Author:
Li Gabriella,Moore Jeffrey K.
Abstract
AbstractAlthough cold temperatures have long been used to depolymerize microtubules, how temperature specifically affects the polymerization and depolymerization activities of tubulin proteins and how these lead to changes in microtubule networks in cells has not been established. We investigated these questions in budding yeast, an organism found in diverse environments and therefore predicted to exhibit dynamic microtubules across a broad range of temperatures. We measured the dynamics of GFP-labeled microtubules in living cells and found that lowering the temperature from 37°C to 10°C decreased the rates of both polymerization and depolymerization, decreased the amount of polymer assembled before catastrophes and decreased the frequency of microtubule emergence from nucleation sites. Lowering to 4°C caused rapid loss of almost all microtubule polymer. We provide evidence that these effects on microtubule dynamics may be explained in part by changes in the co-factor-dependent conformational dynamics of tubulin proteins. Ablation of tubulin-binding co-factors further sensitizes cells and their microtubules to low temperatures, and we highlight a specific role for TBCB/Alf1 in microtubule maintenance at low temperatures. Finally, we show that inhibiting the maturation cycle of tubulin by using a point mutant in β-tubulin confers hyper-stable microtubules at low temperatures, rescues the requirement for TBCB/Alf1, and improves the cold tolerance of the yeast. Together, these results reveal an unappreciated step in the tubulin cycle in cells and suggest that this step may be a key limiting factor in the thermal tolerance of organisms.
Publisher
Cold Spring Harbor Laboratory
Reference70 articles.
1. Aiken , Jayne , Georgia Buscaglia , A. Sophie Aiken , Jeffrey K. Moore , and Emily A. Bates . 2019. “Tubulin Mutations in Brain Development Disorders: Why Haploinsufficiency Does Not Explain TUBA1A Tubulinopathies.” Cytoskeleton.
2. Genome-Wide Analysis Reveals Novel and Discrete Functions for Tubulin Carboxy-Terminal Tails;Current Biology: CB,2014
3. High-Resolution Microtubule Structures Reveal the Structural Transitions in αβ-Tubulin upon GTP Hydrolysis
4. Amberg , David C. , Daniel J. Burke , and Jeffrey N. Strathern . 2000. Methods in Yeast Genetics: A Cold Spring Harbor Laboratory Course Manual, 2005 Edition. Cold Spring Harbor Laboratory Press.
5. Interactions of tubulin with vinblastine and guanosine triphosphate