Potential role for a novel AP180-related protein during endocytosis in MDCK cells.
Clathrin assembly protein, AP180, was originally identified as a brain-specific protein localized to the presynaptic junction. AP180 acts to limit vesicle size and maintain a pool of releasable synaptic vesicles during rapid recycling. In this study, we show that polarized epithelial Madin-Darby canine kidney (MDCK) cells express two AP180-related proteins: the ubiquitously expressed 62-kDa clathrin assembly lymphoid myeloid leukemia (CALM, AP180-2) protein and a novel high-molecular-weight homolog that we have named AP180-3. Sequence analysis of AP180-3 expressed in MDCK cells shows high homology to AP180 from rat brain. AP180-3 contains conserved motifs found in brain-specific AP180, including the epsin NH2-terminal homology (ENTH) domain, the binding site for the alpha-subunit of AP-2, and DLL repeats. Our studies show that AP180-3 from MDCK cells forms complexes with AP-2 and clathrin and that membrane recruitment of these complexes is modulated by phosphorylation. We demonstrate by immunohistochemistry that AP180-3 is localized to cytoplasmic vesicles in MDCK cells and is also present in tubule epithelial cells from mouse kidney. We observed by immunodetection that a high-molecular-weight AP180-related protein is expressed in numerous cells in addition to MDCK cells.
Kusner
Linda
L
Dept. of Physiology and Biophysics, Case Western Reserve University School of Medicine, 10900 Euclid Avenue, Cleveland, OH 44106-4970, USA.
Carlin
Cathleen
C
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004285:Dogs; D004705:Endocytosis / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D034041:Monomeric Clathrin Assembly Proteins / Q000032:analysis / Q000235:genetics / Q000502:physiology*; D020033:Protein Isoforms / Q000235:genetics / Q000502:physiology; D051381:Rats; D017386:Sequence Homology, Amino Acid
D034041:Monomeric Clathrin Assembly Proteins; D020033:Protein Isoforms; C439023:clathrin assembly protein AP180