Tubulin isotype regulation maintains asymmetric requirement for α-tubulin over β-tubulin

Wethekam, Linnea; Moore, Jeffrey K.

doi:10.1083/jcb.202202102

Cited by 4 publications

(9 citation statements)

References 80 publications

(89 reference statements)

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“…β-tubulin overexpression leads to the formation of insoluble aggregates, in contrast, such aggregates do not form when α-tubulin is overexpressed 48 . In both settings, MT functions are highly disrupted 48 . The TBC-DEG binding site to β-tubulin likely evolved to prevent the formation of insoluble polymers of β-tubulin.…”

Section: Discussionmentioning

confidence: 93%

“…Eukaryotic cells maintain stoichiometric α- and β-tubulin expression, which is critical for MT function and homeostasis 47 . The over-expression of α-tubulin is less toxic than the overexpression of β-tubulin for eukaryotic cells 48 . β-tubulin overexpression leads to the formation of insoluble aggregates, in contrast, such aggregates do not form when α-tubulin is overexpressed 48 .…”

Section: Discussionmentioning

confidence: 99%

“…The over-expression of α-tubulin is less toxic than the overexpression of β-tubulin for eukaryotic cells 48 . β-tubulin overexpression leads to the formation of insoluble aggregates, in contrast, such aggregates do not form when α-tubulin is overexpressed 48 . In both settings, MT functions are highly disrupted 48 .…”

Section: Discussionmentioning

confidence: 99%

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Cryo-EM structures of the tubulin cofactors reveal the molecular basis for the biogenesis of alpha/beta-tubulin

Taheri,

Wang,

Singal

et al. 2024

Preprint

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Microtubule polarity and dynamic polymerization originate from the self-association properties of the a-tubulin heterodimer. For decades, it has remained poorly understood how the tubulin cofactors, TBCD, TBCE, TBCC, and the Arl2 GTPase mediate a-tubulin biogenesis from α- and β-tubulins. Here, we use cryogenic electron microscopy to determine structures of tubulin cofactors bound to αβ-tubulin. These structures show that TBCD, TBCE, and Arl2 form a heterotrimeric cage-like TBC-DEG assembly around the a-tubulin heterodimer. TBCD wraps around Arl2 and almost entirely encircles -tubulin, while TBCE forms a lever arm that anchors along the other end of TBCD and rotates α-tubulin. Structures of the TBC-DEG-αβ-tubulin assemblies bound to TBCC reveal the clockwise rotation of the TBCE lever that twists a-tubulin by pulling its C-terminal tail while TBCD holds -tubulin in place. Altogether, these structures uncover transition states in αβ-tubulin biogenesis, suggesting a vise-like mechanism for the GTP-hydrolysis dependent a-tubulin biogenesis mediated by TBC-DEG and TBCC. These structures provide the first evidence of the critical functions of the tubulin cofactors as enzymes that regulate the invariant organization of αβ-tubulin, by catalyzing α- and β-tubulin assembly, disassembly, and subunit exchange which are crucial for regulating the polymerization capacities of αβ-tubulins into microtubules.

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Section: Discussionmentioning

confidence: 93%

Section: Discussionmentioning

confidence: 99%

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Cryo-EM structures of the tubulin cofactors reveal the molecular basis for the biogenesis of alpha/beta-tubulin

Taheri,

Wang,

Singal

et al. 2024

Preprint

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“…To do this we used purified yeast tubulin to build standard curves of αtubulin. Band intensities of cell lysates from log-phase cells grown in rich media were compared to the standard curve and converted from nanograms of protein to molecules per cell (see Material and Methods; Wethekam & Moore, 2023). In cells where we removed the TUB1 intron, the level of α-tubulin across experiments and biological replicates was more consistent and tended to be less than what we observed in wild-type controls (Figure 1B-D).…”

Section: Tub1 Intron Promotes α-Tubulin Expressionmentioning

confidence: 86%

“…Our previous work showed that cells must maintain excess α-tubulin for efficient mitotic timing and to prevent the accumulation of monomeric β-tubulin (Wethekam & Moore, 2023). Conserved features within coding sequences of α-tubulin genes may provide a point of regulation.…”

Section: Introductionmentioning

confidence: 99%

α-tubulin regulation by 5’ introns inS. cerevisiae

Wethekam

Moore

2023

Preprint

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Across eukaryotic genomes, multiple α and β-tubulin genes require regulation to ensure sufficient production of tubulin heterodimers. Features within these gene families that regulate expression remain underexplored. Here we investigate the role of the 5' intron in regulating α-tubulin expression in S. cerevisiae. We find that the intron in the α-tubulin, TUB1, promotes α-tubulin expression and cell fitness during microtubule stress. The role of the TUB1 intron depends on proximity to the TUB1 promoter and sequence features that are distinct from the intron in the alternative α-tubulin isotype, TUB3. These results lead us to perform a screen to identify genes that act with the TUB1 intron. We identified several genes involved in chromatin remodeling, α/β-tubulin heterodimer assembly, and the spindle assembly checkpoint. We propose a model where the TUB1 intron promotes expression from the chromosomal locus, and that this may represent a conserved mechanism for tubulin regulation under conditions that require high levels of tubulin production.

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α-tubulin regulation by 5′ introns in Saccharomyces cerevisiae

Wethekam,

Moore

2023

Genetics

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Across eukaryotic genomes, multiple α- and β-tubulin genes require regulation to ensure sufficient production of tubulin heterodimers. Features within these gene families that regulate expression remain underexplored. Here we investigate the role of the 5’ intron in regulating α-tubulin expression in S. cerevisiae. We find that the intron in the α-tubulin, TUB1, promotes α-tubulin expression and cell fitness during microtubule stress. The role of the TUB1 intron depends on proximity to the TUB1 promoter and sequence features that are distinct from the intron in the alternative α-tubulin isotype, TUB3. These results lead us to perform a screen to identify genes that act with the TUB1 intron. We identified several genes involved in chromatin remodeling, α/β-tubulin heterodimer assembly, and the spindle assembly checkpoint. We propose a model where the TUB1 intron promotes expression from the chromosomal locus, and that this may represent a conserved mechanism for tubulin regulation under conditions that require high levels of tubulin production.

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Tubulin isotype regulation maintains asymmetric requirement for α-tubulin over β-tubulin

Cited by 4 publications

References 80 publications

Cryo-EM structures of the tubulin cofactors reveal the molecular basis for the biogenesis of alpha/beta-tubulin

Cryo-EM structures of the tubulin cofactors reveal the molecular basis for the biogenesis of alpha/beta-tubulin

α-tubulin regulation by 5’ introns inS. cerevisiae

α-tubulin regulation by 5′ introns in Saccharomyces cerevisiae

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