHC-I) molecules and other MHC-I-independent ligands. Mouse cytomegalovirus (MCMV) infection promotes a rapid host-mediated loss of the inhibitory NKR-P1B ligand Clr-b (encoded by Clec2d) on infected cells. Here we provide evidence that an MCMV m145 family member, m153, functions to stabilize cell surface Clr-b during MCMV infection. Ectopic expression of m153 in fibroblasts augments Clr-b cell surface levels. Moreover, infections using m153-deficient MCMV mutants (Δm144-m158 and Δm153) show an accelerated and exacerbated Clr-b downregulation. Importantly, enhanced loss of Clr-b during Δm153 mutant infection reverts to wild-type levels upon exogenous m153 complementation in fibroblasts. While the effects of m153 on Clr-b levels are independent of Clec2d transcription, imaging experiments revealed that the m153 and Clr-b proteins only minimally colocalize within the same subcellular compartments, and tagged versions of the proteins were refractory to coimmunoprecipitation under mild-detergent conditions. Surprisingly, the Δm153 mutant possesses enhanced virulence in vivo, independent of both Clr-b and NKRP1B, suggesting that m153 potentially targets additional host factors. Nevertheless, the present data highlight a unique mechanism by which MCMV modulates NK ligand expression. IMPORTANCE Cytomegaloviruses are betaherpesviruses that in immunocompromised individuals can lead to severe pathologies. These viruses encode various gene products that serve to evade innate immune recognition. NK cells are among the first immune cells that respond to CMV infection and use germ lineencoded NK cell receptors (NKR) to distinguish healthy from virus-infected cells. One such axis that plays a critical role in NK recognition involves the inhibitory NKR-P1B receptor, which engages the host ligand Clr-b, a molecule commonly lost on stressed cells ("missing-self"). In this study, we discovered that mouse CMV utilizes the m153 glycoprotein to circumvent host-mediated Clr-b downregulation, in order to evade NK recognition. These results highlight a novel MCMV-mediated immune evasion strategy.

Original language	English
Article number	e01220-19
Journal	Journal of Virology
Volume	94
Issue number	1
DOIs	https://doi.org/10.1128/JVI.01220-19
Publication status	Published - Jan 1 2020

Bibliographical note

Funding Information:
Institute. I.S.S. was supported by a Science Without Borders scholarship from the Brazilian National Council for Scientific and Technological Development (CNPq). This work was supported by operating grants from the Canadian Institute of Health Research (number 86630 to A.P.M. and J.R.C., number 106491 to J.R.C., and number 136808 to B.T.), by an Investigator in the Pathogenesis of Infectious Disease from the Burroughs Wellcome Fund (number 1007761 to J.R.C.), a European Regional and Development Fund (K.01.1.1.01.0006) awarded to the Scientific Centre of Excellence for Virus Immunology and Vaccines and cofinanced by the European Regional Development Fund (S.J. and A.K.), and by the intramural research program of the NIAID, NIH (ZIA AI001028-11 to D.H.M.).

Funding Information:
O.A.A. was supported by a Postgraduate Scholarship-Doctoral Award from the National Sciences and Engineering Research Council of Canada. Oscar A. Aguilar holds a Postdoctoral Enrichment Program Award from the Burroughs Wellcome Fund (BWF) and is a Cancer Research Institute Irvington Fellow supported by the Cancer Research

Publisher Copyright:
© 2019 American Society for Microbiology.

