Dataset Information

Human ?-Tubulin Isotypes Can Regulate Microtubule Protofilament Number and Stability.

ABSTRACT: Cell biological studies have shown that protofilament number, a fundamental feature of microtubules, can correlate with the expression of different tubulin isotypes. However, it is not known if tubulin isotypes directly control this basic microtubule property. Here, we report high-resolution cryo-EM reconstructions (3.5-3.65 Å) of purified human ?1B/?3 and ?1B/?2B microtubules and find that the ?-tubulin isotype can determine protofilament number. Comparisons of atomic models of 13- and 14-protofilament microtubules reveal how tubulin subunit plasticity, manifested in "accordion-like" distributed structural changes, can accommodate distinct lattice organizations. Furthermore, compared to ?1B/?3 microtubules, ?1B/?2B filaments are more stable to passive disassembly and against depolymerization by MCAK or chTOG, microtubule-associated proteins with distinct mechanisms of action. Mixing tubulin isotypes in different proportions results in microtubules with protofilament numbers and stabilities intermediate to those of isotypically pure filaments. Together, our findings indicate that microtubule protofilament number and stability can be controlled through ?-tubulin isotype composition.

SUBMITTER: Ti SC

PROVIDER: S-EPMC6362463 | biostudies-literature | 2018 Oct

REPOSITORIES: biostudies-literature

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Human β-Tubulin Isotypes Can Regulate Microtubule Protofilament Number and Stability.

Ti Shih-Chieh SC Alushin Gregory M GM Kapoor Tarun M TM

Developmental cell 20180920 2

Cell biological studies have shown that protofilament number, a fundamental feature of microtubules, can correlate with the expression of different tubulin isotypes. However, it is not known if tubulin isotypes directly control this basic microtubule property. Here, we report high-resolution cryo-EM reconstructions (3.5-3.65 Å) of purified human α1B/β3 and α1B/β2B microtubules and find that the β-tubulin isotype can determine protofilament number. Comparisons of atomic models of 13- and 14-proto ...[more]

PMID: 30245156

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Project description:BackgroundThe 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.Methodology/principal findingsWith 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.Conclusion/significanceWe 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.

Dataset Information

Human ?-Tubulin Isotypes Can Regulate Microtubule Protofilament Number and Stability.

Publications

Human β-Tubulin Isotypes Can Regulate Microtubule Protofilament Number and Stability.

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