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The dimeric Golgi protein Gorab binds to Sas6 as a monomer to mediate centriole duplication

The duplication and ninefold symmetry of the Drosophila centriole requires that the cartwheel molecule, Sas6, physically associates with Gorab, a trans-Golgi component. How Gorab achieves these disparate associations is unclear. Here, we use hydrogen–deuterium exchange mass spectrometry to define Go...

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Autores principales: Fatalska, Agnieszka, Stepinac, Emma, Richter, Magdalena, Kovacs, Levente, Pietras, Zbigniew, Puchinger, Martin, Dong, Gang, Dadlez, Michal, Glover, David M
Formato: Online Artículo Texto
Lenguaje:English
Publicado: eLife Sciences Publications, Ltd 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8009671/
https://www.ncbi.nlm.nih.gov/pubmed/33704067
http://dx.doi.org/10.7554/eLife.57241
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author Fatalska, Agnieszka
Stepinac, Emma
Richter, Magdalena
Kovacs, Levente
Pietras, Zbigniew
Puchinger, Martin
Dong, Gang
Dadlez, Michal
Glover, David M
author_facet Fatalska, Agnieszka
Stepinac, Emma
Richter, Magdalena
Kovacs, Levente
Pietras, Zbigniew
Puchinger, Martin
Dong, Gang
Dadlez, Michal
Glover, David M
author_sort Fatalska, Agnieszka
collection PubMed
description The duplication and ninefold symmetry of the Drosophila centriole requires that the cartwheel molecule, Sas6, physically associates with Gorab, a trans-Golgi component. How Gorab achieves these disparate associations is unclear. Here, we use hydrogen–deuterium exchange mass spectrometry to define Gorab’s interacting surfaces that mediate its subcellular localization. We identify a core stabilization sequence within Gorab’s C-terminal coiled-coil domain that enables homodimerization, binding to Rab6, and thereby trans-Golgi localization. By contrast, part of the Gorab monomer’s coiled-coil domain undergoes an antiparallel interaction with a segment of the parallel coiled-coil dimer of Sas6. This stable heterotrimeric complex can be visualized by electron microscopy. Mutation of a single leucine residue in Sas6’s Gorab-binding domain generates a Sas6 variant with a sixteenfold reduced binding affinity for Gorab that cannot support centriole duplication. Thus, Gorab dimers at the Golgi exist in equilibrium with Sas6-associated monomers at the centriole to balance Gorab’s dual role.
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spelling pubmed-80096712021-03-31 The dimeric Golgi protein Gorab binds to Sas6 as a monomer to mediate centriole duplication Fatalska, Agnieszka Stepinac, Emma Richter, Magdalena Kovacs, Levente Pietras, Zbigniew Puchinger, Martin Dong, Gang Dadlez, Michal Glover, David M eLife Cell Biology The duplication and ninefold symmetry of the Drosophila centriole requires that the cartwheel molecule, Sas6, physically associates with Gorab, a trans-Golgi component. How Gorab achieves these disparate associations is unclear. Here, we use hydrogen–deuterium exchange mass spectrometry to define Gorab’s interacting surfaces that mediate its subcellular localization. We identify a core stabilization sequence within Gorab’s C-terminal coiled-coil domain that enables homodimerization, binding to Rab6, and thereby trans-Golgi localization. By contrast, part of the Gorab monomer’s coiled-coil domain undergoes an antiparallel interaction with a segment of the parallel coiled-coil dimer of Sas6. This stable heterotrimeric complex can be visualized by electron microscopy. Mutation of a single leucine residue in Sas6’s Gorab-binding domain generates a Sas6 variant with a sixteenfold reduced binding affinity for Gorab that cannot support centriole duplication. Thus, Gorab dimers at the Golgi exist in equilibrium with Sas6-associated monomers at the centriole to balance Gorab’s dual role. eLife Sciences Publications, Ltd 2021-03-11 /pmc/articles/PMC8009671/ /pubmed/33704067 http://dx.doi.org/10.7554/eLife.57241 Text en © 2021, Fatalska et al http://creativecommons.org/licenses/by/4.0/ http://creativecommons.org/licenses/by/4.0/This article is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use and redistribution provided that the original author and source are credited.
spellingShingle Cell Biology
Fatalska, Agnieszka
Stepinac, Emma
Richter, Magdalena
Kovacs, Levente
Pietras, Zbigniew
Puchinger, Martin
Dong, Gang
Dadlez, Michal
Glover, David M
The dimeric Golgi protein Gorab binds to Sas6 as a monomer to mediate centriole duplication
title The dimeric Golgi protein Gorab binds to Sas6 as a monomer to mediate centriole duplication
title_full The dimeric Golgi protein Gorab binds to Sas6 as a monomer to mediate centriole duplication
title_fullStr The dimeric Golgi protein Gorab binds to Sas6 as a monomer to mediate centriole duplication
title_full_unstemmed The dimeric Golgi protein Gorab binds to Sas6 as a monomer to mediate centriole duplication
title_short The dimeric Golgi protein Gorab binds to Sas6 as a monomer to mediate centriole duplication
title_sort dimeric golgi protein gorab binds to sas6 as a monomer to mediate centriole duplication
topic Cell Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8009671/
https://www.ncbi.nlm.nih.gov/pubmed/33704067
http://dx.doi.org/10.7554/eLife.57241
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