Siah-1b is a direct transcriptional target of p53: identification of the functional p53 responsive element in the siah-1b promoter.
Siah proteins are E3 ubiquitin ligases. They are homologues of the Drosophila seven in absentia (Sina), a protein required for the R7 photoreceptor development. We have previously found that the expression of human siah-1 and its mouse homologue siah-1b are induced by p53 during apoptosis and tumor reversion. So far, no evidence that the siah-1b gene is a direct transcriptional target of p53 has been provided. In the present study we investigate this issue. Northern blot analysis with a specific probe demonstrates an increase in siah-1b transcription on activation of endogenous and inducible exogenous p53. To explore whether this effect is directly mediated by p53 we analyzed 20 kb of chromosome X DNA, containing the siah-1b locus. A p53-binding site was identified in the siah-1b promoter, located at nucleotides -2155/-2103 relative to the translational start site. This site is composed of two half-sites, conforming to the p53-binding consensus sequence but separated by a nonclassical 33-bp spacer. In luciferase assays, p53 induces a substantial increase in siah-1b promoter activity. Gel shift and DNase-I-footprinting studies, combined with mutational analysis and chromatin immunoprecipitation, indicate that p53 effectively binds the siah-1b promoter in vitro and in vivo. Thus, the siah-1b gene is a direct transcriptional target of p53.
Molecular Engines Laboratories, 20 Rue Bouvier, 75011 Paris, France.
Giusy
G
Fiucci
Séverine
S
Beaucourt
Dominique
D
Duflaut
Alexandra
A
Lespagnol
Pamela
P
Stumptner-Cuvelette
Anne
A
Géant
Gilles
G
Buchwalter
Marcel
M
Tuynder
Laurent
L
Susini
Jean-Michel
JM
Lassalle
Christine
C
Wasylyk
Bohdan
B
Wasylyk
Moshe
M
Oren
Robert
R
Amson
Adam
A
Telerman
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D045744:Cell Line, Tumor; D002843:Chromatin / Q000235:genetics / Q000378:metabolism; D016384:Consensus Sequence; D004273:DNA, Neoplasm / Q000235:genetics / Q000378:metabolism; D066298:In Vitro Techniques; D007438:Introns; D051379:Mice; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D015533:Transcriptional Activation; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*; D044767:Ubiquitin-Protein Ligases; D000098563:Seven in Absentia Proteins
D002843:Chromatin; D004273:DNA, Neoplasm; D009687:Nuclear Proteins; D012333:RNA, Messenger; D016159:Tumor Suppressor Protein p53; D044767:Ubiquitin-Protein Ligases; D000098563:Seven in Absentia Proteins