Acetylation of lysine 40 in α-tubulin is not essential in Tetrahymena thermophila

Abstract

In Tetrahymena, at least 17 distinct microtubule structures are assembled from a single primary sequence type of α- and β-tubulin heterodimer, precluding distinctions among microtubular systems based on tubulin primary sequence isotypes. Tetrahymena tubulins also are modified by several types of posttranslational reactions including acetylation of α-tubulin at lysine 40, a modification found in most eukaryotes. In Tetrahymena, axonemal α-tubulin and numerous other microtubules are acetylated. We completely replaced the single type of α-tubulin gene in the macronucleus with a version encoding arginine instead of lysine 40 and therefore cannot be acetylated at this position. No acetylated tubulin was detectable in these transformants using a monoclonal antibody specific for acetylated lysine 40. Surprisingly, mutants lacking detectable acetylated tubulin are indistinguishable from wild-type cells. Thus, acetylation of α-tubulin at lysine 40 is nonessential in Tetrahymena. In addition, isoelectric focusing gel analysis of axonemal tubulin from cells unable to acetylate α-tubulin leads us to conclude that: (a) most or all ciliary α-tubulin is acetylated, (b) other lysines cannot be acetylated to compensate for loss of acetylation at lysine 40, and (c) acetylated α-tubulin molecules in wild-type cells contain one or more additional charge-altering modifications.