Conformational surveillance of Orai1 by a rhomboid intramembrane protease prevents inappropriate CRAC channel activation

Adam G Grieve; Yi-Chun Yeh; Yu-Fen Chang; Hsin-Yi Huang; Lucrezia Zarcone; Johannes Breuning; Nicholas Johnson; Kvido Stříšovský; Marion H Brown; Anant B Parekh; Matthew Freeman

doi:10.1016/j.molcel.2021.10.025

Conformational surveillance of Orai1 by a rhomboid intramembrane protease prevents inappropriate CRAC channel activation

Adam G Grieve^*, Yi-Chun Yeh, Yu-Fen Chang, Hsin-Yi Huang, Lucrezia Zarcone, Johannes Breuning, Nicholas Johnson, Kvido Stříšovský, Marion H Brown, Anant B Parekh, Matthew Freeman^*

^*Corresponding author for this work

School of Biochemistry

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Abstract

Calcium influx through plasma membrane calcium release-activated calcium (CRAC) channels, which are formed of hexamers of Orai1, is a potent trigger for many important biological processes, most notably in T cell-mediated immunity. Through a bioinformatics-led cell biological screen, we have identified Orai1 as a substrate for the rhomboid intramembrane protease RHBDL2. We show that RHBDL2 prevents stochastic calcium signaling in unstimulated cells through conformational surveillance and cleavage of inappropriately activated Orai1. A conserved disease-linked proline residue is responsible for RHBDL2's recognizing the active conformation of Orai1, which is required to sharpen switch-like signaling triggered by store-operated calcium entry. Loss of RHBDL2 control of CRAC channel activity causes severe dysregulation of downstream CRAC channel effectors, including transcription factor activation, inflammatory cytokine expression, and T cell activation. We propose that this surveillance function may represent an ancient activity of rhomboid proteases in degrading unwanted signaling proteins.

Original language	English
Pages (from-to)	4784-4798.e7
Number of pages	23
Journal	Molecular Cell
Volume	81
Issue number	23
Early online date	19 Nov 2021
DOIs	https://doi.org/10.1016/j.molcel.2021.10.025
Publication status	Published - 2 Dec 2021

Bibliographical note

Funding Information:
We thank members of the Freeman lab for their support and advice during the study, particularly Fangfang Lu and Nina Jajcanin-Jozic for their feedback on the manuscript. We thank Pedro Carvalho (Dunn School, Oxford University), Tim Levine (Institute of Ophthalmology, University College London [UCL]), and Francesca Robertson (Biochemistry Department, Oxford University) for critical reading of the manuscript. We also thank Prof. Tim Levine for his guidance with the use of HHpred. We would like to thank Dr. Michael van der Weijer for the generous gift of pLVX plasmids subcloned with zeocin and blasticidin resistance genes. We gratefully acknowledge the support of Dr. Alan Wainman in the light microscopy facility and Dr. Michal Maj in the flow cytometry facility at the Dunn School. This research was supported by the following grants: a MOST grant ( 109-2320-B-006-016-MY3 ) to Y.-C.Y., a CIU Trust grant to J.B., a European Regional Development Fund project ( CZ.02.1.01/0.0/0.0/16_019/0000729 ) and a Czech Science Foundation grant (21-24456S) to K.S., a Medical Research Council (MRC) grant ( LO1047CX ) to A.B.P., a Marie Skłodowska-Curie fellowship (Horizon 2020 Framework Programme 659166 ) to A.G.G.; a Biotechnology and Biological Sciences Research Council (BBSRC) research grant ( BB/RO16771/1 ) to A.G.G. and M.F., and Wellcome Trust Investigator Awards ( 101035/Z/13/Z and 220887/Z/20/Z ) to M.F.

