Abstract
The p53 tumor suppressor protein plays a crucial role in tumorigenesis by controlling cell-cycle progression and apoptosis. We have previously described a transcript designated tumor suppressor activated pathway-6 (TSAP6) that is up-regulated in the p53-inducible cell line, LTR6. Cloning of the murine and human full-length TSAP6 cDNA revealed that it encodes a 488-aa protein with five to six transmembrane domains. This gene is the murine and human homologue of the recently published rat pHyde. Antibodies raised against murine and human TSAP6 recognize a 50- to 55-kDa band induced by p53. Analysis of the TSAP6 promoter identified a functional p53-responsive element. Functional studies demonstrated that TSAP6 antisense cDNA diminished levels of the 50- to 55-kDa protein and decreased significantly the levels of p53-induced apoptosis. Furthermore, TSAP6 small interfering RNA inhibited apoptosis in TSAP6-overexpressing cells. Yeast two-hybrid analysis followed by GST/in vitro-transcribed/translated pull-down assays and in vivo coimmunoprecipitations revealed that TSAP6 associated with Nix, a proapoptotic Bcl-2-related protein and the Myt1 kinase, a negative regulator of the G(2)/M transition. Moreover, TSAP6 enhanced the susceptibility of cells to apoptosis and cooperated with Nix to exacerbate this effect. Cell-cycle studies indicated that TSAP6 could augment Myt1 activity. Overall, these data suggest that TSAP6 may act downstream to p53 to interface apoptosis and cell-cycle progression.
MeSH terms
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Amino Acid Sequence
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Animals
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Apoptosis*
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Blotting, Northern
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Blotting, Western
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Cell Cycle
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Cell Cycle Proteins
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Cloning, Molecular
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DNA, Complementary / metabolism
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Flow Cytometry
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G1 Phase
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G2 Phase
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Glutathione Transferase / metabolism
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HeLa Cells
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Humans
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Luciferases / metabolism
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Membrane Proteins / genetics*
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Membrane Proteins / metabolism*
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Membrane Proteins / physiology*
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Mice
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Mitosis
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Molecular Sequence Data
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Oligonucleotides, Antisense / pharmacology
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Oncogene Proteins*
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Oxidoreductases
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Phosphorylation
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Precipitin Tests
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Promoter Regions, Genetic
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Protein Binding
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Protein Biosynthesis
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Protein Serine-Threonine Kinases / metabolism*
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Protein Structure, Tertiary
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Protein-Tyrosine Kinases / metabolism*
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Proto-Oncogene Proteins c-bcl-2 / metabolism
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Proto-Oncogene Proteins*
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RNA, Messenger / metabolism
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RNA, Small Interfering / metabolism
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Recombinant Fusion Proteins / metabolism
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S Phase
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Sequence Homology, Amino Acid
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Time Factors
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Transcription, Genetic
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Tumor Cells, Cultured
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Tumor Suppressor Protein p53 / metabolism*
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Tumor Suppressor Proteins*
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Two-Hybrid System Techniques
Substances
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BNIP3L protein, human
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Cell Cycle Proteins
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DNA, Complementary
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Membrane Proteins
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Oligonucleotides, Antisense
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Oncogene Proteins
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Proto-Oncogene Proteins
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Proto-Oncogene Proteins c-bcl-2
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RNA, Messenger
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RNA, Small Interfering
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Recombinant Fusion Proteins
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Tumor Suppressor Protein p53
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Tumor Suppressor Proteins
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Oxidoreductases
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STEAP3 protein, human
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TSAP6 protein, mouse
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Luciferases
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Glutathione Transferase
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Myt1 kinase, rat
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Protein-Tyrosine Kinases
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PKMYT1 protein, human
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Protein Serine-Threonine Kinases
Associated data
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GENBANK/AY214461
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GENBANK/AY214462