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TACC3 is a microtubule plus end-tracking protein that promotes axon elongation and also regulates microtubule plus end dynamics in multiple embryonic cell types.

Nwagbara BU, Faris AE, Bearce EA, Erdogan B, Ebbert PT, Evans MF, Rutherford EL, Enzenbacher TB, Lowery LA - Mol. Biol. Cell (2014)

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.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Boston College, Chestnut Hill, MA 02467.

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TACC3 protein is expressed within embryonic neuronal growth cones and promotes axon outgrowth. (A–F) Immunostaining with antibodies to TACC3 (A) and tubulin (B) in cultured embryonic neural crest cells confirm that TACC3 is enriched at the centrosome (C, A′–C′), in addition to being present in puncta throughout the cell. (D–F) TACC3 antibody also strongly stains growth cones. Scale bar, 5 μm. (G) Western blot showing that a morpholino (MO) targeted against TACC3 results in 50% knockdown (KD) by 2 d postfertilization. Bar graph depicts densito­metry of blot shown; however, similar results were obtained with >20 individual Western blots (unpublished data). (H–I) Axon outgrowth parameters after 24 h in culture were quantified in control and TACC3 KD conditions. (H) The numbers of axons per neural tube explant were counted from three independent experiments. (I) The distance from explant to growth cone was measured to calculate the length of axons from three independent experiments. (J–K) Representative phase contrast microscopy images of axons from control and TACC3 KD. (L) Western blot showing that TACC3 levels are increased (overexpressed [OE]) 2 d after injecting embryos with TACC3 mRNA. Bar graph depicts densitometry of blot shown; however, similar results were obtained with >20 individual Western blots (unpublished data). Note that the TACC3 blot in L was exposed for a shorter amount of time than the one in G, which explains the fainter control band in L. (M, N) Axon outgrowth parameters after 18 h in culture were quantified in control and TACC3 OE conditions. (O–P) Representative phase contrast microscopy images of axons from control and TACC3 OE. Box-and-whisker plots indicate the mean (diamond), median, extrema, and quartiles. An unpaired t test was performed to assess significance between conditions. **p < 0.01, ***p < 0.001; n.s., not significant. Bar, 50 μm (J, K, O, P).
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Figure 1: TACC3 protein is expressed within embryonic neuronal growth cones and promotes axon outgrowth. (A–F) Immunostaining with antibodies to TACC3 (A) and tubulin (B) in cultured embryonic neural crest cells confirm that TACC3 is enriched at the centrosome (C, A′–C′), in addition to being present in puncta throughout the cell. (D–F) TACC3 antibody also strongly stains growth cones. Scale bar, 5 μm. (G) Western blot showing that a morpholino (MO) targeted against TACC3 results in 50% knockdown (KD) by 2 d postfertilization. Bar graph depicts densito­metry of blot shown; however, similar results were obtained with >20 individual Western blots (unpublished data). (H–I) Axon outgrowth parameters after 24 h in culture were quantified in control and TACC3 KD conditions. (H) The numbers of axons per neural tube explant were counted from three independent experiments. (I) The distance from explant to growth cone was measured to calculate the length of axons from three independent experiments. (J–K) Representative phase contrast microscopy images of axons from control and TACC3 KD. (L) Western blot showing that TACC3 levels are increased (overexpressed [OE]) 2 d after injecting embryos with TACC3 mRNA. Bar graph depicts densitometry of blot shown; however, similar results were obtained with >20 individual Western blots (unpublished data). Note that the TACC3 blot in L was exposed for a shorter amount of time than the one in G, which explains the fainter control band in L. (M, N) Axon outgrowth parameters after 18 h in culture were quantified in control and TACC3 OE conditions. (O–P) Representative phase contrast microscopy images of axons from control and TACC3 OE. Box-and-whisker plots indicate the mean (diamond), median, extrema, and quartiles. An unpaired t test was performed to assess significance between conditions. **p < 0.01, ***p < 0.001; n.s., not significant. Bar, 50 μm (J, K, O, P).

Mentions: We first performed immunostaining of fixed, cultured embryonic neural crest cells and neuronal growth cones, using an antibody specific to X. laevis TACC3. Within neural crest cells, we observed punctate staining throughout the entire cell (Figure 1, A–C), as well as staining at the centrosome (Figure 1, A′–C′), consistent with previous reports of TACC3 localization (Gergely et al., 2000b; Groisman et al., 2000). We also observed strong labeling within axons and growth cones (Figure 1, D–F), which has not been previously reported, and we confirmed TACC3 presence in neural tissue by Western blot of axon lysates (Figure 1 Supplement). These results suggest that TACC3 may play a role within the growth cone during axon outgrowth.


