title: Overexpression of PREP-1 in F9 teratocarcinoma cells leads to a functionally relevant increase of PBX-2 by preventing its degradation.
abstract: To bind DNA and to be retained in the nucleus, PBX proteins must form heterodimeric complexes with members of the MEINOX family. Therefore the balance between PBX and MEINOX must be an important regulatory feature. We show that overexpression of PREP-1 influences the level of PBX-2 protein maintaining the PREP-1-PBX balance. This effect has important functional consequences. F9 teratocarcinoma cells stably transfected with PREP-1 had an increased DNA binding activity to a PREP-PBX-responsive element. Because PREP-1 binds DNA efficiently only when dimerized to PBX, the increased DNA binding activity suggests that the level of PBX might also have increased. Indeed PREP-1-overexpressing cells had a higher level of PBX-2 and PBX-1b proteins. PBX-2 increase did not depend on increased mRNA level or a higher rate of translation but rather because of a protein stabilization process. Indeed, PBX-2 level drastically decreased after 3 h of cycloheximide treatment in control but not in PREP-1-overexpressing cells and the proteasome inhibitor MG132 prevented PBX-2 decay in control cells. Hence, dimerization with PREP-1 appears to decrease proteasomal degradation of PBX-2. Retinoic acid induces differentiation of F9 teratocarcinoma cells with a cascade synthesis of HOX proteins. In PREP-1-overexpressing cells, HOXb1 induction was more sustained (3 days versus 1 day) and the induced level of MEIS-1b, another TALE (three amino acid loop extension) protein involved in embryonal development, was higher. Thus an increase in PREP-1 leads to changes in the fate-determining HOXb1 and has therefore important functional consequences.
authors.affiliation: Molecular Genetics Unit, Department of Molecular Biology and Functional Genomics, Università Vita Salute San Raffaele and DIBIT, via Olgettina 58, 20132 Milan, Italy.
authors.forename: Elena
authors.initials: E
authors.lastname: Longobardi
authors.forename: Francesco
authors.initials: F
authors.lastname: Blasi
mesh_terms: D000595:Amino Acid Sequence; D015152:Blotting, Northern; D002454:Cell Differentiation; D002467:Cell Nucleus / Q000378:metabolism; D003513:Cycloheximide / Q000494:pharmacology; D003546:Cysteine Endopeptidases / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D004247:DNA / Q000378:metabolism; D019281:Dimerization; D018398:Homeodomain Proteins / Q000096:biosynthesis* / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology*; D015151:Immunoblotting; D007700:Kinetics; D007976:Leupeptins / Q000494:pharmacology; D008954:Models, Biological; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors / Q000378:metabolism; D000075064:Myeloid Ecotropic Viral Integration Site 1 Protein; D009363:Neoplasm Proteins / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex; D011485:Protein Binding; D014176:Protein Biosynthesis; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis* / Q000737:chemistry*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018243:Teratocarcinoma / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection; D014212:Tretinoin / Q000494:pharmacology; D014407:Tumor Cells, Cultured
chemical_list: D018398:Homeodomain Proteins; D007976:Leupeptins; D009097:Multienzyme Complexes; D000075064:Myeloid Ecotropic Viral Integration Site 1 Protein; D009363:Neoplasm Proteins; C090465:PBX2 protein, human; C106682:PKNOX1 protein, human; D011500:Protein Synthesis Inhibitors; D011518:Proto-Oncogene Proteins; D012333:RNA, Messenger; D014212:Tretinoin; D004247:DNA; D003513:Cycloheximide; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde