Toll-like receptor 4 and macrophage scavenger receptor 1 crosstalk regulates phagocytosis of a fungal pathogen

Chinaemerem U. Onyishi; Guillaume E. Desanti; Alex L. Wilkinson; Samuel Lara-Reyna; Eva-Maria Frickel; Gyorgy Fejer; Olivier D. Christophe; Clare E. Bryant; Subhankar Mukhopadhyay; Siamon Gordon; Robin C. May

doi:10.1038/s41467-023-40635-w

Toll-like receptor 4 and macrophage scavenger receptor 1 crosstalk regulates phagocytosis of a fungal pathogen

Chinaemerem U. Onyishi, Guillaume E. Desanti, Alex L. Wilkinson, Samuel Lara-Reyna, Eva-Maria Frickel, Gyorgy Fejer, Olivier D. Christophe, Clare E. Bryant, Subhankar Mukhopadhyay, Siamon Gordon, Robin C. May^*

^*Corresponding author for this work

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Abstract

The opportunistic fungal pathogen Cryptococcus neoformans causes lethal infections in immunocompromised patients. Macrophages are central to the host response to cryptococci; however, it is unclear how C. neoformans is recognised and phagocytosed by macrophages. Here we investigate the role of TLR4 in the non-opsonic phagocytosis of C. neoformans. We find that loss of TLR4 function unexpectedly increases phagocytosis of non-opsonised cryptococci by murine and human macrophages. The increased phagocytosis observed in Tlr4^−/− cells was dampened by pre-treatment of macrophages with oxidised-LDL, a known ligand of scavenger receptors. The scavenger receptor, macrophage scavenger receptor 1 (MSR1) (also known as SR-A1 or CD204) was upregulated in Tlr4^−/− macrophages. Genetic ablation of MSR1 resulted in a 75% decrease in phagocytosis of non-opsonised cryptococci, strongly suggesting that it is a key non-opsonic receptor for this pathogen. We go on to show that MSR1-mediated uptake likely involves the formation of a multimolecular signalling complex involving FcγR leading to SYK, PI3K, p38 and ERK1/2 activation to drive actin remodelling and phagocytosis. Altogether, our data indicate a hitherto unidentified role for TLR4/MSR1 crosstalk in the non-opsonic phagocytosis of C. neoformans.

Original language	English
Article number	4895
Number of pages	13
Journal	Nature Communications
Volume	14
Issue number	1
DOIs	https://doi.org/10.1038/s41467-023-40635-w
Publication status	Published - 14 Aug 2023

Bibliographical note

Acknowledgments:
We thank Maria Makarova for assistance with confocal microscopy, Michael G. Tomlinson for his in-silico analysis of MSR1 and contribution to the discussion, Sean D. Kelly for technical help regarding the establishment of MPI cell lines and Sarah Dimeloe and Nancy Gudgeon for helpful advice on the culture of PBMCs. The MSR1 knockout and MSR1/MARCO double knockout lines were established with the support of NC3Rs Grant NC/V001019/1. Work in C.E.B. lab is supported by grant BB/V000276/1. This work was partly supported by a Royal Society project grant (RGS\R2\202260) to S.M lab. C.U.O is supported by a Ph.D. studentship from the Darwin Trust of Edinburgh. A.L.W. was supported by Ph.D. funding from the Wellcome Trust ‘MIDAS’ doctoral training program. R.C.M. gratefully acknowledges support from the BBSRC and European Research Council Consolidator Award.

Copyright:
© 2023. Springer Nature Limited.

Keywords

Animals
Humans
Mice
Cryptococcosis
Cryptococcus neoformans
Macrophages/microbiology
Phagocytosis
Toll-Like Receptor 4/genetics
Scavenger Receptors, Class A/metabolism

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Cite this

Onyishi, C. U., Desanti, G. E., Wilkinson, A. L., Lara-Reyna, S., Frickel, E.-M., Fejer, G., Christophe, O. D., Bryant, C. E., Mukhopadhyay, S., Gordon, S., & May, R. C. (2023). Toll-like receptor 4 and macrophage scavenger receptor 1 crosstalk regulates phagocytosis of a fungal pathogen. Nature Communications, 14(1), Article 4895. https://doi.org/10.1038/s41467-023-40635-w

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abstract = "The opportunistic fungal pathogen Cryptococcus neoformans causes lethal infections in immunocompromised patients. Macrophages are central to the host response to cryptococci; however, it is unclear how C. neoformans is recognised and phagocytosed by macrophages. Here we investigate the role of TLR4 in the non-opsonic phagocytosis of C. neoformans. We find that loss of TLR4 function unexpectedly increases phagocytosis of non-opsonised cryptococci by murine and human macrophages. The increased phagocytosis observed in Tlr4−/− cells was dampened by pre-treatment of macrophages with oxidised-LDL, a known ligand of scavenger receptors. The scavenger receptor, macrophage scavenger receptor 1 (MSR1) (also known as SR-A1 or CD204) was upregulated in Tlr4−/− macrophages. Genetic ablation of MSR1 resulted in a 75% decrease in phagocytosis of non-opsonised cryptococci, strongly suggesting that it is a key non-opsonic receptor for this pathogen. We go on to show that MSR1-mediated uptake likely involves the formation of a multimolecular signalling complex involving FcγR leading to SYK, PI3K, p38 and ERK1/2 activation to drive actin remodelling and phagocytosis. Altogether, our data indicate a hitherto unidentified role for TLR4/MSR1 crosstalk in the non-opsonic phagocytosis of C. neoformans.",