TACC3 is a microtubule plus end-tracking protein that promotes axon elongation and also regulates microtubule plus end dynamics in multiple embryonic cell types.

Nwagbara BU, Faris AE, Bearce EA, Erdogan B, Ebbert PT, Evans MF, Rutherford EL, Enzenbacher TB, Lowery LA - Mol. Biol. Cell (2014)

TACC3 protein is expressed within embryonic neuronal growth cones and promotes axon outgrowth. (A–F) Immunostaining with antibodies to TACC3 (A) and tubulin (B) in cultured embryonic neural crest cells confirm that TACC3 is enriched at the centrosome (C, A′–C′), in addition to being present in puncta throughout the cell. (D–F) TACC3 antibody also strongly stains growth cones. Scale bar, 5 μm. (G) Western blot showing that a morpholino (MO) targeted against TACC3 results in 50% knockdown (KD) by 2 d postfertilization. Bar graph depicts densito­metry of blot shown; however, similar results were obtained with >20 individual Western blots (unpublished data). (H–I) Axon outgrowth parameters after 24 h in culture were quantified in control and TACC3 KD conditions. (H) The numbers of axons per neural tube explant were counted from three independent experiments. (I) The distance from explant to growth cone was measured to calculate the length of axons from three independent experiments. (J–K) Representative phase contrast microscopy images of axons from control and TACC3 KD. (L) Western blot showing that TACC3 levels are increased (overexpressed [OE]) 2 d after injecting embryos with TACC3 mRNA. Bar graph depicts densitometry of blot shown; however, similar results were obtained with >20 individual Western blots (unpublished data). Note that the TACC3 blot in L was exposed for a shorter amount of time than the one in G, which explains the fainter control band in L. (M, N) Axon outgrowth parameters after 18 h in culture were quantified in control and TACC3 OE conditions. (O–P) Representative phase contrast microscopy images of axons from control and TACC3 OE. Box-and-whisker plots indicate the mean (diamond), median, extrema, and quartiles. An unpaired t test was performed to assess significance between conditions. **p < 0.01, ***p < 0.001; n.s., not significant. Bar, 50 μm (J, K, O, P).
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Related In: Results  -  Collection

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Figure 1: TACC3 protein is expressed within embryonic neuronal growth cones and promotes axon outgrowth. (A–F) Immunostaining with antibodies to TACC3 (A) and tubulin (B) in cultured embryonic neural crest cells confirm that TACC3 is enriched at the centrosome (C, A′–C′), in addition to being present in puncta throughout the cell. (D–F) TACC3 antibody also strongly stains growth cones. Scale bar, 5 μm. (G) Western blot showing that a morpholino (MO) targeted against TACC3 results in 50% knockdown (KD) by 2 d postfertilization. Bar graph depicts densito­metry of blot shown; however, similar results were obtained with >20 individual Western blots (unpublished data). (H–I) Axon outgrowth parameters after 24 h in culture were quantified in control and TACC3 KD conditions. (H) The numbers of axons per neural tube explant were counted from three independent experiments. (I) The distance from explant to growth cone was measured to calculate the length of axons from three independent experiments. (J–K) Representative phase contrast microscopy images of axons from control and TACC3 KD. (L) Western blot showing that TACC3 levels are increased (overexpressed [OE]) 2 d after injecting embryos with TACC3 mRNA. Bar graph depicts densitometry of blot shown; however, similar results were obtained with >20 individual Western blots (unpublished data). Note that the TACC3 blot in L was exposed for a shorter amount of time than the one in G, which explains the fainter control band in L. (M, N) Axon outgrowth parameters after 18 h in culture were quantified in control and TACC3 OE conditions. (O–P) Representative phase contrast microscopy images of axons from control and TACC3 OE. Box-and-whisker plots indicate the mean (diamond), median, extrema, and quartiles. An unpaired t test was performed to assess significance between conditions. **p < 0.01, ***p < 0.001; n.s., not significant. Bar, 50 μm (J, K, O, P).
Mentions: We first performed immunostaining of fixed, cultured embryonic neural crest cells and neuronal growth cones, using an antibody specific to X. laevis TACC3. Within neural crest cells, we observed punctate staining throughout the entire cell (Figure 1, A–C), as well as staining at the centrosome (Figure 1, A′–C′), consistent with previous reports of TACC3 localization (Gergely et al., 2000b; Groisman et al., 2000). We also observed strong labeling within axons and growth cones (Figure 1, D–F), which has not been previously reported, and we confirmed TACC3 presence in neural tissue by Western blot of axon lysates (Figure 1 Supplement). These results suggest that TACC3 may play a role within the growth cone during axon outgrowth.

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.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, Boston College, Chestnut Hill, MA 02467.

Show MeSH
Related in: MedlinePlus