Differential modification of the C-terminal tails of different α-tubulins and their importance for microtubule functionin vivo

Abstract

AbstractMicrotubules (MTs) are built from α-/β-tubulin dimers and used as tracks by kinesin and dynein motors to transport a variety of cargos, such as mRNAs, proteins, and organelles, within the cell. Tubulins are subjected to several post-translational modifications (PTMs). Glutamylation is one of them, and it is responsible for adding one or more glutamic acid residues as branched peptide chains to the C-terminal tails of both α- and β-tubulin. However, very little is known about the specific modifications found on the different tubulin isotypesin vivoand the role of these PTMs in MT transport and other cellular processesin vivo. In this study, we found that inDrosophilaovaries, glutamylation of α-tubulin isotypes occurred clearly on the C-terminal ends of αTub84B and αTub84D (αTub84B/D). In contrast, the ovarian α-tubulin, αTub67C, is not glutamylated. The C-terminal ends of αTub84B/D are glutamylated at several glutamyl side chains in various combinations.Drosophila TTLL5is required for the mono- and poly-glutamylation of ovarian αTub84B/D and with this for the proper localization of glutamylated microtubules. Similarly, proper Kinesin-1 distribution in the germline also depends onTTLL5as well as the refining of Staufen localization and the normal fast ooplasmic streaming with its directional movement, two processes known to depend on Kinesin-1 activities. In the nervous system, the pausing of anterograde axonal transport of mitochondria is affected by an enzymatically dead mutant ofTTLL5. Our results demonstratein vivoroles ofTTLL5in differential glutamylation of α-tubulins and point to thein vivoimportance of α-tubulin glutamylation for Kinesin-1-dependent processes.Summaryα-tubulin glutamylation was established in the C-terminal domain ofDrosophilaαTub84B and αTub84D (αTub84B/D). Multiple glutamyl residues were pinpointed in this domain. The female germline α-tubulin, αTub67C, is, however, not glutamylated.TTLL5is required for mono- and poly-glutamylation of αTub84B/D.TTLL5is required for the proper Kinesin heavy chain (Khc) distribution in the germline, and the kinesin-based refinement of Staufen localization and ooplasmic streaming during late oogenesis.TTLL5is needed for the pausing of anterograde trafficking of mitochondria in axons.