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EVC-EVC2 complex stability and ciliary targeting are regulated by modification with ubiquitin and SUMO

Ellis van Creveld syndrome and Weyers acrofacial dysostosis are two rare genetic diseases affecting skeletal development. They are both ciliopathies, as they are due to malfunction of primary cilia, microtubule-based plasma membrane protrusions that function as cellular antennae and are required for...

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Autores principales: Barbeito, Pablo, Martin-Morales, Raquel, Palencia-Campos, Adrian, Cerrolaza, Juan, Rivas-Santos, Celia, Gallego-Colastra, Leticia, Caparros-Martin, Jose Antonio, Martin-Bravo, Carolina, Martin-Hurtado, Ana, Sánchez-Bellver, Laura, Marfany, Gemma, Ruiz-Perez, Victor L., Garcia-Gonzalo, Francesc R.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2023
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10413113/
https://www.ncbi.nlm.nih.gov/pubmed/37576597
http://dx.doi.org/10.3389/fcell.2023.1190258
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author Barbeito, Pablo
Martin-Morales, Raquel
Palencia-Campos, Adrian
Cerrolaza, Juan
Rivas-Santos, Celia
Gallego-Colastra, Leticia
Caparros-Martin, Jose Antonio
Martin-Bravo, Carolina
Martin-Hurtado, Ana
Sánchez-Bellver, Laura
Marfany, Gemma
Ruiz-Perez, Victor L.
Garcia-Gonzalo, Francesc R.
author_facet Barbeito, Pablo
Martin-Morales, Raquel
Palencia-Campos, Adrian
Cerrolaza, Juan
Rivas-Santos, Celia
Gallego-Colastra, Leticia
Caparros-Martin, Jose Antonio
Martin-Bravo, Carolina
Martin-Hurtado, Ana
Sánchez-Bellver, Laura
Marfany, Gemma
Ruiz-Perez, Victor L.
Garcia-Gonzalo, Francesc R.
author_sort Barbeito, Pablo
collection PubMed
description Ellis van Creveld syndrome and Weyers acrofacial dysostosis are two rare genetic diseases affecting skeletal development. They are both ciliopathies, as they are due to malfunction of primary cilia, microtubule-based plasma membrane protrusions that function as cellular antennae and are required for Hedgehog signaling, a key pathway during skeletal morphogenesis. These ciliopathies are caused by mutations affecting the EVC-EVC2 complex, a transmembrane protein heterodimer that regulates Hedgehog signaling from inside primary cilia. Despite the importance of this complex, the mechanisms underlying its stability, targeting and function are poorly understood. To address this, we characterized the endogenous EVC protein interactome in control and Evc-null cells. This proteomic screen confirmed EVC’s main known interactors (EVC2, IQCE, EFCAB7), while revealing new ones, including USP7, a deubiquitinating enzyme involved in Hedgehog signaling. We therefore looked at EVC-EVC2 complex ubiquitination. Such ubiquitination exists but is independent of USP7 (and of USP48, also involved in Hh signaling). We did find, however, that monoubiquitination of EVC-EVC2 cytosolic tails greatly reduces their protein levels. On the other hand, modification of EVC-EVC2 cytosolic tails with the small ubiquitin-related modifier SUMO3 has a different effect, enhancing complex accumulation at the EvC zone, immediately distal to the ciliary transition zone, possibly via increased binding to the EFCAB7-IQCE complex. Lastly, we find that EvC zone targeting of EVC-EVC2 depends on two separate EFCAB7-binding motifs within EVC2’s Weyers-deleted peptide. Only one of these motifs had been characterized previously, so we have mapped the second herein. Altogether, our data shed light on EVC-EVC2 complex regulatory mechanisms, with implications for ciliopathies.