ASJC Scopus Subject Areas

Microbiology
Immunology
Insect Science
Virology

PubMed: MeSH publication types

Journal Article
Research Support, N.I.H., Intramural
Research Support, Non-U.S. Gov't

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10.1128/JVI.01220-19

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Aguilar, O. A., Sampaio, I. S., Rahim, M. M. A., Samaniego, J. D., Tilahun, M. E., Krishnamoorthy, M., Popović, B., Babić, M., Krmpotić, A., Treanor, B., Margulies, D. H., Allan, D. S. J., Makrigiannis, A. P., Jonjić, S., & Carlyle, J. R. (2020). Mouse cytomegalovirus m153 protein stabilizes expression of the inhibitory NKR-P1B ligand Clr-b. Journal of Virology, 94(1), Article e01220-19. https://doi.org/10.1128/JVI.01220-19

Aguilar, OA, Sampaio, IS, Rahim, MMA, Samaniego, JD, Tilahun, ME, Krishnamoorthy, M, Popović, B, Babić, M, Krmpotić, A, Treanor, B, Margulies, DH, Allan, DSJ, Makrigiannis, AP, Jonjić, S & Carlyle, JR 2020, 'Mouse cytomegalovirus m153 protein stabilizes expression of the inhibitory NKR-P1B ligand Clr-b', Journal of Virology, vol. 94, no. 1, e01220-19. https://doi.org/10.1128/JVI.01220-19

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title = "Mouse cytomegalovirus m153 protein stabilizes expression of the inhibitory NKR-P1B ligand Clr-b",

abstract = "Natural killer (NK) cells are a subset of innate lymphoid cells (ILC) capable of recognizing stressed and infected cells through multiple germ line-encoded receptor-ligand interactions. Missing-self recognition involves NK cell sensing of the loss of host-encoded inhibitory ligands on target cells, including MHC class I (MHC-I) molecules and other MHC-I-independent ligands. Mouse cytomegalovirus (MCMV) infection promotes a rapid host-mediated loss of the inhibitory NKR-P1B ligand Clr-b (encoded by Clec2d) on infected cells. Here we provide evidence that an MCMV m145 family member, m153, functions to stabilize cell surface Clr-b during MCMV infection. Ectopic expression of m153 in fibroblasts augments Clr-b cell surface levels. Moreover, infections using m153-deficient MCMV mutants (Δm144-m158 and Δm153) show an accelerated and exacerbated Clr-b downregulation. Importantly, enhanced loss of Clr-b during Δm153 mutant infection reverts to wild-type levels upon exogenous m153 complementation in fibroblasts. While the effects of m153 on Clr-b levels are independent of Clec2d transcription, imaging experiments revealed that the m153 and Clr-b proteins only minimally colocalize within the same subcellular compartments, and tagged versions of the proteins were refractory to coimmunoprecipitation under mild-detergent conditions. Surprisingly, the Δm153 mutant possesses enhanced virulence in vivo, independent of both Clr-b and NKRP1B, suggesting that m153 potentially targets additional host factors. Nevertheless, the present data highlight a unique mechanism by which MCMV modulates NK ligand expression. IMPORTANCE Cytomegaloviruses are betaherpesviruses that in immunocompromised individuals can lead to severe pathologies. These viruses encode various gene products that serve to evade innate immune recognition. NK cells are among the first immune cells that respond to CMV infection and use germ lineencoded NK cell receptors (NKR) to distinguish healthy from virus-infected cells. One such axis that plays a critical role in NK recognition involves the inhibitory NKR-P1B receptor, which engages the host ligand Clr-b, a molecule commonly lost on stressed cells ({"}missing-self{"}). In this study, we discovered that mouse CMV utilizes the m153 glycoprotein to circumvent host-mediated Clr-b downregulation, in order to evade NK recognition. These results highlight a novel MCMV-mediated immune evasion strategy.",

author = "Aguilar, {Oscar A.} and Sampaio, {Isabella S.} and Rahim, {Mir Munir A.} and Samaniego, {Jackeline D.} and Tilahun, {Mulualem E.} and Mithunah Krishnamoorthy and Branka Popovi{\'c} and Marina Babi{\'c} and Astrid Krmpoti{\'c} and Bebhinn Treanor and Margulies, {David H.} and Allan, {David S.J.} and Makrigiannis, {Andrew P.} and Stipan Jonji{\'c} and Carlyle, {James R.}",

