MyD88 TIR domain higher-order assembly interactions revealed by microcrystal electron diffraction and serial femtosecond crystallography

Max T.B. Clabbers; Susannah Holmes; Timothy W. Muusse; Parimala R. Vajjhala; Sara J. Thygesen; Alpeshkumar K. Malde; Dominic J.B. Hunter; Tristan I. Croll; Leonie Flueckiger; Jeffrey D. Nanson; Md Habibur Rahaman; Andrew Aquila; Mark S. Hunter; Mengning Liang; Chun Hong Yoon; Jingjing Zhao; Nadia A. Zatsepin; Brian Abbey; Emma Sierecki; Yann Gambin; Katryn J. Stacey; Connie Darmanin; Bostjan Kobe; Hongyi Xu; Thomas Ve

doi:10.1038/s41467-021-22590-6

MyD88 TIR domain higher-order assembly interactions revealed by microcrystal electron diffraction and serial femtosecond crystallography

Max T.B. Clabbers, Susannah Holmes, Timothy W. Muusse, Parimala R. Vajjhala, Sara J. Thygesen, Alpeshkumar K. Malde, Dominic J.B. Hunter, Tristan I. Croll, Leonie Flueckiger, Jeffrey D. Nanson, Md Habibur Rahaman, Andrew Aquila, Mark S. Hunter, Mengning Liang, Chun Hong Yoon, Jingjing Zhao, Nadia A. Zatsepin, Brian Abbey, Emma Sierecki, Yann GambinKatryn J. Stacey, Connie Darmanin^*, Bostjan Kobe^*, Hongyi Xu^*, Thomas Ve^*

^*Corresponding author for this work

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

74 Citations (Scopus)

Abstract

MyD88 and MAL are Toll-like receptor (TLR) adaptors that signal to induce pro-inflammatory cytokine production. We previously observed that the TIR domain of MAL (MAL^TIR) forms filaments in vitro and induces formation of crystalline higher-order assemblies of the MyD88 TIR domain (MyD88^TIR). These crystals are too small for conventional X-ray crystallography, but are ideally suited to structure determination by microcrystal electron diffraction (MicroED) and serial femtosecond crystallography (SFX). Here, we present MicroED and SFX structures of the MyD88^TIR assembly, which reveal a two-stranded higher-order assembly arrangement of TIR domains analogous to that seen previously for MAL^TIR. We demonstrate via mutagenesis that the MyD88^TIR assembly interfaces are critical for TLR4 signaling in vivo, and we show that MAL promotes unidirectional assembly of MyD88^TIR. Collectively, our studies provide structural and mechanistic insight into TLR signal transduction and allow a direct comparison of the MicroED and SFX techniques.

Original language	English
Article number	2578
Number of pages	14
Journal	Nature Communications
Volume	12
Issue number	1
Early online date	10 May 2021
DOIs	https://doi.org/10.1038/s41467-021-22590-6
Publication status	Published - Dec 2021
Externally published	Yes

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Clabbers, M. T. B., Holmes, S., Muusse, T. W., Vajjhala, P. R., Thygesen, S. J., Malde, A. K., Hunter, D. J. B., Croll, T. I., Flueckiger, L., Nanson, J. D., Rahaman, M. H., Aquila, A., Hunter, M. S., Liang, M., Yoon, C. H., Zhao, J., Zatsepin, N. A., Abbey, B., Sierecki, E., ... Ve, T. (2021). MyD88 TIR domain higher-order assembly interactions revealed by microcrystal electron diffraction and serial femtosecond crystallography. Nature Communications, 12(1), Article 2578. https://doi.org/10.1038/s41467-021-22590-6

@article{57717c371f3242e1a88b63b8eb40802d,

title = "MyD88 TIR domain higher-order assembly interactions revealed by microcrystal electron diffraction and serial femtosecond crystallography",

abstract = "MyD88 and MAL are Toll-like receptor (TLR) adaptors that signal to induce pro-inflammatory cytokine production. We previously observed that the TIR domain of MAL (MALTIR) forms filaments in vitro and induces formation of crystalline higher-order assemblies of the MyD88 TIR domain (MyD88TIR). These crystals are too small for conventional X-ray crystallography, but are ideally suited to structure determination by microcrystal electron diffraction (MicroED) and serial femtosecond crystallography (SFX). Here, we present MicroED and SFX structures of the MyD88TIR assembly, which reveal a two-stranded higher-order assembly arrangement of TIR domains analogous to that seen previously for MALTIR. We demonstrate via mutagenesis that the MyD88TIR assembly interfaces are critical for TLR4 signaling in vivo, and we show that MAL promotes unidirectional assembly of MyD88TIR. Collectively, our studies provide structural and mechanistic insight into TLR signal transduction and allow a direct comparison of the MicroED and SFX techniques.",

