Publications

• MEC-17 is an alpha-tubulin acetyltransferase.

  Akella J.S., Wloga D., Kim J., Starostina N.G., Lyons-Abbott S., Morrissette N.S., Dougan S.T., Kipreos E.T., Gaertig J.

  In most eukaryotic cells, subsets of microtubules are adapted for specific functions by post-translational modifications (PTMs) of tubulin subunits. Acetylation of the epsilon-amino group of K40 on alpha-tubulin is a conserved PTM on the luminal side of microtubules that was discovered in the flag ... >> More

  In most eukaryotic cells, subsets of microtubules are adapted for specific functions by post-translational modifications (PTMs) of tubulin subunits. Acetylation of the epsilon-amino group of K40 on alpha-tubulin is a conserved PTM on the luminal side of microtubules that was discovered in the flagella of Chlamydomonas reinhardtii. Studies on the significance of microtubule acetylation have been limited by the undefined status of the alpha-tubulin acetyltransferase. Here we show that MEC-17, a protein related to the Gcn5 histone acetyltransferases and required for the function of touch receptor neurons in Caenorhabditis elegans, acts as a K40-specific acetyltransferase for alpha-tubulin. In vitro, MEC-17 exclusively acetylates K40 of alpha-tubulin. Disruption of the Tetrahymena MEC-17 gene phenocopies the K40R alpha-tubulin mutation and makes microtubules more labile. Depletion of MEC-17 in zebrafish produces phenotypes consistent with neuromuscular defects. In C. elegans, MEC-17 and its paralogue W06B11.1 are redundantly required for acetylation of MEC-12 alpha-tubulin, and contribute to the function of touch receptor neurons partly via MEC-12 acetylation and partly via another function, possibly by acetylating another protein. In summary, we identify MEC-17 as an enzyme that acetylates the K40 residue of alpha-tubulin, the only PTM known to occur on the luminal surface of microtubules. << Less

  Nature 467:218-222(2010) [PubMed] [EuropePMC]

• Structure of the alpha-tubulin acetyltransferase, alphaTAT1, and implications for tubulin-specific acetylation.

  Friedmann D.R., Aguilar A., Fan J., Nachury M.V., Marmorstein R.

  Protein acetylation is an important posttranslational modification with the recent identification of new substrates and enzymes, new links to disease, and modulators of protein acetylation for therapy. α-Tubulin acetyltransferase (αTAT1) is the major α-tubulin lysine-40 (K40) acetyltransferase in ... >> More

  Protein acetylation is an important posttranslational modification with the recent identification of new substrates and enzymes, new links to disease, and modulators of protein acetylation for therapy. α-Tubulin acetyltransferase (αTAT1) is the major α-tubulin lysine-40 (K40) acetyltransferase in mammals, nematodes, and protozoa, and its activity plays a conserved role in several microtubule-based processes. Here, we present the X-ray crystal structure of the human αTAT1/acetyl-CoA complex. Together with structure-based mutagenesis, enzymatic analysis, and functional studies in cells, we elucidate the catalytic mechanism and mode of tubulin-specific acetylation. We find that αTAT1 has an overall fold similar to the Gcn5 histone acetyltransferase but contains a relatively wide substrate binding groove and unique structural elements that play important roles in α-tubulin-specific acetylation. Conserved aspartic acid and cysteine residues play important catalytic roles through a ternary complex mechanism. αTAT1 mutations have analogous effects on tubulin acetylation in vitro and in cells, demonstrating that it is the central determining factor of α-tubulin K40 acetylation levels in vivo. Together, these studies provide general insights into distinguishing features between histone and tubulin acetyltransferases, and they have specific implications for understanding the molecular basis of tubulin acetylation and for developing small molecule modulators of microtubule acetylation for therapy. << Less

  Proc. Natl. Acad. Sci. U.S.A. 109:19655-19660(2012) [PubMed] [EuropePMC]

• Molecular basis for age-dependent microtubule acetylation by tubulin acetyltransferase.