note = "Funding Information: Institute. I.S.S. was supported by a Science Without Borders scholarship from the Brazilian National Council for Scientific and Technological Development (CNPq). This work was supported by operating grants from the Canadian Institute of Health Research (number 86630 to A.P.M. and J.R.C., number 106491 to J.R.C., and number 136808 to B.T.), by an Investigator in the Pathogenesis of Infectious Disease from the Burroughs Wellcome Fund (number 1007761 to J.R.C.), a European Regional and Development Fund (K.01.1.1.01.0006) awarded to the Scientific Centre of Excellence for Virus Immunology and Vaccines and cofinanced by the European Regional Development Fund (S.J. and A.K.), and by the intramural research program of the NIAID, NIH (ZIA AI001028-11 to D.H.M.). Funding Information: O.A.A. was supported by a Postgraduate Scholarship-Doctoral Award from the National Sciences and Engineering Research Council of Canada. Oscar A. Aguilar holds a Postdoctoral Enrichment Program Award from the Burroughs Wellcome Fund (BWF) and is a Cancer Research Institute Irvington Fellow supported by the Cancer Research Publisher Copyright: {\textcopyright} 2019 American Society for Microbiology.",

year = "2020",

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doi = "10.1128/JVI.01220-19",

language = "English",

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T1 - Mouse cytomegalovirus m153 protein stabilizes expression of the inhibitory NKR-P1B ligand Clr-b

AU - Aguilar, Oscar A.

AU - Sampaio, Isabella S.

AU - Rahim, Mir Munir A.

AU - Samaniego, Jackeline D.

AU - Tilahun, Mulualem E.

AU - Krishnamoorthy, Mithunah

AU - Popović, Branka

AU - Babić, Marina

AU - Krmpotić, Astrid

AU - Treanor, Bebhinn

AU - Margulies, David H.

AU - Allan, David S.J.

AU - Makrigiannis, Andrew P.

AU - Jonjić, Stipan

AU - Carlyle, James R.

N1 - Funding Information: Institute. I.S.S. was supported by a Science Without Borders scholarship from the Brazilian National Council for Scientific and Technological Development (CNPq). This work was supported by operating grants from the Canadian Institute of Health Research (number 86630 to A.P.M. and J.R.C., number 106491 to J.R.C., and number 136808 to B.T.), by an Investigator in the Pathogenesis of Infectious Disease from the Burroughs Wellcome Fund (number 1007761 to J.R.C.), a European Regional and Development Fund (K.01.1.1.01.0006) awarded to the Scientific Centre of Excellence for Virus Immunology and Vaccines and cofinanced by the European Regional Development Fund (S.J. and A.K.), and by the intramural research program of the NIAID, NIH (ZIA AI001028-11 to D.H.M.). Funding Information: O.A.A. was supported by a Postgraduate Scholarship-Doctoral Award from the National Sciences and Engineering Research Council of Canada. Oscar A. Aguilar holds a Postdoctoral Enrichment Program Award from the Burroughs Wellcome Fund (BWF) and is a Cancer Research Institute Irvington Fellow supported by the Cancer Research Publisher Copyright: © 2019 American Society for Microbiology.

