Characterization and functional analysis of the porcine lactoferrin gene promoter.
Lactoferrin, a ferric binding glycoprotein found in milk, can possibly prevent microbial infection of the mammary gland and gastrointestinal tract. To define the regulation of the porcine lactoferrin gene (pLTF), we cloned its 5'-flanking region from a porcine liver genomic library and analyzed the 5' upstream region of approx. 4kb, two exons, and an intron. The transcription start site was localized by primer extension to residue G, which is 41 nucleotides upstream from the ATG start codon. The pLTF 5'-flanking region possesses several putative cis-acting regulatory elements found in both housekeeping and inducible genes; to define their function, they were inserted into a chloramphenicol acetyltransferase reporter construct. The region up to -156 sufficed for basic promoter activity, whereas the region up to -780 was required for maximal promoter activity in porcine testis cells (STcells), kidney cells (PK15 cells) and human mammary epithelial cells (HBL-100 cells). Detailed analysis of this proximal region by DNase I footprinting and electrophoretic mobility shift assays reveals that the ubiquitous factors SP1, AP2 and the mammary gland-specific factor (MGF) might play significant roles in regulating the transcription of the pLTF gene.
Department of Life Science, National Tsing-Hua University, Hsinchu, Taiwan, ROC.
S R
SR
Wang
J
J
Lin
I C
IC
Cheng
T Y
TY
Lin
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001940:Breast / Q000737:chemistry / Q000378:metabolism; D002457:Cell Extracts / Q000737:chemistry; D016384:Consensus Sequence / Q000235:genetics; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005260:Female; D005796:Genes / Q000235:genetics*; D007668:Kidney / Q000737:chemistry / Q000166:cytology / Q000378:metabolism; D007781:Lactoferrin / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D008297:Male; D008894:Milk Proteins; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding; D050799:STAT5 Transcription Factor; D017422:Sequence Analysis, DNA; D017384:Sequence Deletion / Q000235:genetics; D016329:Sp1 Transcription Factor / Q000737:chemistry / Q000378:metabolism; D013552:Swine / Q000235:genetics*; D013737:Testis / Q000737:chemistry / Q000166:cytology / Q000378:metabolism; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism; D050656:Transcription Factor AP-2; D014157:Transcription Factors / Q000737:chemistry / Q000378:metabolism; D014158:Transcription, Genetic / Q000235:genetics
D002457:Cell Extracts; D004268:DNA-Binding Proteins; D008894:Milk Proteins; D009687:Nuclear Proteins; D050799:STAT5 Transcription Factor; D016329:Sp1 Transcription Factor; D015534:Trans-Activators; D050656:Transcription Factor AP-2; D014157:Transcription Factors; D004247:DNA; D007781:Lactoferrin