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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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Inactivation of dynein by the m74–2 intermediate chain antibody, as monitored by the redistribution of lysosomes to the cell periphery. Lysosomes were loaded with rhodamine-dextran. (A and C) Paired phase contrast (A) and fluorescence (C) images of Xenopus fibroblast taken just after antibody injection. (B and D) The same cell 24 min later (Video 1, available at http://www.jcb.org/cgi/content/full/jcb.200105051/DC1).
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fig3: Inactivation of dynein by the m74–2 intermediate chain antibody, as monitored by the redistribution of lysosomes to the cell periphery. Lysosomes were loaded with rhodamine-dextran. (A and C) Paired phase contrast (A) and fluorescence (C) images of Xenopus fibroblast taken just after antibody injection. (B and D) The same cell 24 min later (Video 1, available at http://www.jcb.org/cgi/content/full/jcb.200105051/DC1).

Mentions: To investigate the possible role of dynein in microtubule-linked signaling to adhesion sites, we used two methods to block dynein–cargo interactions: (a) injection with a function-blocking antibody against the dynein intermediate chain (Steffen et al., 1997); and (b) transfection with dynamitin (Burkhardt et al., 1997). An indicator of a block in dynein-linked activity is the dispersion of lysosomes to the cell periphery (Burkhardt et al., 1997). As shown in Fig. 3 (Video 1, available at http://www.jcb.org/cgi/content/full/jcb.200105051/DC1), injection of cells with the m74–2 anti-dynein antibody caused the efficient scattering of rhodamine dextran–loaded lysosomes away from the centrosomal region, consistent with a block of dynein activity in the Xenopus cells. This effect was observed in all injected cells. Under the same conditions, no change in the general distribution or dynamics of microtubules was observed in the antibody-injected cells, as compared with controls (unpublished data).


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)

Inactivation of dynein by the m74–2 intermediate chain antibody, as monitored by the redistribution of lysosomes to the cell periphery. Lysosomes were loaded with rhodamine-dextran. (A and C) Paired phase contrast (A) and fluorescence (C) images of Xenopus fibroblast taken just after antibody injection. (B and D) The same cell 24 min later (Video 1, 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

fig3: Inactivation of dynein by the m74–2 intermediate chain antibody, as monitored by the redistribution of lysosomes to the cell periphery. Lysosomes were loaded with rhodamine-dextran. (A and C) Paired phase contrast (A) and fluorescence (C) images of Xenopus fibroblast taken just after antibody injection. (B and D) The same cell 24 min later (Video 1, available at http://www.jcb.org/cgi/content/full/jcb.200105051/DC1).
Mentions: To investigate the possible role of dynein in microtubule-linked signaling to adhesion sites, we used two methods to block dynein–cargo interactions: (a) injection with a function-blocking antibody against the dynein intermediate chain (Steffen et al., 1997); and (b) transfection with dynamitin (Burkhardt et al., 1997). An indicator of a block in dynein-linked activity is the dispersion of lysosomes to the cell periphery (Burkhardt et al., 1997). As shown in Fig. 3 (Video 1, available at http://www.jcb.org/cgi/content/full/jcb.200105051/DC1), injection of cells with the m74–2 anti-dynein antibody caused the efficient scattering of rhodamine dextran–loaded lysosomes away from the centrosomal region, consistent with a block of dynein activity in the Xenopus cells. This effect was observed in all injected cells. Under the same conditions, no change in the general distribution or dynamics of microtubules was observed in the antibody-injected cells, as compared with controls (unpublished data).

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