Chemical-genetic inhibition of a sensitized mutant myosin Vb demonstrates a role in peripheral-pericentriolar membrane traffic

^*McLaughlin Research Institute, Great Falls, MT 59405; ^†Universidade Federal do Estado do Rio de Janeiro, Universidade Estácio de Sá, Universidade Castelo Branco, CEP 22290-280, Rio de Janeiro, Brazil; ^‡Department of Cellular and Molecular Pharmacology, University of California, San Francisco, CA 94143; ^§Departments of Surgery and Cell and Developmental Biology, Vanderbilt University School of Medicine and Nashville Veterans Affairs Medical Center, Nashville, TN 37235; ^¶Genomics Institute of the Novartis Research Foundation, San Diego, CA 92121; and ^∥Oregon Hearing Research Center, Oregon Health and Science University, and Vollum Institute, Portland, OR 97239

Edited by James A. Spudich, Stanford University School of Medicine, Stanford, CA, and approved December 10, 2003 (received for review September 12, 2003)

Abstract

Selective, in situ inhibition of individual unconventional myosins is a powerful approach to determine their specific physiological functions. Here, we report the engineering of a myosin Vb mutant that still hydrolyzes ATP, yet is selectively sensitized to an N ⁶-substituted ADP analog that inhibits its activity, causing it to remain tightly bound to actin. Inhibition of the sensitized mutant causes inhibition of accumulation of transferrin in the cytoplasm and increases levels of plasma-membrane transferrin receptor, suggesting that myosin Vb functions in traffic between peripheral and pericentrosomal compartments.

Footnotes

↵ ** To whom correspondence should be addressed at: McLaughlin Research Institute, 1520 23rd Street South, Great Falls, MT 59405. E-mail: umbjm{at}montana.edu.
This paper was submitted directly (Track II) to the PNAS office.
Abbreviations: PE-ADP, N ⁶-(2-phenylethyl)-ADP; TfR, transferrin receptor.