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Biochemically validated structural model of the 15‐subunit intraflagellar transport complex IFT‐B

Cilia are ubiquitous eukaryotic organelles impotant for cellular motility, signaling, and sensory reception. Cilium formation requires intraflagellar transport of structural and signaling components and involves 22 different proteins organized into intraflagellar transport (IFT) complexes IFT‐A and...

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Autores principales: Petriman, Narcis A, Loureiro‐López, Marta, Taschner, Michael, Zacharia, Nevin K, Georgieva, Magdalena M, Boegholm, Niels, Wang, Jiaolong, Mourão, André, Russell, Robert B, Andersen, Jens S, Lorentzen, Esben
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9753473/
https://www.ncbi.nlm.nih.gov/pubmed/36354106
http://dx.doi.org/10.15252/embj.2022112440
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author Petriman, Narcis A
Loureiro‐López, Marta
Taschner, Michael
Zacharia, Nevin K
Georgieva, Magdalena M
Boegholm, Niels
Wang, Jiaolong
Mourão, André
Russell, Robert B
Andersen, Jens S
Lorentzen, Esben
author_facet Petriman, Narcis A
Loureiro‐López, Marta
Taschner, Michael
Zacharia, Nevin K
Georgieva, Magdalena M
Boegholm, Niels
Wang, Jiaolong
Mourão, André
Russell, Robert B
Andersen, Jens S
Lorentzen, Esben
author_sort Petriman, Narcis A
collection PubMed
description Cilia are ubiquitous eukaryotic organelles impotant for cellular motility, signaling, and sensory reception. Cilium formation requires intraflagellar transport of structural and signaling components and involves 22 different proteins organized into intraflagellar transport (IFT) complexes IFT‐A and IFT‐B that are transported by molecular motors. The IFT‐B complex constitutes the backbone of polymeric IFT trains carrying cargo between the cilium and the cell body. Currently, high‐resolution structures are only available for smaller IFT‐B subcomplexes leaving > 50% structurally uncharacterized. Here, we used Alphafold to structurally model the 15‐subunit IFT‐B complex. The model was validated using cross‐linking/mass‐spectrometry data on reconstituted IFT‐B complexes, X‐ray scattering in solution, diffraction from crystals as well as site‐directed mutagenesis and protein‐binding assays. The IFT‐B structure reveals an elongated and highly flexible complex consistent with cryo‐electron tomographic reconstructions of IFT trains. The IFT‐B complex organizes into IFT‐B1 and IFT‐B2 parts with binding sites for ciliary cargo and the inactive IFT dynein motor, respectively. Interestingly, our results are consistent with two different binding sites for IFT81/74 on IFT88/70/52/46 suggesting the possibility of different structural architectures for the IFT‐B1 complex. Our data present a structural framework to understand IFT‐B complex assembly, function, and ciliopathy variants.
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spelling pubmed-97534732022-12-23 Biochemically validated structural model of the 15‐subunit intraflagellar transport complex IFT‐B Petriman, Narcis A Loureiro‐López, Marta Taschner, Michael Zacharia, Nevin K Georgieva, Magdalena M Boegholm, Niels Wang, Jiaolong Mourão, André Russell, Robert B Andersen, Jens S Lorentzen, Esben EMBO J Articles Cilia are ubiquitous eukaryotic organelles impotant for cellular motility, signaling, and sensory reception. Cilium formation requires intraflagellar transport of structural and signaling components and involves 22 different proteins organized into intraflagellar transport (IFT) complexes IFT‐A and IFT‐B that are transported by molecular motors. The IFT‐B complex constitutes the backbone of polymeric IFT trains carrying cargo between the cilium and the cell body. Currently, high‐resolution structures are only available for smaller IFT‐B subcomplexes leaving > 50% structurally uncharacterized. Here, we used Alphafold to structurally model the 15‐subunit IFT‐B complex. The model was validated using cross‐linking/mass‐spectrometry data on reconstituted IFT‐B complexes, X‐ray scattering in solution, diffraction from crystals as well as site‐directed mutagenesis and protein‐binding assays. The IFT‐B structure reveals an elongated and highly flexible complex consistent with cryo‐electron tomographic reconstructions of IFT trains. The IFT‐B complex organizes into IFT‐B1 and IFT‐B2 parts with binding sites for ciliary cargo and the inactive IFT dynein motor, respectively. Interestingly, our results are consistent with two different binding sites for IFT81/74 on IFT88/70/52/46 suggesting the possibility of different structural architectures for the IFT‐B1 complex. Our data present a structural framework to understand IFT‐B complex assembly, function, and ciliopathy variants. John Wiley and Sons Inc. 2022-11-10 /pmc/articles/PMC9753473/ /pubmed/36354106 http://dx.doi.org/10.15252/embj.2022112440 Text en © 2022 The Authors. Published under the terms of the CC BY NC ND 4.0 license. https://creativecommons.org/licenses/by-nc-nd/4.0/This is an open access article under the terms of the http://creativecommons.org/licenses/by-nc-nd/4.0/ (https://creativecommons.org/licenses/by-nc-nd/4.0/) License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non‐commercial and no modifications or adaptations are made.
spellingShingle Articles
Petriman, Narcis A
Loureiro‐López, Marta
Taschner, Michael
Zacharia, Nevin K
Georgieva, Magdalena M
Boegholm, Niels
Wang, Jiaolong
Mourão, André
Russell, Robert B
Andersen, Jens S
Lorentzen, Esben
Biochemically validated structural model of the 15‐subunit intraflagellar transport complex IFT‐B
title Biochemically validated structural model of the 15‐subunit intraflagellar transport complex IFT‐B
title_full Biochemically validated structural model of the 15‐subunit intraflagellar transport complex IFT‐B
title_fullStr Biochemically validated structural model of the 15‐subunit intraflagellar transport complex IFT‐B
title_full_unstemmed Biochemically validated structural model of the 15‐subunit intraflagellar transport complex IFT‐B
title_short Biochemically validated structural model of the 15‐subunit intraflagellar transport complex IFT‐B
title_sort biochemically validated structural model of the 15‐subunit intraflagellar transport complex ift‐b
topic Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9753473/
https://www.ncbi.nlm.nih.gov/pubmed/36354106
http://dx.doi.org/10.15252/embj.2022112440
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