The ePHD protein SPBP interacts with TopBP1 and together they co-operate to stimulate Ets1-mediated transcription.
SPBP (Stromelysin-1 PDGF responsive element binding protein) is a ubiquitously expressed 220 kDa nuclear protein shown to enhance or repress the transcriptional activity of various transcription factors. A yeast two-hybrid screen, with the extended plant homeodomain (ePHD) of SPBP as bait, identified TopBP1 (topoisomerase II beta-binding protein 1) as a candidate interaction partner of SPBP. TopBP1 has eight BRCA1 carboxy-terminal (BRCT) domains and is involved in DNA replication, DNA damage responses and in the regulation of gene expression. The interaction between SPBP and TopBP1 was confirmed in vitro and in vivo, and was found to be mediated by the ePHD domain of SPBP and the BRCT6 domain of TopBP1. Both SPBP and TopBP1 enhanced the transcriptional activity of Ets1 on the c-myc P1P2- and matrix metalloproteinase-3 (MMP3) promoters. Together they displayed a more than additive effect. Both proteins were associated with these promoters. The involvement of TopBP1 was dependent on the serine 1159 phosphorylation site, known to be important for transcriptional activation. Depletion of endogenous SPBP by siRNA treatment reduced MMP3 secretion by 50% in phorbol ester-stimulated human fibroblasts. Taken together, our results show that TopBP1 and SPBP interact physically and functionally to co-operate as co-activators of Ets1.
Sjøttem
Eva
E
Biochemistry Department and Department of Molecular Biotechnology, Institute of Medical Biology, University of Tromsø, 9037 Tromsø, Norway.
Rekdal
Cecilie
C
Svineng
Gunbjørg
G
Johnsen
Sylvia Sagen
SS
Klenow
Helle
H
Uglehus
Rebecca Dale
RD
Johansen
Terje
T
D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D005347:Fibroblasts / Q000201:enzymology; D006367:HeLa Cells; D006801:Humans; D019278:Matrix Metalloproteinase 3 / Q000235:genetics / Q000378:metabolism; D009687:Nuclear Proteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D017434:Protein Structure, Tertiary; D050784:Proto-Oncogene Protein c-ets-1 / Q000378:metabolism*; D015534:Trans-Activators; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D015533:Transcriptional Activation; D020798:Two-Hybrid System Techniques
D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D050784:Proto-Oncogene Protein c-ets-1; C431552:RAI1 protein, human; C114750:TCF20 protein, human; C110513:TOPBP1 protein, human; D015534:Trans-Activators; D014157:Transcription Factors; D019278:Matrix Metalloproteinase 3