Main

Most mammalian cells possess two subsets of microtubules: dynamic microtubules with a half-life of 5–10 min, and stable microtubules that have a half-life of hours, and which contain one or more types of post-translationally modified tubulin2. One of these modifications, detyrosination, accumulates in stable microtubules but does not cause microtubule stabilization3,4,5,6,7. For other tubulin modifications, however, the case is less clear.

Hubbert et al.1 did not investigate whether changes in tubulin acetylation alter microtubule stability. To test this, we treated wound-edge, serum-starved NIH 3T3 fibroblasts, which have few stable microtubules6,8, with inhibitors of HDAC6 and used resistance to depolymerization by nocodazole and accumulation of detyrosinated tubulin as assays for increased stable microtubules6,8. Cells treated with trichostatin A (TSA), an inhibitor of HDAC6, showed an increase in microtubule acetylation1 (Fig. 1a, b, insets), but not in the detyrosination of microtubules compared with untreated cells (Fig. 1a, b). Cells treated with sodium butyrate, a deacetylase inhibitor that does not affect HDAC6 activity1, did not increase either acetylation or detyrosination of microtubules (results not shown).

Figure 1: Immunofluorescent images of serum-starved, wounded NIH 3T3 fibroblasts, showing that increased acetylation of tubulin does not stabilize microtubules in these cells.
figure 1

Cells were incubated with trichostatin A (TSA; 5 µM; 4 h; a, c) or without TSA (b, d); cells in d were treated with 10 µM lysophosphatidic acid (LPA). Cells were fixed and immunostained for detyrosinated tubulin6,8 (a, b), acetylated tubulin10 (insets) or bulk tubulin (c, d). a, b, TSA increases microtubule acetylation (insets) but does not increase microtubule detyrosination compared with untreated controls (a, b). c, d, TSA does not increase the number of microtubules that are resistant to nocodazole (c; 10 µM; 30 min), whereas cells treated with LPA have nocodazole-resistant microtubules (d). Arrows show stable, modified microtubules orientated towards the leading edge. Scale bars, 15 µm.

Serum-starved cells treated with TSA did not contain nocodazole-resistant microtubules either (Fig. 1c), in contrast to cells treated with a physiological stimulator of stable microtubules, lysophosphatidic acid (LPA)6,8 (Fig. 1d). LPA-treated cells had more acetylated microtubules (results not shown).

These results indicate that increased tubulin acetylation does not increase levels of stable microtubules; rather, microtubules must be stabilized by other mechanisms (such as capping7) and then these stable microtubules accumulate acetylated tubulin, just as they accumulate detyrosinated tubulin. This is consistent with results showing that tubulin acetylation has no effect on microtubule assembly in vitro9 and that acetylated tubulin is only detectable in long-lived stable microtubules in vivo10.

Hubbert et al. found that HDAC6 overexpression enhances cell motility1. Our results imply that this increase in cell motility is not caused by changes in levels of stable microtubules, but by changes in the acetylation of tubulin (or of an as-yet-unidentified protein). Migrating wound-edge fibroblasts contain stable, post-translationally modified microtubules that are orientated towards the cell's leading edge2,6,8 (Fig. 1d), and these may direct organelles and other important cellular components to the leading edge.

Detyrosinated tubulin seems to have an enhanced affinity for kinesin in vitro11, and could be involved in kinesin-dependent recruitment of intermediate filaments to microtubules12 and in the recycling of endocytic vesicles13. Perhaps acetylation will also turn out to affect the activity of microtubule-associated proteins or motors on microtubules.