Subcellular localization and dynamics of MysPDZ (Myo18A) in live mammalian cells.
MysPDZ is an unconventional myosin belonging to the class XVIII myosin containing a KE (lysine and glutamine)-rich domain and a PDZ domain, which codistributes with actin fibers partially without any canonical actin binding sequence in its myosin head domain. Recently, we reported the identification of a novel isoform of MysPDZ lacking these domains and exhibiting subcellular localization and expression profile different from the original form of MysPDZ. In order to delineate domains directing the subcellular localization of MysPDZ, we performed co-immunoprecipitation experiments and image analyses using mutants of MysPDZ fused with enhanced yellow fluorescent protein. Co-immunoprecipitation analyses showed that MysPDZ can self-associate through its C-terminus coiled-coil domain and the KE-rich domain mediates the interaction with actin. We observed by image analyses that the codistribution with actin fibers and the localization in inner surface of cell membrane of MysPDZ are controlled by the KE-rich domain and the PDZ domain, respectively. Time lapse video microscopy showed that MysPDZ in the cytoplasm moves randomly and rapidly within short range and is allocated to a subcellular compartment without ATP hydrolysis by MysPDZ. This suggests that MysPDZ is a protein which is unlike most unconventional myosins. Our study uncovers a novel role of the KE-rich and PDZ domains in directing subcellular localization and also contributes to a better understanding of functional differences in MysPDZ isoforms.
Mori
Kentaro
K
Department of Cell Biology, Institute of Development, Aging and Cancer, Tohoku University, 4-1, Seiryo-machi, Aoba-ku, Sendai, Miyagi 980-8575, Japan.
Matsuda
Ken-ichi
K
Furusawa
Tadashi
T
Kawata
Mitsuhiro
M
Inoue
Toshiaki
T
Obinata
Masuo
M
D000199:Actins / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D002470:Cell Survival; D036681:Fluorescence Recovery After Photobleaching; D006801:Humans; D047468:Immunoprecipitation; D051379:Mice; D018715:Microscopy, Video; D009154:Mutation / Q000235:genetics; D009218:Myosins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D021381:Protein Transport; D013997:Time Factors
D000199:Actins; C407101:Myo18a protein, mouse; D020033:Protein Isoforms; D009218:Myosins