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Assembly of flagellar radial spoke proteins in Chlamydomonas: identification of the axoneme binding domain of radial spoke protein 3
Radial spokes of the eukaryotic flagellum extend from the A tubule of each outer doublet microtubule toward the central pair microtubules. In the paralyzed flagella mutant of Chlamydomonas pf14, a mutation in the gene for one of 17 polypeptides that comprise the radial spokes results in flagella tha...
Formato: | Texto |
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Lenguaje: | English |
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The Rockefeller University Press
1993
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2119812/ https://www.ncbi.nlm.nih.gov/pubmed/8408197 |
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collection | PubMed |
description | Radial spokes of the eukaryotic flagellum extend from the A tubule of each outer doublet microtubule toward the central pair microtubules. In the paralyzed flagella mutant of Chlamydomonas pf14, a mutation in the gene for one of 17 polypeptides that comprise the radial spokes results in flagella that lack all 17 spoke components. The defective gene product, radial spoke protein 3 (RSP3), is, therefore, pivotal to the assembly of the entire spoke and may attach the spoke to the axoneme. We have synthesized RSP3 in vitro and assayed its binding to axonemes from pf14 cells to determine if RSP3 can attach to spokeless axonemes. In vitro, RSP3 binds to pf14 axonemes, but not to wild-type axonemes or microtubules polymerized from purified chick brain tubulin. The sole axoneme binding domain of RSP3 is located within amino acids 1-85 of the 516 amino acid protein; deletion of these amino acids abolishes binding by RSP3. Fusion of amino acids 1-85 or 42-85 to an unrelated protein confers complete or partial binding activity, respectively, to the fusion protein. Transformation of pf14 cells with mutagenized RSP3 genes indicates that amino acids 18-87 of RSP3 are important to its function, but that the carboxy-terminal 140 amino acids can be deleted with little effect on radial spoke assembly or flagellar motility. |
format | Text |
id | pubmed-2119812 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 1993 |
publisher | The Rockefeller University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-21198122008-05-01 Assembly of flagellar radial spoke proteins in Chlamydomonas: identification of the axoneme binding domain of radial spoke protein 3 J Cell Biol Articles Radial spokes of the eukaryotic flagellum extend from the A tubule of each outer doublet microtubule toward the central pair microtubules. In the paralyzed flagella mutant of Chlamydomonas pf14, a mutation in the gene for one of 17 polypeptides that comprise the radial spokes results in flagella that lack all 17 spoke components. The defective gene product, radial spoke protein 3 (RSP3), is, therefore, pivotal to the assembly of the entire spoke and may attach the spoke to the axoneme. We have synthesized RSP3 in vitro and assayed its binding to axonemes from pf14 cells to determine if RSP3 can attach to spokeless axonemes. In vitro, RSP3 binds to pf14 axonemes, but not to wild-type axonemes or microtubules polymerized from purified chick brain tubulin. The sole axoneme binding domain of RSP3 is located within amino acids 1-85 of the 516 amino acid protein; deletion of these amino acids abolishes binding by RSP3. Fusion of amino acids 1-85 or 42-85 to an unrelated protein confers complete or partial binding activity, respectively, to the fusion protein. Transformation of pf14 cells with mutagenized RSP3 genes indicates that amino acids 18-87 of RSP3 are important to its function, but that the carboxy-terminal 140 amino acids can be deleted with little effect on radial spoke assembly or flagellar motility. The Rockefeller University Press 1993-10-01 /pmc/articles/PMC2119812/ /pubmed/8408197 Text en This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 4.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/4.0/). |
spellingShingle | Articles Assembly of flagellar radial spoke proteins in Chlamydomonas: identification of the axoneme binding domain of radial spoke protein 3 |
title | Assembly of flagellar radial spoke proteins in Chlamydomonas: identification of the axoneme binding domain of radial spoke protein 3 |
title_full | Assembly of flagellar radial spoke proteins in Chlamydomonas: identification of the axoneme binding domain of radial spoke protein 3 |
title_fullStr | Assembly of flagellar radial spoke proteins in Chlamydomonas: identification of the axoneme binding domain of radial spoke protein 3 |
title_full_unstemmed | Assembly of flagellar radial spoke proteins in Chlamydomonas: identification of the axoneme binding domain of radial spoke protein 3 |
title_short | Assembly of flagellar radial spoke proteins in Chlamydomonas: identification of the axoneme binding domain of radial spoke protein 3 |
title_sort | assembly of flagellar radial spoke proteins in chlamydomonas: identification of the axoneme binding domain of radial spoke protein 3 |
topic | Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2119812/ https://www.ncbi.nlm.nih.gov/pubmed/8408197 |