Publisher Copyright:
© 2021 The Author(s)

Keywords

rhomboid protease
Orai1
RHBDL2
transmembrane
calcium
T cell
signalling

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Grieve, A. G., Yeh, Y.-C., Chang, Y.-F., Huang, H.-Y., Zarcone, L., Breuning, J., Johnson, N., Stříšovský, K., Brown, M. H., Parekh, A. B., & Freeman, M. (2021). Conformational surveillance of Orai1 by a rhomboid intramembrane protease prevents inappropriate CRAC channel activation. Molecular Cell, 81(23), 4784-4798.e7. https://doi.org/10.1016/j.molcel.2021.10.025

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abstract = "Calcium influx through plasma membrane calcium release-activated calcium (CRAC) channels, which are formed of hexamers of Orai1, is a potent trigger for many important biological processes, most notably in T cell-mediated immunity. Through a bioinformatics-led cell biological screen, we have identified Orai1 as a substrate for the rhomboid intramembrane protease RHBDL2. We show that RHBDL2 prevents stochastic calcium signaling in unstimulated cells through conformational surveillance and cleavage of inappropriately activated Orai1. A conserved disease-linked proline residue is responsible for RHBDL2's recognizing the active conformation of Orai1, which is required to sharpen switch-like signaling triggered by store-operated calcium entry. Loss of RHBDL2 control of CRAC channel activity causes severe dysregulation of downstream CRAC channel effectors, including transcription factor activation, inflammatory cytokine expression, and T cell activation. We propose that this surveillance function may represent an ancient activity of rhomboid proteases in degrading unwanted signaling proteins.",

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note = "Funding Information: We thank members of the Freeman lab for their support and advice during the study, particularly Fangfang Lu and Nina Jajcanin-Jozic for their feedback on the manuscript. We thank Pedro Carvalho (Dunn School, Oxford University), Tim Levine (Institute of Ophthalmology, University College London [UCL]), and Francesca Robertson (Biochemistry Department, Oxford University) for critical reading of the manuscript. We also thank Prof. Tim Levine for his guidance with the use of HHpred. We would like to thank Dr. Michael van der Weijer for the generous gift of pLVX plasmids subcloned with zeocin and blasticidin resistance genes. We gratefully acknowledge the support of Dr. Alan Wainman in the light microscopy facility and Dr. Michal Maj in the flow cytometry facility at the Dunn School. This research was supported by the following grants: a MOST grant ( 109-2320-B-006-016-MY3 ) to Y.-C.Y., a CIU Trust grant to J.B., a European Regional Development Fund project ( CZ.02.1.01/0.0/0.0/16_019/0000729 ) and a Czech Science Foundation grant (21-24456S) to K.S., a Medical Research Council (MRC) grant ( LO1047CX ) to A.B.P., a Marie Sk{\l}odowska-Curie fellowship (Horizon 2020 Framework Programme 659166 ) to A.G.G.; a Biotechnology and Biological Sciences Research Council (BBSRC) research grant ( BB/RO16771/1 ) to A.G.G. and M.F., and Wellcome Trust Investigator Awards ( 101035/Z/13/Z and 220887/Z/20/Z ) to M.F. Publisher Copyright: {\textcopyright} 2021 The Author(s)",

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doi = "10.1016/j.molcel.2021.10.025",

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T1 - Conformational surveillance of Orai1 by a rhomboid intramembrane protease prevents inappropriate CRAC channel activation

