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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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Enlargement of focal adhesions following microtubule disassembly. Figure shows the same Xenopus fibroblast injected with TAMRA-vinculin, before (A) and after (B) treatment for 3 h with 2,5 μM nocodazole. (Video 8, available at http://www.jcb.org/cgi/content/full/jcb.200105051/DC1.)
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fig1: Enlargement of focal adhesions following microtubule disassembly. Figure shows the same Xenopus fibroblast injected with TAMRA-vinculin, before (A) and after (B) treatment for 3 h with 2,5 μM nocodazole. (Video 8, available at http://www.jcb.org/cgi/content/full/jcb.200105051/DC1.)

Mentions: Major changes in the size and distribution of substrate adhesion sites in fibroblasts occur after the disassembly of microtubules by colchicine or nocodazole (Bershadsky et al., 1996; Enomoto, 1996; Kaverina et al., 1997, 1999). As shown in Fig. 1 , microtubule disassembly by nocodazole resulted in the growth of adhesion sites over a 3-h period, as well as in the characteristic depolarization of cell shape that has been seen in other fibroblast types. Measurement of contact size and number in vinculin-injected cells imaged at zero time and 3 h later confirmed the dramatic shift toward larger and fewer adhesion sites in nocodazole (Fig. 2) . Time-lapse imaging of the same cells revealed the dynamics of adhesion site reorganization toward the depolarized state. Accordingly, adhesion site growth occurred by the enlargement of single adhesions as well as by the fusion of adhesions by sliding (Video 8, available at http://www.jcb.org/cgi/content/full/jcb.200105051/DC1). Both of these effects are attributable to a depolarized equalization and an increase in contractility in the actin cytoskeleton (see Discussion) (Danowski, 1989; Riveline et al., 2001).


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)

Enlargement of focal adhesions following microtubule disassembly. Figure shows the same Xenopus fibroblast injected with TAMRA-vinculin, before (A) and after (B) treatment for 3 h with 2,5 μM nocodazole. (Video 8, available at http://www.jcb.org/cgi/content/full/jcb.200105051/DC1.)
© Copyright Policy
Related In: Results  -  Collection

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

fig1: Enlargement of focal adhesions following microtubule disassembly. Figure shows the same Xenopus fibroblast injected with TAMRA-vinculin, before (A) and after (B) treatment for 3 h with 2,5 μM nocodazole. (Video 8, available at http://www.jcb.org/cgi/content/full/jcb.200105051/DC1.)
Mentions: Major changes in the size and distribution of substrate adhesion sites in fibroblasts occur after the disassembly of microtubules by colchicine or nocodazole (Bershadsky et al., 1996; Enomoto, 1996; Kaverina et al., 1997, 1999). As shown in Fig. 1 , microtubule disassembly by nocodazole resulted in the growth of adhesion sites over a 3-h period, as well as in the characteristic depolarization of cell shape that has been seen in other fibroblast types. Measurement of contact size and number in vinculin-injected cells imaged at zero time and 3 h later confirmed the dramatic shift toward larger and fewer adhesion sites in nocodazole (Fig. 2) . Time-lapse imaging of the same cells revealed the dynamics of adhesion site reorganization toward the depolarized state. Accordingly, adhesion site growth occurred by the enlargement of single adhesions as well as by the fusion of adhesions by sliding (Video 8, available at http://www.jcb.org/cgi/content/full/jcb.200105051/DC1). Both of these effects are attributable to a depolarized equalization and an increase in contractility in the actin cytoskeleton (see Discussion) (Danowski, 1989; Riveline et al., 2001).

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