The FAM deubiquitylating enzyme localizes to multiple points of protein trafficking in epithelia, where it associates with E-cadherin and beta-catenin.
Ubiquitylation is a necessary step in the endocytosis and lysosomal trafficking of many plasma membrane proteins and can also influence protein trafficking in the biosynthetic pathway. Although a molecular understanding of ubiquitylation in these processes is beginning to emerge, very little is known about the role deubiquitylation may play. Fat Facets in mouse (FAM) is substrate-specific deubiquitylating enzyme highly expressed in epithelia where it interacts with its substrate, beta-catenin. Here we show, in the polarized intestinal epithelial cell line T84, FAM localized to multiple points of protein trafficking. FAM interacted with beta-catenin and E-cadherin in T84 cells but only in subconfluent cultures. FAM extensively colocalized with beta-catenin in cytoplasmic puncta but not at sites of cell-cell contact as well as immunoprecipitating with beta-catenin and E-cadherin from a higher molecular weight complex ( approximately 500 kDa). At confluence FAM neither colocalized with, nor immunoprecipitated, beta-catenin or E-cadherin, which were predominantly in a larger molecular weight complex ( approximately 2 MDa) at the cell surface. Overexpression of FAM in MCF-7 epithelial cells resulted in increased beta-catenin levels, which localized to the plasma membrane. Expression of E-cadherin in L-cell fibroblasts resulted in the relocalization of FAM from the Golgi to cytoplasmic puncta. These data strongly suggest that FAM associates with E-cadherin and beta-catenin during trafficking to the plasma membrane.
Murray
Rachael Z
RZ
Child Health Research Institute, North Adelaide, SA 5006, Australia.
Jolly
Lachlan A
LA
Wood
Stephen A
SA
D000185:Actinin / Q000378:metabolism; D015820:Cadherins / Q000096:biosynthesis*; D002448:Cell Adhesion; D002460:Cell Line; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D002850:Chromatography, Gel; D003593:Cytoplasm / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000096:biosynthesis* / Q000378:metabolism; D003902:Detergents / Q000494:pharmacology; D010450:Endopeptidases / Q000096:biosynthesis* / Q000502:physiology; D004848:Epithelium / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism; D006801:Humans; D013058:Mass Spectrometry; D008856:Microscopy, Fluorescence; D009218:Myosins / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D011233:Precipitin Tests; D021381:Protein Transport; D015534:Trans-Activators / Q000096:biosynthesis* / Q000378:metabolism; D014162:Transfection; D025801:Ubiquitin / Q000378:metabolism; D043222:Ubiquitin Thiolesterase; D051176:beta Catenin
C495270:CTNNB1 protein, human; D015820:Cadherins; D003598:Cytoskeletal Proteins; D003902:Detergents; D015534:Trans-Activators; D025801:Ubiquitin; D051176:beta Catenin; D000185:Actinin; D010450:Endopeptidases; D043222:Ubiquitin Thiolesterase; C404328:Usp9x protein, mouse; D009218:Myosins