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Ndel1 disfavors dynein–dynactin–adaptor complex formation in two distinct ways
Dynein is the primary minus-end-directed microtubule motor protein. To achieve activation, dynein binds to the dynactin complex and an adaptor to form the “activated dynein complex.” The protein Lis1 aids activation by binding to dynein and promoting its association with dynactin and the adaptor. Nd...
Autores principales: | , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Society for Biochemistry and Molecular Biology
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10248797/ https://www.ncbi.nlm.nih.gov/pubmed/37086789 http://dx.doi.org/10.1016/j.jbc.2023.104735 |
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author | Garrott, Sharon R. Gillies, John P. Siva, Aravintha Little, Saffron R. El Jbeily, Rita DeSantis, Morgan E. |
author_facet | Garrott, Sharon R. Gillies, John P. Siva, Aravintha Little, Saffron R. El Jbeily, Rita DeSantis, Morgan E. |
author_sort | Garrott, Sharon R. |
collection | PubMed |
description | Dynein is the primary minus-end-directed microtubule motor protein. To achieve activation, dynein binds to the dynactin complex and an adaptor to form the “activated dynein complex.” The protein Lis1 aids activation by binding to dynein and promoting its association with dynactin and the adaptor. Ndel1 and its paralog Nde1 are dynein- and Lis1-binding proteins that help control dynein localization within the cell. Cell-based assays suggest that Ndel1–Nde1 also work with Lis1 to promote dynein activation, although the underlying mechanism is unclear. Using purified proteins and quantitative binding assays, here we found that the C-terminal region of Ndel1 contributes to dynein binding and negatively regulates binding to Lis1. Using single-molecule imaging and protein biochemistry, we observed that Ndel1 inhibits dynein activation in two distinct ways. First, Ndel1 disfavors the formation of the activated dynein complex. We found that phosphomimetic mutations in the C-terminal domain of Ndel1 increase its ability to inhibit dynein–dynactin–adaptor complex formation. Second, we observed that Ndel1 interacts with dynein and Lis1 simultaneously and sequesters Lis1 away from its dynein-binding site. In doing this, Ndel1 prevents Lis1-mediated dynein activation. Together, our work suggests that in vitro, Ndel1 is a negative regulator of dynein activation, which contrasts with cellular studies where Ndel1 promotes dynein activity. To reconcile our findings with previous work, we posit that Ndel1 functions to scaffold dynein and Lis1 together while keeping dynein in an inhibited state. We speculate that Ndel1 release can be triggered in cellular settings to allow for timed dynein activation. |
format | Online Article Text |
id | pubmed-10248797 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | American Society for Biochemistry and Molecular Biology |
record_format | MEDLINE/PubMed |
spelling | pubmed-102487972023-06-09 Ndel1 disfavors dynein–dynactin–adaptor complex formation in two distinct ways Garrott, Sharon R. Gillies, John P. Siva, Aravintha Little, Saffron R. El Jbeily, Rita DeSantis, Morgan E. J Biol Chem Research Article Dynein is the primary minus-end-directed microtubule motor protein. To achieve activation, dynein binds to the dynactin complex and an adaptor to form the “activated dynein complex.” The protein Lis1 aids activation by binding to dynein and promoting its association with dynactin and the adaptor. Ndel1 and its paralog Nde1 are dynein- and Lis1-binding proteins that help control dynein localization within the cell. Cell-based assays suggest that Ndel1–Nde1 also work with Lis1 to promote dynein activation, although the underlying mechanism is unclear. Using purified proteins and quantitative binding assays, here we found that the C-terminal region of Ndel1 contributes to dynein binding and negatively regulates binding to Lis1. Using single-molecule imaging and protein biochemistry, we observed that Ndel1 inhibits dynein activation in two distinct ways. First, Ndel1 disfavors the formation of the activated dynein complex. We found that phosphomimetic mutations in the C-terminal domain of Ndel1 increase its ability to inhibit dynein–dynactin–adaptor complex formation. Second, we observed that Ndel1 interacts with dynein and Lis1 simultaneously and sequesters Lis1 away from its dynein-binding site. In doing this, Ndel1 prevents Lis1-mediated dynein activation. Together, our work suggests that in vitro, Ndel1 is a negative regulator of dynein activation, which contrasts with cellular studies where Ndel1 promotes dynein activity. To reconcile our findings with previous work, we posit that Ndel1 functions to scaffold dynein and Lis1 together while keeping dynein in an inhibited state. We speculate that Ndel1 release can be triggered in cellular settings to allow for timed dynein activation. American Society for Biochemistry and Molecular Biology 2023-04-21 /pmc/articles/PMC10248797/ /pubmed/37086789 http://dx.doi.org/10.1016/j.jbc.2023.104735 Text en © 2023 The Authors https://creativecommons.org/licenses/by/4.0/This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Research Article Garrott, Sharon R. Gillies, John P. Siva, Aravintha Little, Saffron R. El Jbeily, Rita DeSantis, Morgan E. Ndel1 disfavors dynein–dynactin–adaptor complex formation in two distinct ways |
title | Ndel1 disfavors dynein–dynactin–adaptor complex formation in two distinct ways |
title_full | Ndel1 disfavors dynein–dynactin–adaptor complex formation in two distinct ways |
title_fullStr | Ndel1 disfavors dynein–dynactin–adaptor complex formation in two distinct ways |
title_full_unstemmed | Ndel1 disfavors dynein–dynactin–adaptor complex formation in two distinct ways |
title_short | Ndel1 disfavors dynein–dynactin–adaptor complex formation in two distinct ways |
title_sort | ndel1 disfavors dynein–dynactin–adaptor complex formation in two distinct ways |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10248797/ https://www.ncbi.nlm.nih.gov/pubmed/37086789 http://dx.doi.org/10.1016/j.jbc.2023.104735 |
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