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spelling pubmed-104131132023-08-11 EVC-EVC2 complex stability and ciliary targeting are regulated by modification with ubiquitin and SUMO Barbeito, Pablo Martin-Morales, Raquel Palencia-Campos, Adrian Cerrolaza, Juan Rivas-Santos, Celia Gallego-Colastra, Leticia Caparros-Martin, Jose Antonio Martin-Bravo, Carolina Martin-Hurtado, Ana Sánchez-Bellver, Laura Marfany, Gemma Ruiz-Perez, Victor L. Garcia-Gonzalo, Francesc R. Front Cell Dev Biol Cell and Developmental Biology Ellis van Creveld syndrome and Weyers acrofacial dysostosis are two rare genetic diseases affecting skeletal development. They are both ciliopathies, as they are due to malfunction of primary cilia, microtubule-based plasma membrane protrusions that function as cellular antennae and are required for Hedgehog signaling, a key pathway during skeletal morphogenesis. These ciliopathies are caused by mutations affecting the EVC-EVC2 complex, a transmembrane protein heterodimer that regulates Hedgehog signaling from inside primary cilia. Despite the importance of this complex, the mechanisms underlying its stability, targeting and function are poorly understood. To address this, we characterized the endogenous EVC protein interactome in control and Evc-null cells. This proteomic screen confirmed EVC’s main known interactors (EVC2, IQCE, EFCAB7), while revealing new ones, including USP7, a deubiquitinating enzyme involved in Hedgehog signaling. We therefore looked at EVC-EVC2 complex ubiquitination. Such ubiquitination exists but is independent of USP7 (and of USP48, also involved in Hh signaling). We did find, however, that monoubiquitination of EVC-EVC2 cytosolic tails greatly reduces their protein levels. On the other hand, modification of EVC-EVC2 cytosolic tails with the small ubiquitin-related modifier SUMO3 has a different effect, enhancing complex accumulation at the EvC zone, immediately distal to the ciliary transition zone, possibly via increased binding to the EFCAB7-IQCE complex. Lastly, we find that EvC zone targeting of EVC-EVC2 depends on two separate EFCAB7-binding motifs within EVC2’s Weyers-deleted peptide. Only one of these motifs had been characterized previously, so we have mapped the second herein. Altogether, our data shed light on EVC-EVC2 complex regulatory mechanisms, with implications for ciliopathies. Frontiers Media S.A. 2023-07-27 /pmc/articles/PMC10413113/ /pubmed/37576597 http://dx.doi.org/10.3389/fcell.2023.1190258 Text en Copyright © 2023 Barbeito, Martin-Morales, Palencia-Campos, Cerrolaza, Rivas-Santos, Gallego-Colastra, Caparros-Martin, Martin-Bravo, Martin-Hurtado, Sánchez-Bellver, Marfany, Ruiz-Perez and Garcia-Gonzalo. https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Cell and Developmental Biology
Barbeito, Pablo
Martin-Morales, Raquel
Palencia-Campos, Adrian
Cerrolaza, Juan
Rivas-Santos, Celia
Gallego-Colastra, Leticia
Caparros-Martin, Jose Antonio
Martin-Bravo, Carolina
Martin-Hurtado, Ana
Sánchez-Bellver, Laura
Marfany, Gemma
Ruiz-Perez, Victor L.
Garcia-Gonzalo, Francesc R.
EVC-EVC2 complex stability and ciliary targeting are regulated by modification with ubiquitin and SUMO
title EVC-EVC2 complex stability and ciliary targeting are regulated by modification with ubiquitin and SUMO
title_full EVC-EVC2 complex stability and ciliary targeting are regulated by modification with ubiquitin and SUMO
title_fullStr EVC-EVC2 complex stability and ciliary targeting are regulated by modification with ubiquitin and SUMO
title_full_unstemmed EVC-EVC2 complex stability and ciliary targeting are regulated by modification with ubiquitin and SUMO
title_short EVC-EVC2 complex stability and ciliary targeting are regulated by modification with ubiquitin and SUMO
title_sort evc-evc2 complex stability and ciliary targeting are regulated by modification with ubiquitin and sumo
topic Cell and Developmental Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10413113/
https://www.ncbi.nlm.nih.gov/pubmed/37576597
http://dx.doi.org/10.3389/fcell.2023.1190258
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