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author = "Onyishi, {Chinaemerem U.} and Desanti, {Guillaume E.} and Wilkinson, {Alex L.} and Samuel Lara-Reyna and Eva-Maria Frickel and Gyorgy Fejer and Christophe, {Olivier D.} and Bryant, {Clare E.} and Subhankar Mukhopadhyay and Siamon Gordon and May, {Robin C.}",

note = "Acknowledgments: We thank Maria Makarova for assistance with confocal microscopy, Michael G. Tomlinson for his in-silico analysis of MSR1 and contribution to the discussion, Sean D. Kelly for technical help regarding the establishment of MPI cell lines and Sarah Dimeloe and Nancy Gudgeon for helpful advice on the culture of PBMCs. The MSR1 knockout and MSR1/MARCO double knockout lines were established with the support of NC3Rs Grant NC/V001019/1. Work in C.E.B. lab is supported by grant BB/V000276/1. This work was partly supported by a Royal Society project grant (RGS\R2\202260) to S.M lab. C.U.O is supported by a Ph.D. studentship from the Darwin Trust of Edinburgh. A.L.W. was supported by Ph.D. funding from the Wellcome Trust {\textquoteleft}MIDAS{\textquoteright} doctoral training program. R.C.M. gratefully acknowledges support from the BBSRC and European Research Council Consolidator Award. Copyright: {\textcopyright} 2023. Springer Nature Limited.",

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AU - Lara-Reyna, Samuel

AU - Frickel, Eva-Maria

AU - Fejer, Gyorgy

AU - Christophe, Olivier D.

AU - Bryant, Clare E.

AU - Mukhopadhyay, Subhankar

AU - Gordon, Siamon

AU - May, Robin C.

N1 - Acknowledgments: We thank Maria Makarova for assistance with confocal microscopy, Michael G. Tomlinson for his in-silico analysis of MSR1 and contribution to the discussion, Sean D. Kelly for technical help regarding the establishment of MPI cell lines and Sarah Dimeloe and Nancy Gudgeon for helpful advice on the culture of PBMCs. The MSR1 knockout and MSR1/MARCO double knockout lines were established with the support of NC3Rs Grant NC/V001019/1. Work in C.E.B. lab is supported by grant BB/V000276/1. This work was partly supported by a Royal Society project grant (RGS\R2\202260) to S.M lab. C.U.O is supported by a Ph.D. studentship from the Darwin Trust of Edinburgh. A.L.W. was supported by Ph.D. funding from the Wellcome Trust ‘MIDAS’ doctoral training program. R.C.M. gratefully acknowledges support from the BBSRC and European Research Council Consolidator Award. Copyright: © 2023. Springer Nature Limited.

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N2 - The opportunistic fungal pathogen Cryptococcus neoformans causes lethal infections in immunocompromised patients. Macrophages are central to the host response to cryptococci; however, it is unclear how C. neoformans is recognised and phagocytosed by macrophages. Here we investigate the role of TLR4 in the non-opsonic phagocytosis of C. neoformans. We find that loss of TLR4 function unexpectedly increases phagocytosis of non-opsonised cryptococci by murine and human macrophages. The increased phagocytosis observed in Tlr4−/− cells was dampened by pre-treatment of macrophages with oxidised-LDL, a known ligand of scavenger receptors. The scavenger receptor, macrophage scavenger receptor 1 (MSR1) (also known as SR-A1 or CD204) was upregulated in Tlr4−/− macrophages. Genetic ablation of MSR1 resulted in a 75% decrease in phagocytosis of non-opsonised cryptococci, strongly suggesting that it is a key non-opsonic receptor for this pathogen. We go on to show that MSR1-mediated uptake likely involves the formation of a multimolecular signalling complex involving FcγR leading to SYK, PI3K, p38 and ERK1/2 activation to drive actin remodelling and phagocytosis. Altogether, our data indicate a hitherto unidentified role for TLR4/MSR1 crosstalk in the non-opsonic phagocytosis of C. neoformans.

AB - The opportunistic fungal pathogen Cryptococcus neoformans causes lethal infections in immunocompromised patients. Macrophages are central to the host response to cryptococci; however, it is unclear how C. neoformans is recognised and phagocytosed by macrophages. Here we investigate the role of TLR4 in the non-opsonic phagocytosis of C. neoformans. We find that loss of TLR4 function unexpectedly increases phagocytosis of non-opsonised cryptococci by murine and human macrophages. The increased phagocytosis observed in Tlr4−/− cells was dampened by pre-treatment of macrophages with oxidised-LDL, a known ligand of scavenger receptors. The scavenger receptor, macrophage scavenger receptor 1 (MSR1) (also known as SR-A1 or CD204) was upregulated in Tlr4−/− macrophages. Genetic ablation of MSR1 resulted in a 75% decrease in phagocytosis of non-opsonised cryptococci, strongly suggesting that it is a key non-opsonic receptor for this pathogen. We go on to show that MSR1-mediated uptake likely involves the formation of a multimolecular signalling complex involving FcγR leading to SYK, PI3K, p38 and ERK1/2 activation to drive actin remodelling and phagocytosis. Altogether, our data indicate a hitherto unidentified role for TLR4/MSR1 crosstalk in the non-opsonic phagocytosis of C. neoformans.

KW - Animals

KW - Humans

KW - Mice

KW - Cryptococcosis

KW - Cryptococcus neoformans

KW - Macrophages/microbiology

KW - Phagocytosis

KW - Toll-Like Receptor 4/genetics

KW - Scavenger Receptors, Class A/metabolism

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VL - 14

JO - Nature Communications

JF - Nature Communications

IS - 1

M1 - 4895

ER -

Toll-like receptor 4 and macrophage scavenger receptor 1 crosstalk regulates phagocytosis of a fungal pathogen

Abstract

Bibliographical note

Keywords

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