Rab8-optineurin-myosin VI: analysis of interactions and functions in the secretory pathway.
The small GTPase Rab8 has been shown to regulate polarized membrane trafficking pathways from the TGN to the cell surface. Optineurin is an effector protein of Rab8 and a binding partner of the actin-based motor protein myosin VI. We used various approaches to study the interactions between myosin VI and its binding partners and to analyze their role(s) in intracellular membrane trafficking pathways. In this chapter, we describe the use of the mammalian two-hybrid assay to demonstrate protein-protein interactions and to identify binding sites. We describe a secretion assay that was used in combination with RNA interference technology to analyze the function of myosin VI, optineurin, and Rab8 in exocytic membrane trafficking pathways.
Chibalina
Margarita V
MV
Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom.
Roberts
Rhys C
RC
Arden
Susan D
SD
Kendrick-Jones
John
J
Buss
Folma
F
D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D016466:CHO Cells; D018797:Cell Cycle Proteins; D006224:Cricetinae; D003412:Cricetulus; D006367:HeLa Cells; D006801:Humans; D026901:Membrane Transport Proteins; D051379:Mice; D018995:Myosin Heavy Chains / Q000378:metabolism*; D034741:RNA, Small Interfering; D035582:Transcription Factor TFIIIA / Q000378:metabolism*; D014162:Transfection / Q000379:methods; D020798:Two-Hybrid System Techniques; D020691:rab GTP-Binding Proteins / Q000378:metabolism*
D018797:Cell Cycle Proteins; D026901:Membrane Transport Proteins; C409460:OPTN protein, human; D034741:RNA, Small Interfering; D035582:Transcription Factor TFIIIA; C089785:myosin VI; D000469:Alkaline Phosphatase; D018995:Myosin Heavy Chains; D020691:rab GTP-Binding Proteins