author = "Clabbers, {Max T.B.} and Susannah Holmes and Muusse, {Timothy W.} and Vajjhala, {Parimala R.} and Thygesen, {Sara J.} and Malde, {Alpeshkumar K.} and Hunter, {Dominic J.B.} and Croll, {Tristan I.} and Leonie Flueckiger and Nanson, {Jeffrey D.} and Rahaman, {Md Habibur} and Andrew Aquila and Hunter, {Mark S.} and Mengning Liang and Yoon, {Chun Hong} and Jingjing Zhao and Zatsepin, {Nadia A.} and Brian Abbey and Emma Sierecki and Yann Gambin and Stacey, {Katryn J.} and Connie Darmanin and Bostjan Kobe and Hongyi Xu and Thomas Ve",

note = "Publisher Copyright: {\textcopyright} 2021, The Author(s).",

year = "2021",

month = dec,

doi = "10.1038/s41467-021-22590-6",

language = "English",

volume = "12",

journal = "Nature Communications",

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Clabbers, MTB, Holmes, S, Muusse, TW, Vajjhala, PR, Thygesen, SJ, Malde, AK, Hunter, DJB, Croll, TI, Flueckiger, L, Nanson, JD, Rahaman, MH, Aquila, A, Hunter, MS, Liang, M, Yoon, CH, Zhao, J, Zatsepin, NA, Abbey, B, Sierecki, E, Gambin, Y, Stacey, KJ, Darmanin, C, Kobe, B, Xu, H & Ve, T 2021, 'MyD88 TIR domain higher-order assembly interactions revealed by microcrystal electron diffraction and serial femtosecond crystallography', Nature Communications, vol. 12, no. 1, 2578. https://doi.org/10.1038/s41467-021-22590-6

TY - JOUR

T1 - MyD88 TIR domain higher-order assembly interactions revealed by microcrystal electron diffraction and serial femtosecond crystallography

AU - Clabbers, Max T.B.

AU - Holmes, Susannah

AU - Muusse, Timothy W.

AU - Vajjhala, Parimala R.

AU - Thygesen, Sara J.

AU - Malde, Alpeshkumar K.

AU - Hunter, Dominic J.B.

AU - Croll, Tristan I.

AU - Flueckiger, Leonie

AU - Nanson, Jeffrey D.

AU - Rahaman, Md Habibur

AU - Aquila, Andrew

AU - Hunter, Mark S.

AU - Liang, Mengning

AU - Yoon, Chun Hong

AU - Zhao, Jingjing

AU - Zatsepin, Nadia A.

AU - Abbey, Brian

AU - Sierecki, Emma

AU - Gambin, Yann

AU - Stacey, Katryn J.

AU - Darmanin, Connie

AU - Kobe, Bostjan

AU - Xu, Hongyi

AU - Ve, Thomas

PY - 2021/12

Y1 - 2021/12

N2 - MyD88 and MAL are Toll-like receptor (TLR) adaptors that signal to induce pro-inflammatory cytokine production. We previously observed that the TIR domain of MAL (MALTIR) forms filaments in vitro and induces formation of crystalline higher-order assemblies of the MyD88 TIR domain (MyD88TIR). These crystals are too small for conventional X-ray crystallography, but are ideally suited to structure determination by microcrystal electron diffraction (MicroED) and serial femtosecond crystallography (SFX). Here, we present MicroED and SFX structures of the MyD88TIR assembly, which reveal a two-stranded higher-order assembly arrangement of TIR domains analogous to that seen previously for MALTIR. We demonstrate via mutagenesis that the MyD88TIR assembly interfaces are critical for TLR4 signaling in vivo, and we show that MAL promotes unidirectional assembly of MyD88TIR. Collectively, our studies provide structural and mechanistic insight into TLR signal transduction and allow a direct comparison of the MicroED and SFX techniques.

AB - MyD88 and MAL are Toll-like receptor (TLR) adaptors that signal to induce pro-inflammatory cytokine production. We previously observed that the TIR domain of MAL (MALTIR) forms filaments in vitro and induces formation of crystalline higher-order assemblies of the MyD88 TIR domain (MyD88TIR). These crystals are too small for conventional X-ray crystallography, but are ideally suited to structure determination by microcrystal electron diffraction (MicroED) and serial femtosecond crystallography (SFX). Here, we present MicroED and SFX structures of the MyD88TIR assembly, which reveal a two-stranded higher-order assembly arrangement of TIR domains analogous to that seen previously for MALTIR. We demonstrate via mutagenesis that the MyD88TIR assembly interfaces are critical for TLR4 signaling in vivo, and we show that MAL promotes unidirectional assembly of MyD88TIR. Collectively, our studies provide structural and mechanistic insight into TLR signal transduction and allow a direct comparison of the MicroED and SFX techniques.

UR - http://www.scopus.com/inward/record.url?scp=85105587344&partnerID=8YFLogxK

U2 - 10.1038/s41467-021-22590-6

DO - 10.1038/s41467-021-22590-6

M3 - Article

C2 - 33972532

AN - SCOPUS:85105587344

SN - 2041-1723

VL - 12

JO - Nature Communications

JF - Nature Communications

IS - 1

M1 - 2578

ER -

MyD88 TIR domain higher-order assembly interactions revealed by microcrystal electron diffraction and serial femtosecond crystallography

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