Characterization of estrogen-responsive mouse lactoferrin promoter.
Mouse lactoferrin is expressed in a variety of tissues under different types of control. To understand how molecular mechanisms govern the mode of lactoferrin expression, we isolated and characterized the 5'-flanking region of the lactoferrin gene. Several clones containing lactoferrin gene fragments were isolated from a mouse (129/J) genomic library including clone lambda J14, which contains a 7.5-kilobase pair 5'-flanking sequence. Sequence analysis of the region flanking the transcription initiation site revealed the following: a TATA-like sequence, two CAAT boxes, three GC boxes including one within the first intron, an AP2 site, seven PU boxes, an AC-rich region, a B1 sequence, and an estrogen-responsive element consensus sequence over-lapping with a chicken ovalbumin upstream promoter-binding element. Footprinting analysis demonstrated that several regions, including the putative estrogen-responsive element region, in the 5'-flanking sequence were protected from DNase I digestion. Promoter fragments were cloned into a chloramphenicol acetyltransferase receptor plasmid to study functional activity. The mouse lactoferrin gene promoter was active in human endometrium carcinoma RL 95-2 cells and in rat glioma C6 cells. Multiple upstream elements modulated the basal transcriptional promoter activity. The transcription level directed by this minimal promoter was controlled by both positive (between -1739 and -922) and negative (between -2644 and -1739, and between -589 and -291) regulatory sequences. A tissue-specific regulatory sequence was critical for the establishment of lactoferrin expression in human endometrium carcinoma cells, but not in rat glioma cells located between -1739 and -922. Reporter plasmid 0.6 mL14-CAT, containing the estrogen-responsive element sequence, was estrogen-responsive in the presence of estrogen receptor in human endometrium carcinoma RL 95-2 cells.
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Developmental Endocrinology and Pharmacology Section, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709.
Teng
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D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D016889:Endometrial Neoplasms; D004967:Estrogens / Q000502:physiology*; D005260:Female; D005809:Genes, Regulator; D006801:Humans; D007438:Introns; D007781:Lactoferrin / Q000235:genetics*; D008099:Liver / Q000187:drug effects / Q000502:physiology*; D051379:Mice; D008815:Mice, Inbred Strains; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000187:drug effects; D051381:Rats; D011960:Receptors, Estrogen / Q000032:analysis; D012045:Regulatory Sequences, Nucleic Acid; D015183:Restriction Mapping; D016385:TATA Box; D014162:Transfection
D004967:Estrogens; D011960:Receptors, Estrogen; D007781:Lactoferrin