Distinct functional roles of β-tubulin isotypes in microtubule arrays of Tetrahymena thermophila, a model single-celled organism.

<h4>Background</h4>The multi-tubulin hypothesis proposes that each tubulin isotype performs a unique role, or subset of roles, in the universe of microtubule function(s). To test this hypothesis, we are investigating the functions of the recently discovered, noncanonical β-like tubulins...

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Autores principales:	Sandra Pucciarelli, Patrizia Ballarini, Daniela Sparvoli, Sabrina Barchetta, Ting Yu, H William Detrich, Cristina Miceli
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2012
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/ee52771e8d9f4484bfeefaf8e1810247
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spelling	oai:doaj.org-article:ee52771e8d9f4484bfeefaf8e18102472021-11-18T07:14:37ZDistinct functional roles of β-tubulin isotypes in microtubule arrays of Tetrahymena thermophila, a model single-celled organism.1932-620310.1371/journal.pone.0039694https://doaj.org/article/ee52771e8d9f4484bfeefaf8e18102472012-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/22745812/?tool=EBIhttps://doaj.org/toc/1932-6203<h4>Background</h4>The multi-tubulin hypothesis proposes that each tubulin isotype performs a unique role, or subset of roles, in the universe of microtubule function(s). To test this hypothesis, we are investigating the functions of the recently discovered, noncanonical β-like tubulins (BLTs) of the ciliate, Tetrahymena thermophila. Tetrahymena forms 17 distinct microtubular structures whose assembly had been thought to be based on single α- and β-isotypes. However, completion of the macronuclear genome sequence of Tetrahymena demonstrated that this ciliate possessed a β-tubulin multigene family: two synonymous genes (BTU1 and BTU2) encode the canonical β-tubulin, BTU2, and six genes (BLT1-6) yield five divergent β-tubulin isotypes. In this report, we examine the structural features and functions of two of the BLTs (BLT1 and BLT4) and compare them to those of BTU2.<h4>Methodology/principal findings</h4>With respect to BTU2, BLT1 and BLT4 had multiple sequence substitutions in their GTP-binding sites, in their interaction surfaces, and in their microtubule-targeting motifs, which together suggest that they have specialized functions. To assess the roles of these tubulins in vivo, we transformed Tetrahymena with expression vectors that direct the synthesis of GFP-tagged versions of the isotypes. We show that GFP-BLT1 and GFP-BLT4 were not detectable in somatic cilia and basal bodies, whereas GFP-BTU2 strongly labeled these structures. During cell division, GFP-BLT1 and GFP-BLT4, but not GFP-BTU2, were incorporated into the microtubule arrays of the macronucleus and into the mitotic apparatus of the micronucleus. GFP-BLT1 also participated in formation of the microtubules of the meiotic apparatus of the micronucleus during conjugation. Partitioning of the isotypes between nuclear and ciliary microtubules was confirmed biochemically.<h4>Conclusion/significance</h4>We conclude that Tetrahymena uses a family of distinct β-tubulin isotypes to construct subsets of functionally different microtubules, a result that provides strong support for the multi-tubulin hypothesis.Sandra PucciarelliPatrizia BallariniDaniela SparvoliSabrina BarchettaTing YuH William DetrichCristina MiceliPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 7, Iss 6, p e39694 (2012)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Sandra Pucciarelli Patrizia Ballarini Daniela Sparvoli Sabrina Barchetta Ting Yu H William Detrich Cristina Miceli Distinct functional roles of β-tubulin isotypes in microtubule arrays of Tetrahymena thermophila, a model single-celled organism.
description	<h4>Background</h4>The multi-tubulin hypothesis proposes that each tubulin isotype performs a unique role, or subset of roles, in the universe of microtubule function(s). To test this hypothesis, we are investigating the functions of the recently discovered, noncanonical β-like tubulins (BLTs) of the ciliate, Tetrahymena thermophila. Tetrahymena forms 17 distinct microtubular structures whose assembly had been thought to be based on single α- and β-isotypes. However, completion of the macronuclear genome sequence of Tetrahymena demonstrated that this ciliate possessed a β-tubulin multigene family: two synonymous genes (BTU1 and BTU2) encode the canonical β-tubulin, BTU2, and six genes (BLT1-6) yield five divergent β-tubulin isotypes. In this report, we examine the structural features and functions of two of the BLTs (BLT1 and BLT4) and compare them to those of BTU2.<h4>Methodology/principal findings</h4>With respect to BTU2, BLT1 and BLT4 had multiple sequence substitutions in their GTP-binding sites, in their interaction surfaces, and in their microtubule-targeting motifs, which together suggest that they have specialized functions. To assess the roles of these tubulins in vivo, we transformed Tetrahymena with expression vectors that direct the synthesis of GFP-tagged versions of the isotypes. We show that GFP-BLT1 and GFP-BLT4 were not detectable in somatic cilia and basal bodies, whereas GFP-BTU2 strongly labeled these structures. During cell division, GFP-BLT1 and GFP-BLT4, but not GFP-BTU2, were incorporated into the microtubule arrays of the macronucleus and into the mitotic apparatus of the micronucleus. GFP-BLT1 also participated in formation of the microtubules of the meiotic apparatus of the micronucleus during conjugation. Partitioning of the isotypes between nuclear and ciliary microtubules was confirmed biochemically.<h4>Conclusion/significance</h4>We conclude that Tetrahymena uses a family of distinct β-tubulin isotypes to construct subsets of functionally different microtubules, a result that provides strong support for the multi-tubulin hypothesis.
format	article
author	Sandra Pucciarelli Patrizia Ballarini Daniela Sparvoli Sabrina Barchetta Ting Yu H William Detrich Cristina Miceli
author_facet	Sandra Pucciarelli Patrizia Ballarini Daniela Sparvoli Sabrina Barchetta Ting Yu H William Detrich Cristina Miceli
author_sort	Sandra Pucciarelli
title	Distinct functional roles of β-tubulin isotypes in microtubule arrays of Tetrahymena thermophila, a model single-celled organism.
title_short	Distinct functional roles of β-tubulin isotypes in microtubule arrays of Tetrahymena thermophila, a model single-celled organism.
title_full	Distinct functional roles of β-tubulin isotypes in microtubule arrays of Tetrahymena thermophila, a model single-celled organism.
title_fullStr	Distinct functional roles of β-tubulin isotypes in microtubule arrays of Tetrahymena thermophila, a model single-celled organism.
title_full_unstemmed	Distinct functional roles of β-tubulin isotypes in microtubule arrays of Tetrahymena thermophila, a model single-celled organism.
title_sort	distinct functional roles of β-tubulin isotypes in microtubule arrays of tetrahymena thermophila, a model single-celled organism.
publisher	Public Library of Science (PLoS)
publishDate	2012
url	https://doaj.org/article/ee52771e8d9f4484bfeefaf8e1810247
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Distinct functional roles of β-tubulin isotypes in microtubule arrays of Tetrahymena thermophila, a model single-celled organism.

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