Murine CENPF interacts with syntaxin 4 in the regulation of vesicular transport.
Syntaxin 4 is a component of the SNARE complex that regulates membrane docking and fusion. Using a yeast two-hybrid screen, we identify a novel interaction between syntaxin 4 and cytoplasmic murine CENPF, a protein previously demonstrated to associate with the microtubule network and SNAP-25. The binding domain for syntaxin 4 in CENPF was defined by yeast two-hybrid assay and co-immunoprecipitation. Confocal analyses in cell culture reveal a high degree of colocalization between endogenously expressed proteins in interphase cells. Additionally, the endogenous SNARE proteins can be isolated as a complex with CENPF in immunoprecipitation experiments. Further analyses demonstrate that murine CENPF and syntaxin 4 colocalize with components of plasma membrane recycling: SNAP-25 and VAMP2. Depletion of endogenous CENPF disrupts GLUT4 trafficking whereas expression of a dominant-negative form of CENPF inhibits cell coupling. Taken together, these studies demonstrate that CENPF provides a direct link between proteins of the SNARE system and the microtubule network and indicate a diverse role for murine CENPF in vesicular transport.
Pooley
Ryan D
RD
Stahlman Cardiovascular Research Laboratories, Program for Developmental Biology, and Department of Medicine, Vanderbilt University Medical Center, Nashville, TN 37232-6300, USA.
Moynihan
Katherine L
KL
Soukoulis
Victor
V
Reddy
Samyukta
S
Francis
Richard
R
Lo
Cecilia
C
Ma
Li-Jun
LJ
Bader
David M
DM
D000818:Animals; D001692:Biological Transport / Q000502:physiology; D019556:COS Cells; D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism; D002522:Chlorocebus aethiops; D002868:Chromosomal Proteins, Non-Histone / Q000235:genetics / Q000378:metabolism*; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism*; D051275:Glucose Transporter Type 4 / Q000235:genetics / Q000378:metabolism; D051379:Mice; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism*; D008870:Microtubules / Q000235:genetics / Q000378:metabolism; D041681:NIH 3T3 Cells; D050765:Qa-SNARE Proteins / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics; D050825:Synaptosomal-Associated Protein 25 / Q000235:genetics / Q000378:metabolism; D022161:Transport Vesicles / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques; D050988:Vesicle-Associated Membrane Protein 2 / Q000235:genetics / Q000378:metabolism
D002868:Chromosomal Proteins, Non-Histone; D051275:Glucose Transporter Type 4; D008840:Microfilament Proteins; D050765:Qa-SNARE Proteins; C495693:Slc2a4 protein, mouse; D050825:Synaptosomal-Associated Protein 25; D050988:Vesicle-Associated Membrane Protein 2; C094772:centromere protein F; C494808:vesicle-associated membrane protein 2, mouse