AU - Grieve, Adam G

AU - Yeh, Yi-Chun

AU - Chang, Yu-Fen

AU - Huang, Hsin-Yi

AU - Zarcone, Lucrezia

AU - Breuning, Johannes

AU - Johnson, Nicholas

AU - Stříšovský, Kvido

AU - Brown, Marion H

AU - Parekh, Anant B

AU - Freeman, Matthew

N1 - Funding Information: We thank members of the Freeman lab for their support and advice during the study, particularly Fangfang Lu and Nina Jajcanin-Jozic for their feedback on the manuscript. We thank Pedro Carvalho (Dunn School, Oxford University), Tim Levine (Institute of Ophthalmology, University College London [UCL]), and Francesca Robertson (Biochemistry Department, Oxford University) for critical reading of the manuscript. We also thank Prof. Tim Levine for his guidance with the use of HHpred. We would like to thank Dr. Michael van der Weijer for the generous gift of pLVX plasmids subcloned with zeocin and blasticidin resistance genes. We gratefully acknowledge the support of Dr. Alan Wainman in the light microscopy facility and Dr. Michal Maj in the flow cytometry facility at the Dunn School. This research was supported by the following grants: a MOST grant ( 109-2320-B-006-016-MY3 ) to Y.-C.Y., a CIU Trust grant to J.B., a European Regional Development Fund project ( CZ.02.1.01/0.0/0.0/16_019/0000729 ) and a Czech Science Foundation grant (21-24456S) to K.S., a Medical Research Council (MRC) grant ( LO1047CX ) to A.B.P., a Marie Skłodowska-Curie fellowship (Horizon 2020 Framework Programme 659166 ) to A.G.G.; a Biotechnology and Biological Sciences Research Council (BBSRC) research grant ( BB/RO16771/1 ) to A.G.G. and M.F., and Wellcome Trust Investigator Awards ( 101035/Z/13/Z and 220887/Z/20/Z ) to M.F. Publisher Copyright: © 2021 The Author(s)

PY - 2021/12/2

Y1 - 2021/12/2

N2 - Calcium influx through plasma membrane calcium release-activated calcium (CRAC) channels, which are formed of hexamers of Orai1, is a potent trigger for many important biological processes, most notably in T cell-mediated immunity. Through a bioinformatics-led cell biological screen, we have identified Orai1 as a substrate for the rhomboid intramembrane protease RHBDL2. We show that RHBDL2 prevents stochastic calcium signaling in unstimulated cells through conformational surveillance and cleavage of inappropriately activated Orai1. A conserved disease-linked proline residue is responsible for RHBDL2's recognizing the active conformation of Orai1, which is required to sharpen switch-like signaling triggered by store-operated calcium entry. Loss of RHBDL2 control of CRAC channel activity causes severe dysregulation of downstream CRAC channel effectors, including transcription factor activation, inflammatory cytokine expression, and T cell activation. We propose that this surveillance function may represent an ancient activity of rhomboid proteases in degrading unwanted signaling proteins.

AB - Calcium influx through plasma membrane calcium release-activated calcium (CRAC) channels, which are formed of hexamers of Orai1, is a potent trigger for many important biological processes, most notably in T cell-mediated immunity. Through a bioinformatics-led cell biological screen, we have identified Orai1 as a substrate for the rhomboid intramembrane protease RHBDL2. We show that RHBDL2 prevents stochastic calcium signaling in unstimulated cells through conformational surveillance and cleavage of inappropriately activated Orai1. A conserved disease-linked proline residue is responsible for RHBDL2's recognizing the active conformation of Orai1, which is required to sharpen switch-like signaling triggered by store-operated calcium entry. Loss of RHBDL2 control of CRAC channel activity causes severe dysregulation of downstream CRAC channel effectors, including transcription factor activation, inflammatory cytokine expression, and T cell activation. We propose that this surveillance function may represent an ancient activity of rhomboid proteases in degrading unwanted signaling proteins.

KW - rhomboid protease

KW - Orai1

KW - RHBDL2

KW - transmembrane

KW - calcium

KW - T cell

KW - signalling

U2 - 10.1016/j.molcel.2021.10.025

DO - 10.1016/j.molcel.2021.10.025

M3 - Article (Academic Journal)

C2 - 34800360

SN - 1097-2765

VL - 81

SP - 4784-4798.e7

JO - Molecular Cell

JF - Molecular Cell

IS - 23

ER -

Conformational surveillance of Orai1 by a rhomboid intramembrane protease prevents inappropriate CRAC channel activation

Abstract

Bibliographical note

Keywords

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