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The Calpain-7 protease functions together with the ESCRT-III protein IST1 within the midbody to regulate the timing and completion of abscission

The Endosomal Sorting Complexes Required for Transport (ESCRT) machinery mediates the membrane fission step that completes cytokinetic abscission and separates dividing cells. Filaments composed of ESCRT-III subunits constrict membranes of the intercellular bridge midbody to the abscission point. Th...

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Autores principales: Paine, Elliott L, Skalicky, Jack J, Whitby, Frank G, Mackay, Douglas R, Ullman, Katharine S, Hill, Christopher P, Sundquist, Wesley I
Formato: Online Artículo Texto
Lenguaje:English
Publicado: eLife Sciences Publications, Ltd 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10586806/
https://www.ncbi.nlm.nih.gov/pubmed/37772788
http://dx.doi.org/10.7554/eLife.84515
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author Paine, Elliott L
Skalicky, Jack J
Whitby, Frank G
Mackay, Douglas R
Ullman, Katharine S
Hill, Christopher P
Sundquist, Wesley I
author_facet Paine, Elliott L
Skalicky, Jack J
Whitby, Frank G
Mackay, Douglas R
Ullman, Katharine S
Hill, Christopher P
Sundquist, Wesley I
author_sort Paine, Elliott L
collection PubMed
description The Endosomal Sorting Complexes Required for Transport (ESCRT) machinery mediates the membrane fission step that completes cytokinetic abscission and separates dividing cells. Filaments composed of ESCRT-III subunits constrict membranes of the intercellular bridge midbody to the abscission point. These filaments also bind and recruit cofactors whose activities help execute abscission and/or delay abscission timing in response to mitotic errors via the NoCut/Abscission checkpoint. We previously showed that the ESCRT-III subunit IST1 binds the cysteine protease Calpain-7 (CAPN7) and that CAPN7 is required for both efficient abscission and NoCut checkpoint maintenance (Wenzel et al., 2022). Here, we report biochemical and crystallographic studies showing that the tandem microtubule-interacting and trafficking (MIT) domains of CAPN7 bind simultaneously to two distinct IST1 MIT interaction motifs. Structure-guided point mutations in either CAPN7 MIT domain disrupted IST1 binding in vitro and in cells, and depletion/rescue experiments showed that the CAPN7-IST1 interaction is required for (1) CAPN7 recruitment to midbodies, (2) efficient abscission, and (3) NoCut checkpoint arrest. CAPN7 proteolytic activity is also required for abscission and checkpoint maintenance. Hence, IST1 recruits CAPN7 to midbodies, where its proteolytic activity is required to regulate and complete abscission.
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spelling pubmed-105868062023-10-20 The Calpain-7 protease functions together with the ESCRT-III protein IST1 within the midbody to regulate the timing and completion of abscission Paine, Elliott L Skalicky, Jack J Whitby, Frank G Mackay, Douglas R Ullman, Katharine S Hill, Christopher P Sundquist, Wesley I eLife Cell Biology The Endosomal Sorting Complexes Required for Transport (ESCRT) machinery mediates the membrane fission step that completes cytokinetic abscission and separates dividing cells. Filaments composed of ESCRT-III subunits constrict membranes of the intercellular bridge midbody to the abscission point. These filaments also bind and recruit cofactors whose activities help execute abscission and/or delay abscission timing in response to mitotic errors via the NoCut/Abscission checkpoint. We previously showed that the ESCRT-III subunit IST1 binds the cysteine protease Calpain-7 (CAPN7) and that CAPN7 is required for both efficient abscission and NoCut checkpoint maintenance (Wenzel et al., 2022). Here, we report biochemical and crystallographic studies showing that the tandem microtubule-interacting and trafficking (MIT) domains of CAPN7 bind simultaneously to two distinct IST1 MIT interaction motifs. Structure-guided point mutations in either CAPN7 MIT domain disrupted IST1 binding in vitro and in cells, and depletion/rescue experiments showed that the CAPN7-IST1 interaction is required for (1) CAPN7 recruitment to midbodies, (2) efficient abscission, and (3) NoCut checkpoint arrest. CAPN7 proteolytic activity is also required for abscission and checkpoint maintenance. Hence, IST1 recruits CAPN7 to midbodies, where its proteolytic activity is required to regulate and complete abscission. eLife Sciences Publications, Ltd 2023-09-29 /pmc/articles/PMC10586806/ /pubmed/37772788 http://dx.doi.org/10.7554/eLife.84515 Text en © 2023, Paine et al https://creativecommons.org/licenses/by/4.0/This article is distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use and redistribution provided that the original author and source are credited.
spellingShingle Cell Biology
Paine, Elliott L
Skalicky, Jack J
Whitby, Frank G
Mackay, Douglas R
Ullman, Katharine S
Hill, Christopher P
Sundquist, Wesley I
The Calpain-7 protease functions together with the ESCRT-III protein IST1 within the midbody to regulate the timing and completion of abscission
title The Calpain-7 protease functions together with the ESCRT-III protein IST1 within the midbody to regulate the timing and completion of abscission
title_full The Calpain-7 protease functions together with the ESCRT-III protein IST1 within the midbody to regulate the timing and completion of abscission
title_fullStr The Calpain-7 protease functions together with the ESCRT-III protein IST1 within the midbody to regulate the timing and completion of abscission
title_full_unstemmed The Calpain-7 protease functions together with the ESCRT-III protein IST1 within the midbody to regulate the timing and completion of abscission
title_short The Calpain-7 protease functions together with the ESCRT-III protein IST1 within the midbody to regulate the timing and completion of abscission
title_sort calpain-7 protease functions together with the escrt-iii protein ist1 within the midbody to regulate the timing and completion of abscission
topic Cell Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10586806/
https://www.ncbi.nlm.nih.gov/pubmed/37772788
http://dx.doi.org/10.7554/eLife.84515
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