Oxidative Stress-Induced Structural Defects within the Microtubule Lattice Induce Tubulin Acetylation by ATAT1

Abstract

Microtubules are dynamic, filamentous polymers of α/β-tubulin heterodimers that form a complex network throughout the cytoplasm. In cells, the microtubule cytoskeleton is subject to numerous post-translational modifications that confer functional specialization. Unlike most post-translational modifications that occur at the tubulin C-terminus, the acetylation of -tubulin (Lys40) occurs within the hollow microtubule lumen. Recent work suggests that structural defects, or ‘holes’ within the microtubule lattice, allow α-tubulin acetyltransferase 1 (αTAT1), the enzyme responsible for microtubule acetylation, to enter the hollow microtubule lumen, where it catalyzes tubulin acetylation. Using cell-free reconstitution experiments, fluorescence microscopy, and transmission electron microscopy, we found that pathological oxidative stress alters microtubule structure by introducing holes and structural defects within the lattice. Further, we found that oxidative stress accelerates acetylation of tubulin subunits within the microtubule lumen, but has little effect on the rate of microtubule detyrosination, which occurs on the external microtubule surface. Our cell-free reconstitution experiments and computational simulations together suggest that it is the location of aTAT1 binding within the microtubule lumen, and the lattice damage caused by oxidative stress, that leads to increased acetylation in oxidative stress-treated microtubules. Consistent with these results, oxidative stress promotes microtubule acetylation, but not detyrosination, in cells. This work provides insight into mechanisms through which pathological oxidative stress biochemically modifies and remodels the microtubule network in cells.