Multiple initiators and C/EBP binding sites are involved in transcription from the TATA-less rat XDH/XO basal promoter.
In the present study, we have explored further the organization of the TATA-less rat xanthine dehydrogenase/oxidase gene (XDH/XO). A DNase I hypersensitive site has been identified which it colocalizes with the basal promoter reported previously [Chow et al. (1994) Nucleic Acids Res., 22, 1846-1854]. Gel mobility shift assays indicate the presence of multiple binding factors located in the promoter. At least six footprints were detected of which two have been shown to be C/EBP binding sites. Members of the C/EBP-alpha and C/EBP-beta, but not C/EBP-delta, family are able to bind to these two sites. Deletional and mutational studies revealed that C/EBP binding is not essential for the basal level of transcription initiation of this promoter. Much of the transcriptional activity resides in the -102 to -7 DNA fragment, which contains all initiator activity which acts unidirectionally. Within this fragment, four putative initiator elements could be identified; interestingly, the linear integrity of these initiators is important for efficient transcription of the XDH/XO gene. Separation of the initiators leads to a complete loss of transcription activity; however, this loss could be partially restored by the introduction of an Sp1 binding site upstream of the separated initiators. Despite a difference in usage/frequency of initiation at the various initiators, primer extension analyses reveal similar positions for transcription initiations in both XDH/XO reporter constructs and in the endogenous XDH/XO gene. The differential usage of initiators may imply a possible post-transcriptional regulation for the XDH/XO gene.
Chow
C W
CW
Department of Molecular Physiology and Biophysics, School of Medicine, Vanderbilt University, Nashville, TN 37232, USA.
Clark
M P
MP
Rinaldo
J E
JE
Chalkley
R
R
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D022762:CCAAT-Enhancer-Binding Proteins; D004247:DNA / Q000235:genetics; D015342:DNA Probes / Q000235:genetics; D004268:DNA-Binding Proteins / Q000378:metabolism*; D066298:In Vitro Techniques; D008099:Liver / Q000378:metabolism; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics; D051381:Rats; D016385:TATA Box / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D014968:Xanthine Dehydrogenase / Q000235:genetics*; D014969:Xanthine Oxidase / Q000235:genetics*
D022762:CCAAT-Enhancer-Binding Proteins; D015342:DNA Probes; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D012333:RNA, Messenger; D014157:Transcription Factors; D004247:DNA; D014968:Xanthine Dehydrogenase; D014969:Xanthine Oxidase