The putative tumor suppressor BIN1 is a short-lived nuclear phosphoprotein, the localization of which is altered in malignant cells.
BIN1 is a putative tumor suppressor that was identified in a genetic screen for polypeptides that interact with the MYC oncoprotein. Using a set of six monoclonal antibodies, we identified and examined biochemical features and localization of cellular BIN1. Epitope mapping indicated that a putative nuclear localization motif and the MYC-binding domain were among the regions recognized by five antibodies. In immunoprecipitation and Western analyses, cellular BIN1 was identified in human and rodent cells as a monomeric phosphoprotein of M(r) approximately 70,000. Pulse-chase experiments showed that BIN1 was short-lived, with a half-life of approximately 2 h. Cell immunofluorescence experiments revealed overlapping but unique nuclear localization patterns distinguished by two different antibodies. In normal cells, BIN1 was predominantly nucleoplasmic but was also present in a subnuclear compartment. Conversely, in a panel of tumor cells that expressed BIN1, the predominant localization was the subnuclear compartment. Taken together, the results suggested that the antibodies recognized different isoforms or conformations of BIN1, the localization of which varied between normal and tumor cells. This study will facilitate further analysis of the structure and regulation of BIN1 in normal and malignant cells.
Wechsler-Reya
R
R
The Wistar Institute, Philadelphia, Pennsylvania 19104, USA.
Elliott
K
K
Herlyn
M
M
Prendergast
G C
GC
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D000911:Antibodies, Monoclonal; D019556:COS Cells; D002352:Carrier Proteins / Q000032:analysis / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D003001:Cloning, Molecular; D019084:Fluorescent Antibody Technique, Indirect; D016147:Genes, Tumor Suppressor / Q000276:immunology / Q000502:physiology; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D009369:Neoplasms / Q000378:metabolism*; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins / Q000032:analysis / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins
D048868:Adaptor Proteins, Signal Transducing; D000911:Antibodies, Monoclonal; C101410:BIN1 protein, human; C109221:Bin1 protein, mouse; D002352:Carrier Proteins; D009419:Nerve Tissue Proteins; D009687:Nuclear Proteins; D011993:Recombinant Fusion Proteins; D025521:Tumor Suppressor Proteins