The Ras target AF-6 is a substrate of the fam deubiquitinating enzyme.
The Ras target AF-6 has been shown to serve as one of the peripheral components of cell-cell adhesions, and is thought to participate in cell-cell adhesion regulation downstream of Ras. We here purified an AF-6-interacting protein with a molecular mass of approximately 220 kD (p220) to investigate the function of AF-6 at cell-cell adhesions. The peptide sequences of p220 were identical to the amino acid sequences of mouse Fam. Fam is homologous to a deubiquitinating enzyme in Drosophila, the product of the fat facets gene. Recent genetic analyses indicate that the deubiquitinating activity of the fat facets product plays a critical role in controlling the cell fate. We found that Fam accumulated at the cell-cell contact sites of MDCKII cells, but not at free ends of plasma membranes. Fam was partially colocalized with AF-6 and interacted with AF-6 in vivo and in vitro. We also showed that AF-6 was ubiquitinated in intact cells, and that Fam prevented the ubiquitination of AF-6.
Taya
S
S
Division of Signal Transduction, Nara Institute of Science and Technology, Ikoma 630-0101, Japan.
Yamamoto
T
T
Kano
K
K
Kawano
Y
Y
Iwamatsu
A
A
Tsuchiya
T
T
Tanaka
K
K
Kanai-Azuma
M
M
Wood
S A
SA
Mattick
J S
JS
Kaibuchi
K
K
D000818:Animals; D001921:Brain / Q000378:metabolism; D002417:Cattle; D002448:Cell Adhesion / Q000502:physiology*; D002460:Cell Line; D003546:Cysteine Endopeptidases / Q000502:physiology; D029721:Drosophila Proteins; D010450:Endopeptidases / Q000502:physiology*; D005455:Fluorescent Antibody Technique; D011905:Genes, ras / Q000235:genetics*; D007150:Immunohistochemistry; D016547:Kinesins / Q000502:physiology*; D007976:Leupeptins / Q000494:pharmacology; D009097:Multienzyme Complexes / Q000502:physiology; D009218:Myosins / Q000502:physiology*; D046988:Proteasome Endopeptidase Complex; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection / Q000235:genetics; D043222:Ubiquitin Thiolesterase; D014452:Ubiquitins / Q000378:metabolism
C490657:Afdn protein, mouse; D029721:Drosophila Proteins; D007976:Leupeptins; D009097:Multienzyme Complexes; D011993:Recombinant Fusion Proteins; D014452:Ubiquitins; C092747:cno protein, Drosophila; C063261:acetylleucyl-leucyl-norleucinal; D010450:Endopeptidases; D043222:Ubiquitin Thiolesterase; C404328:Usp9x protein, mouse; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D009218:Myosins; D016547:Kinesins