  Szyk A., Deaconescu A.M., Spector J., Goodman B., Valenstein M.L., Ziolkowska N.E., Kormendi V., Grigorieff N., Roll-Mecak A.

  Acetylation of α-tubulin Lys40 by tubulin acetyltransferase (TAT) is the only known posttranslational modification in the microtubule lumen. It marks stable microtubules and is required for polarity establishment and directional migration. Here, we elucidate the mechanistic underpinnings for TAT a ... >> More

  Acetylation of α-tubulin Lys40 by tubulin acetyltransferase (TAT) is the only known posttranslational modification in the microtubule lumen. It marks stable microtubules and is required for polarity establishment and directional migration. Here, we elucidate the mechanistic underpinnings for TAT activity and its preference for microtubules with slow turnover. 1.35 Å TAT cocrystal structures with bisubstrate analogs constrain TAT action to the microtubule lumen and reveal Lys40 engaged in a suboptimal active site. Assays with diverse tubulin polymers show that TAT is stimulated by microtubule interprotofilament contacts. Unexpectedly, despite the confined intraluminal location of Lys40, TAT efficiently scans the microtubule bidirectionally and acetylates stochastically without preference for ends. First-principles modeling and single-molecule measurements demonstrate that TAT catalytic activity, not constrained luminal diffusion, is rate limiting for acetylation. Thus, because of its preference for microtubules over free tubulin and its modest catalytic rate, TAT can function as a slow clock for microtubule lifetimes. << Less

  Cell 157:1405-1415(2014) [PubMed] [EuropePMC]

• Luminal localization of alpha-tubulin K40 acetylation by cryo-EM analysis of fab-labeled microtubules.

  Soppina V., Herbstman J.F., Skiniotis G., Verhey K.J.

  The αβ-tubulin subunits of microtubules can undergo a variety of evolutionarily-conserved post-translational modifications (PTMs) that provide functional specialization to subsets of cellular microtubules. Acetylation of α-tubulin residue Lysine-40 (K40) has been correlated with increased microtub ... >> More

  The αβ-tubulin subunits of microtubules can undergo a variety of evolutionarily-conserved post-translational modifications (PTMs) that provide functional specialization to subsets of cellular microtubules. Acetylation of α-tubulin residue Lysine-40 (K40) has been correlated with increased microtubule stability, intracellular transport, and ciliary assembly, yet a mechanistic understanding of how acetylation influences these events is lacking. Using the anti-acetylated tubulin antibody 6-11B-1 and electron cryo-microscopy, we demonstrate that the K40 acetylation site is located inside the microtubule lumen and thus cannot directly influence events on the microtubule surface, including kinesin-1 binding. Surprisingly, the monoclonal 6-11B-1 antibody recognizes both acetylated and deacetylated microtubules. These results suggest that acetylation induces structural changes in the K40-containing loop that could have important functional consequences on microtubule stability, bending, and subunit interactions. This work has important implications for acetylation and deacetylation reaction mechanisms as well as for interpreting experiments based on 6-11B-1 labeling. << Less

  PLoS ONE 7:E48204-E48204(2012) [PubMed] [EuropePMC]

• Alpha-tubulin acetylase activity in isolated Chlamydomonas flagella.

  Greer K., Maruta H., L'Hernault S.W., Rosenbaum J.L.

  We have previously shown that the alpha-tubulin of Chlamydomonas flagella is synthesized as a precursor which is modified by acetylation in the flagellum during flagellar assembly. In this report, we show the presence of an alpha-tubulin acetylase activity in isolated Chlamydomonas flagella that i ... >> More

  We have previously shown that the alpha-tubulin of Chlamydomonas flagella is synthesized as a precursor which is modified by acetylation in the flagellum during flagellar assembly. In this report, we show the presence of an alpha-tubulin acetylase activity in isolated Chlamydomonas flagella that is highly specific for alpha-tubulin of both mammalian brain and Chlamydomonas. << Less

  J Cell Biol 101:2081-2084(1985) [PubMed] [EuropePMC]