PY - 2020/1/1

Y1 - 2020/1/1

N2 - Natural killer (NK) cells are a subset of innate lymphoid cells (ILC) capable of recognizing stressed and infected cells through multiple germ line-encoded receptor-ligand interactions. Missing-self recognition involves NK cell sensing of the loss of host-encoded inhibitory ligands on target cells, including MHC class I (MHC-I) molecules and other MHC-I-independent ligands. Mouse cytomegalovirus (MCMV) infection promotes a rapid host-mediated loss of the inhibitory NKR-P1B ligand Clr-b (encoded by Clec2d) on infected cells. Here we provide evidence that an MCMV m145 family member, m153, functions to stabilize cell surface Clr-b during MCMV infection. Ectopic expression of m153 in fibroblasts augments Clr-b cell surface levels. Moreover, infections using m153-deficient MCMV mutants (Δm144-m158 and Δm153) show an accelerated and exacerbated Clr-b downregulation. Importantly, enhanced loss of Clr-b during Δm153 mutant infection reverts to wild-type levels upon exogenous m153 complementation in fibroblasts. While the effects of m153 on Clr-b levels are independent of Clec2d transcription, imaging experiments revealed that the m153 and Clr-b proteins only minimally colocalize within the same subcellular compartments, and tagged versions of the proteins were refractory to coimmunoprecipitation under mild-detergent conditions. Surprisingly, the Δm153 mutant possesses enhanced virulence in vivo, independent of both Clr-b and NKRP1B, suggesting that m153 potentially targets additional host factors. Nevertheless, the present data highlight a unique mechanism by which MCMV modulates NK ligand expression. IMPORTANCE Cytomegaloviruses are betaherpesviruses that in immunocompromised individuals can lead to severe pathologies. These viruses encode various gene products that serve to evade innate immune recognition. NK cells are among the first immune cells that respond to CMV infection and use germ lineencoded NK cell receptors (NKR) to distinguish healthy from virus-infected cells. One such axis that plays a critical role in NK recognition involves the inhibitory NKR-P1B receptor, which engages the host ligand Clr-b, a molecule commonly lost on stressed cells ("missing-self"). In this study, we discovered that mouse CMV utilizes the m153 glycoprotein to circumvent host-mediated Clr-b downregulation, in order to evade NK recognition. These results highlight a novel MCMV-mediated immune evasion strategy.

AB - Natural killer (NK) cells are a subset of innate lymphoid cells (ILC) capable of recognizing stressed and infected cells through multiple germ line-encoded receptor-ligand interactions. Missing-self recognition involves NK cell sensing of the loss of host-encoded inhibitory ligands on target cells, including MHC class I (MHC-I) molecules and other MHC-I-independent ligands. Mouse cytomegalovirus (MCMV) infection promotes a rapid host-mediated loss of the inhibitory NKR-P1B ligand Clr-b (encoded by Clec2d) on infected cells. Here we provide evidence that an MCMV m145 family member, m153, functions to stabilize cell surface Clr-b during MCMV infection. Ectopic expression of m153 in fibroblasts augments Clr-b cell surface levels. Moreover, infections using m153-deficient MCMV mutants (Δm144-m158 and Δm153) show an accelerated and exacerbated Clr-b downregulation. Importantly, enhanced loss of Clr-b during Δm153 mutant infection reverts to wild-type levels upon exogenous m153 complementation in fibroblasts. While the effects of m153 on Clr-b levels are independent of Clec2d transcription, imaging experiments revealed that the m153 and Clr-b proteins only minimally colocalize within the same subcellular compartments, and tagged versions of the proteins were refractory to coimmunoprecipitation under mild-detergent conditions. Surprisingly, the Δm153 mutant possesses enhanced virulence in vivo, independent of both Clr-b and NKRP1B, suggesting that m153 potentially targets additional host factors. Nevertheless, the present data highlight a unique mechanism by which MCMV modulates NK ligand expression. IMPORTANCE Cytomegaloviruses are betaherpesviruses that in immunocompromised individuals can lead to severe pathologies. These viruses encode various gene products that serve to evade innate immune recognition. NK cells are among the first immune cells that respond to CMV infection and use germ lineencoded NK cell receptors (NKR) to distinguish healthy from virus-infected cells. One such axis that plays a critical role in NK recognition involves the inhibitory NKR-P1B receptor, which engages the host ligand Clr-b, a molecule commonly lost on stressed cells ("missing-self"). In this study, we discovered that mouse CMV utilizes the m153 glycoprotein to circumvent host-mediated Clr-b downregulation, in order to evade NK recognition. These results highlight a novel MCMV-mediated immune evasion strategy.

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Mouse cytomegalovirus m153 protein stabilizes expression of the inhibitory NKR-P1B ligand Clr-b

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PubMed: MeSH publication types

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