Rab GTPases pp 73-83 | Cite as

Analysis of the Interactions Between Rab GTPases and Class V Myosins

  • Andrew J. Lindsay
  • Stéphanie Miserey-Lenkei
  • Bruno GoudEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 1298)


Myosins are actin-based motor proteins that are involved in a wide variety of cellular processes such as membrane transport, muscle contraction, and cell division. Humans have over 40 myosins that can be placed into 18 classes, the malfunctioning of a number of which can lead to disease. There are three members of the human class V myosin family, myosins Va, Vb, and Vc. People lacking functional myosin Va suffer from a rare autosomal recessive disease called Griscelli’s Syndrome type I (GS1) that is characterized by severe neurological defects and partial albinism. Mutations in the myosin Vb gene lead to an epithelial disorder called microvillus inclusion disease (MVID) that is often fatal in infants. The class V myosins have been implicated in the transport of diverse cargoes such as melanosomes in pigment cells, synaptic vesicles in neurons, RNA transcripts in a variety of cell types, and organelles such as the endoplasmic reticulum. The Rab GTPases play a critical role in recruiting class V myosins to their cargo. We recently published a study in which we used the yeast two-hybrid system to systematically test myosin Va for its ability to interact with each member of the human Rab GTPase family. We present here a detailed description of this yeast two-hybrid “living chip” assay. Furthermore, we present a protocol for validating positive interactions obtained from this screen by coimmunoprecipitation.

Key words

Myosin V Rab GTPases Protein–protein interactions Yeast two-hybrid Coimmunoprecipitation 



The work of AJL for this publication was supported by a joint Health Research Board/Marie Curie Mobility Fellowship (MCPD/2009/6) and a Science Foundation Ireland Programme Grant (09/IN1/B2629) to Mary W. McCaffrey, UCC. BG was supported by the Institut Curie, the Centre National de la Recherche Scientifique, and the Agence Nationale pour la Recherche (ANR Grant 2010 BLAN 122902).


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Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Andrew J. Lindsay
    • 1
  • Stéphanie Miserey-Lenkei
    • 2
  • Bruno Goud
    • 2
    Email author
  1. 1.Molecular Cell Biology Laboratory, School of Biochemistry and Cell Biology, Biosciences InstituteUniversity College CorkCorkIreland
  2. 2.Molecular Mechanisms of Intracellular Transport, Unité Mixte de Recherche 144, Centre National de la Recherche ScientifiqueInstitut CurieParisFrance

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