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Modulation of substrate adhesion dynamics via microtubule targeting requires kinesin-1.

Krylyshkina O, Kaverina I, Kranewitter W, Steffen W, Alonso MC, Cross RA, Small JV - J. Cell Biol. (2002)

Bottom Line: Small. 1999.Cell Biol. 146:1033-1043).In comparison, a block of kinesin-1 activity, either via microinjection of the SUK-4 antibody or of a kinesin-1 heavy chain construct mutated in the motor domain, induced a dramatic increase in the size and reduction in number of substrate adhesions, mimicking the effect observed after microtubule disruption by nocodazole.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Biology, Austrian Academy of Sciences, Billrothsthstrasse 11, Salzburg 5020, Austria.

ABSTRACT
Recent studies have shown that the targeting of substrate adhesions by microtubules promotes adhesion site disassembly (Kaverina, I., O. Krylyshkina, and J.V. Small. 1999. J. Cell Biol. 146:1033-1043). It was accordingly suggested that microtubules serve to convey a signal to adhesion sites to modulate their turnover. Because microtubule motors would be the most likely candidates for effecting signal transmission, we have investigated the consequence of blocking microtubule motor activity on adhesion site dynamics. Using a function-blocking antibody as well as dynamitin overexpression, we found that a block in dynein-cargo interaction induced no change in adhesion site dynamics in Xenopus fibroblasts. In comparison, a block of kinesin-1 activity, either via microinjection of the SUK-4 antibody or of a kinesin-1 heavy chain construct mutated in the motor domain, induced a dramatic increase in the size and reduction in number of substrate adhesions, mimicking the effect observed after microtubule disruption by nocodazole. Blockage of kinesin activity had no influence on either the ability of microtubules to target substrate adhesions or on microtubule polymerisation dynamics. We conclude that conventional kinesin is not required for the guidance of microtubules into substrate adhesions, but is required for the focal delivery of a component(s) that retards their growth or promotes their disassembly.

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Microtubules distribution in Xenopus fibroblasts is unaffected by kinesin inhibition. Cells were preinjected with Cy-3–tubulin (control, middle) and additionally injected with 20 μM T93N (left) or SUK-4 antibody (right).
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fig8: Microtubules distribution in Xenopus fibroblasts is unaffected by kinesin inhibition. Cells were preinjected with Cy-3–tubulin (control, middle) and additionally injected with 20 μM T93N (left) or SUK-4 antibody (right).

Mentions: For cells transfected with GFP-tubulin, we could first show that the general distribution of microtubules in Xenopus fibroblasts was unaffected by injection of the SUK-4 antibody or 20 μM T93N kinesin (Fig. 8) . At higher concentrations of T93N kinesin (100 μM) we observed bundling of microtubules, as was also seen in cells strongly expressing the GFP-tagged T93N construct (unpublished data); hence, the choice of the lower concentration for the kinesin inhibition experiments. Table I presents the results of measurements of microtubule dynamics for control and injected cells transfected with GFP-tubulin. As shown, injection with SUK-4 Ab or 20 μM T93N kinesin had no measurable influence on microtubule dynamics, the values for control and injected cells being comparable to those documented for other cell types (Shelden and Wadsworth, 1993).


Modulation of substrate adhesion dynamics via microtubule targeting requires kinesin-1.

Krylyshkina O, Kaverina I, Kranewitter W, Steffen W, Alonso MC, Cross RA, Small JV - J. Cell Biol. (2002)

Microtubules distribution in Xenopus fibroblasts is unaffected by kinesin inhibition. Cells were preinjected with Cy-3–tubulin (control, middle) and additionally injected with 20 μM T93N (left) or SUK-4 antibody (right).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2199234&req=5

fig8: Microtubules distribution in Xenopus fibroblasts is unaffected by kinesin inhibition. Cells were preinjected with Cy-3–tubulin (control, middle) and additionally injected with 20 μM T93N (left) or SUK-4 antibody (right).
Mentions: For cells transfected with GFP-tubulin, we could first show that the general distribution of microtubules in Xenopus fibroblasts was unaffected by injection of the SUK-4 antibody or 20 μM T93N kinesin (Fig. 8) . At higher concentrations of T93N kinesin (100 μM) we observed bundling of microtubules, as was also seen in cells strongly expressing the GFP-tagged T93N construct (unpublished data); hence, the choice of the lower concentration for the kinesin inhibition experiments. Table I presents the results of measurements of microtubule dynamics for control and injected cells transfected with GFP-tubulin. As shown, injection with SUK-4 Ab or 20 μM T93N kinesin had no measurable influence on microtubule dynamics, the values for control and injected cells being comparable to those documented for other cell types (Shelden and Wadsworth, 1993).

Bottom Line: Small. 1999.Cell Biol. 146:1033-1043).In comparison, a block of kinesin-1 activity, either via microinjection of the SUK-4 antibody or of a kinesin-1 heavy chain construct mutated in the motor domain, induced a dramatic increase in the size and reduction in number of substrate adhesions, mimicking the effect observed after microtubule disruption by nocodazole.

View Article: PubMed Central - PubMed

Affiliation: Institute of Molecular Biology, Austrian Academy of Sciences, Billrothsthstrasse 11, Salzburg 5020, Austria.

ABSTRACT
Recent studies have shown that the targeting of substrate adhesions by microtubules promotes adhesion site disassembly (Kaverina, I., O. Krylyshkina, and J.V. Small. 1999. J. Cell Biol. 146:1033-1043). It was accordingly suggested that microtubules serve to convey a signal to adhesion sites to modulate their turnover. Because microtubule motors would be the most likely candidates for effecting signal transmission, we have investigated the consequence of blocking microtubule motor activity on adhesion site dynamics. Using a function-blocking antibody as well as dynamitin overexpression, we found that a block in dynein-cargo interaction induced no change in adhesion site dynamics in Xenopus fibroblasts. In comparison, a block of kinesin-1 activity, either via microinjection of the SUK-4 antibody or of a kinesin-1 heavy chain construct mutated in the motor domain, induced a dramatic increase in the size and reduction in number of substrate adhesions, mimicking the effect observed after microtubule disruption by nocodazole. Blockage of kinesin activity had no influence on either the ability of microtubules to target substrate adhesions or on microtubule polymerisation dynamics. We conclude that conventional kinesin is not required for the guidance of microtubules into substrate adhesions, but is required for the focal delivery of a component(s) that retards their growth or promotes their disassembly.

Show MeSH
Related in: MedlinePlus