Weakening of the repressive YY-1 site on the thrombospondin-1 promoter via c-Jun/YY-1 interaction.
Thrombospondin-1 (TSP-1) level is tightly regulated at the transcriptional level. To determine the detailed molecular mechanisms of TSP-1 expression, nine serial 5'-deletion constructs of the human genomic tsp-1 promoter (nucleotides -2,220 to +756) were prepared, inserted into luciferase reporter plasmids, and transiently transfected into the Hep3B human hepatocarcinoma cell. Among the nine 5'-deletion constructs, pTSP-Luc-4 (-767 approximately +756) had consistently decreased luciferase activity with or without PMA stimulation, whereas a further truncated construct [pTSP-Luc-4' (-407 approximately +756)] had increased levels of expression. By searching the nucleotides from -767 to -407, a consensus binding sequence (5'-CCATTTT-3') for the repressor Yin Yang-1 (YY-1) at nucleotide -440 was identified. The suppression induced by this site was weakened in the presence of the region upstream of nucleotide -767 (pTSP-Luc-1 and -2). Nuclear protein directly bound to an oligonucleotide containing the repressive YY-1 sequence but the binding capacity of the sequence was decreased by the increased c-Jun levels. Moreover, proteins immunoprecipitated with anti-YY-1 revealed an interaction between c-Jun and YY-1 factor. These data suggest that the repressive YY-1 site of the tsp-1 promoter could not be functional via activating positive cis-elements on the upstream from this site and weakened via c-Jun/YY-1 interactions.
Kang
Jung-Hoon
JH
Department of Biochemistry, College of Medicine, The Catholic University of Korea, 505 Banpo-dong, Socho-gu, Seoul, 137-701, Korea.
Chang
Seo-Yoon
SY
Yeom
Dong-Hoon
DH
Kim
Soo-A
SA
Um
Soo-Jong
SJ
Hong
Kyong-Ja
KJ
D001665:Binding Sites / Q000235:genetics; D045744:Cell Line, Tumor; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015536:Down-Regulation / Q000235:genetics; D024202:Electrophoretic Mobility Shift Assay; D050981:Erythroid-Specific DNA-Binding Factors; D017930:Genes, Reporter / Q000235:genetics; D006801:Humans; D008156:Luciferases / Q000032:analysis / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics*; D016755:Proto-Oncogene Proteins c-jun / Q000235:genetics / Q000378:metabolism*; D012097:Repressor Proteins / Q000378:metabolism*; D017384:Sequence Deletion / Q000235:genetics; D019700:Thrombospondin 1 / Q000235:genetics* / Q000378:metabolism; D018808:Transcription Factor AP-1 / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D050986:YY1 Transcription Factor
D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; D016755:Proto-Oncogene Proteins c-jun; D012097:Repressor Proteins; D019700:Thrombospondin 1; D018808:Transcription Factor AP-1; D014157:Transcription Factors; D050986:YY1 Transcription Factor; D008156:Luciferases