Differential modifications of the C-terminal tails of α-tubulin isoforms and their in vivo importance for kinesin function in Drosophila

Abstract

Microtubules (MTs) are built from -/{beta}-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 - and {beta}-tubulin. However, very little is known about the specific modifications found on the different tubulin isotypes in vivo and the role of these PTMs in MT transport and other cellular processes in vivo. In this study, we found that in Drosophila ovaries, glutamylation of -tubulin isotypes occurred clearly on the C-terminal ends of Tub84B and Tub84D (Tub84B/D). In contrast, the ovarian -tubulin, Tub67C, is not glutamylated. The C-terminal ends of Tub84B/D are glutamylated at several glutamyl side chains in various combinations. Drosophila TTLL5 is required for the mono- and poly-glutamylation of ovarian Tub84B/D and with this for the proper localization of glutamylated microtubules. Similarly, proper Kinesin-1 distribution in the germline also depends on TTLL5 as 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 of TTLL5. Our results demonstrate in vivo roles of TTLL5 in differential glutamylation of -tubulins and point to the in vivo importance of -tubulin glutamylation for Kinesin-1-dependent processes. SummaryO_LI-tubulin glutamylation was established in the C-terminal domain of Drosophila Tub84B and Tub84D (Tub84B/D). Multiple glutamyl residues were pinpointed in this domain. The female germline -tubulin, Tub67C, is, however, not glutamylated. C_LIO_LITTLL5 is required for mono- and poly-glutamylation of Tub84B/D. C_LIO_LITTLL5 is 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. C_LIO_LITTLL5 is needed for the pausing of anterograde trafficking of mitochondria in axons. C_LI