TACC3 is a microtubule plus end-tracking protein that promotes axon elongation and also regulates microtubule plus end dynamics in multiple embryonic cell types.
Bottom Line: Using high-resolution live-imaging data on tagged +TIPs, we show that TACC3 localizes to the extreme microtubule plus end, where it lies distal to the microtubule polymerization marker EB1 and directly overlaps with the microtubule polymerase XMAP215.TACC3 also plays a role in regulating XMAP215 stability and localizing XMAP215 to microtubule plus ends.Taken together, our results implicate TACC3 as a +TIP that functions with XMAP215 to regulate microtubule plus end dynamics.
Affiliation: Department of Biology, Boston College, Chestnut Hill, MA 02467.Show MeSH
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Mentions: Plus end tracking of GFP-TACC3 was not restricted to neural-derived cells, as fibroblasts isolated from mesodermal somite tissue also showed clear plus end tracking (Figure 4, A–C, and Figure 4 Supplemental Movies 1 and 2). We also observed on several occasions that shrinking MTs paused concurrently with obvious accumulation of GFP-TACC3 (Figure 4D). This was most apparent in the mesodermal and neural crest cells, as these had greater numbers of MTs to examine. The plot profiles and distribution of GFP-TACC3 on MT plus ends in fibroblasts were similar to that of the neural cell types, with mean length of GFP-TACC3 plus end accumulation being 0.67 μm (Figure 4, E and F, data measured from 55 MTs). GFP-TACC3 accumulation was detectable on 100% of growing MTs, 70% of paused MTs, and 25% of shrinking MTs (Figure 4G; see also Figure 4 Supplement for an example of GFP-TACC3 on shrinking MT). Measuring fluorescence intensity values of GFP-TACC3 localized at the tips of MTs demonstrated that whereas GFP-TACC3 was observable on one-fourth of shrinking MTs (Figure 4G), mean GFP intensities were ∼40% less on shrinking MTs than on growing MTs. In addition, the maximum intensity of GFP-TACC3 fluorescence on shrinking MTs (106 fluorescence units; top whisker in Figure 4H, box plot) was close to half that of the maximum intensity of fluorescence on growing MTs (172 fluorescence units; Figure 4H), demonstrating that the maximum number of GFP-TACC3 molecules that localize to shrinking MTs is not as great as those that can bind to growing MT plus ends. Thus our results demonstrate that TACC3 can act as a +TIP in multiple vertebrate embryonic cell types and also support the notion that TACC3 may either promote MT polymerization or reduce MT catastrophe.
Affiliation: Department of Biology, Boston College, Chestnut Hill, MA 02467.