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MBC in Press, published online ahead of print March 14, 2007
Mol. Biol. Cell 10.1091/mbc.E06-06-0524

A more recent version of this article appeared on May 1, 2007
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Submitted on June 19, 2006
Revised on February 23, 2007
Accepted on March 1, 2007

Kif5B and Kifc1 Interact and Are Required for Motility and Fission of Early Endocytic Vesicles in Mouse Liver

Sangeeta Nath,*{dagger} Eustratios Bananis,*{dagger} Souvik Sarkar,*{dagger} Richard J. Stockert,* Ann O. Sperry,{ddagger} John W. Murray,*{dagger} and Allan W. Wolkoff*{dagger}

{dagger}Department of Anatomy and Structural Biology and *Marion Bessin Liver Research Center, Albert Einstein College of Medicine, Bronx, NY 10461; {ddagger}Department of Anatomy and Cell Biology, Brody School of Medicine at East Carolina University, Greenville, NC 27858

Monitoring Editor: Erika Holzbaur

Early endocytic vesicles loaded with Texas red asialoorosomucoid were prepared from mouse liver. These vesicles bound to microtubules in vitro and upon ATP addition moved bidirectionally, frequently undergoing fission into two daughter vesicles. There was no effect of vanadate (inhibitor of dynein) on motility, while AMP-PNP (kinesin inhibitor) was highly inhibitory. Studies with specific antibodies confirmed that dynein was not associated with these vesicles, and that Kif5B and the minus-end kinesin, Kifc1 mediated their plus- and minus-end motility, respectively. Over 90% of vesicles associated with Kifc1 also contained Kif5B, and inhibition of Kifc1 with antibody resulted in enhancement of plus-end directed motility. There was reduced vesicle fission when either Kifc1 or Kif5B activity was inhibited by antibody, indicating that the opposing forces resulting from activity of both motors are required for fission to occur. Immunoprecipitation of native Kif5B by FLAG antibody following expression of FLAG-Kifc1 in 293T cells indicates that these two motors can interact with each other. Whether they interact directly or through a complex of potential regulatory proteins will need to be clarified in future studies. However, the present studies show that coordinated activity of these kinesins is essential for motility and processing of early endocytic vesicles.

Address correspondence to: Allan W. Wolkoff (wolkoff{at}aecom.yu.edu)






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