Abstract
Two proteins implicated in inherited deafness, myosin IIIa1, a plus-end-directed motor2, and espin3,4,5,6, an actin-bundling protein containing the actin-monomer-binding motif WH2, have been shown to influence the length of mechanosensory stereocilia7,8. Here we report that espin 1, an ankyrin repeat-containing isoform of espin6, colocalizes with myosin IIIa at stereocilia tips and interacts with a unique conserved domain of myosin IIIa. We show that combined overexpression of these proteins causes greater elongation of stereocilia, compared with overexpression of either myosin IIIa alone or espin 1 alone. When these two proteins were co-expressed in the fibroblast-like COS-7 cell line they induced a tenfold elongation of filopodia. This extraordinary filopodia elongation results from the transport of espin 1 to the plus ends of F-actin by myosin IIIa and depends on espin 1 WH2 activity. This study provides the basis for understanding the role of myosin IIIa and espin 1 in regulating stereocilia length, and presents a physiological example where myosins can boost elongation of actin protrusions by transporting actin regulatory factors to the plus ends of actin filaments.
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Acknowledgements
We thank Chi W. Pak for discussions and for the suggestion of mutations in the WH2 motif, Mark Schneider and Saeeda Latham for initial help with experiments and for discussions related to this work, Martin Horak for advice on cloning procedures, and Ronald Petralia for comments on the manuscript. This work was supported by NIDCD, DIR, NIH and in part by NIH grants to A.C.D. (no. EY003575) and to C.M.Y. (no. EY016419).
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R.C.M. designed probes and experiments, performed the GST pulldowns, cell culture, immunocytochemistry and transfections; F.T.S. performed the dissections, cell and organotypic cultures, immunohistochemistry, transfections and confocal imaging; G.W.D. and A.C.D. designed probes, performed transfections and contributed to the experimental design; U.M. performed image and statistical analyses; A.D.S. characterized antibody and DNA probes; C.M.Y. and J.E.M. generated myosin IIIa kinase dead cDNA, purified and performed kinase and motor activity assays; B.K. performed electron microscopy, designed experiments and analysed the results together with all the other authors. All authors discussed and helped to prepare the manuscript.
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Salles, F., Merritt, R., Manor, U. et al. Myosin IIIa boosts elongation of stereocilia by transporting espin 1 to the plus ends of actin filaments. Nat Cell Biol 11, 443–450 (2009). https://doi.org/10.1038/ncb1851
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DOI: https://doi.org/10.1038/ncb1851
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