Megakaryocyte-specific positive regulatory sequence 5' to the human PF4 gene.
Platelet factor 4 (PF4) is only expressed in platelets and is an appropriate marker for studying megakaryocytic differentiation. We previously characterized cDNA and genomic clones for human PF4 (hPF4) and now present transient expression studies defining the promoter of the gene. 12-O-tetradecanoyl-phorbol-13- acetate (TPA) induces megakaryocytic differentiation of human erythroleukemia (HEL) cells, providing an excellent model system for the study of megakaryocyte-specific promoter activity. Luciferase reporter-gene constructs containing sequences from -2074 to +49 were used to map regions that may regulate PF4 gene expression. The sequence in the region -239 to -107 increased basal promoter activity by four- to five-fold in TPA-induced HEL cells. The sequence between -239 and -107 contains 53 consecutive thymidine residues. Functional studies using constructs in this region show that poly(T) and the region -187 to -107 are necessary for the total increase in activity in TPA-induced HEL cells. Mobility-shift assays show that the poly(T) tract binds TPA-inducible proteins. The results suggest a complex promoter for the PF4 gene involving a basal nonspecific promoter element between -107 and +49, a positive promoter element between -239 and -107 binding specific nuclear proteins from megakaryocyte-lineage cells, and a silencer-like region between -2074 and -1653.
Ramachandran
B
B
Division of Hematology, Children's Hospital of Philadelphia, PA.
Surrey
S
S
Schwartz
E
E
D001483:Base Sequence; D002874:Chromosome Mapping; D004247:DNA / Q000737:chemistry; D004742:Enhancer Elements, Genetic; D015870:Gene Expression; D017930:Genes, Reporter; D006801:Humans; D004915:Leukemia, Erythroblastic, Acute / Q000378:metabolism; D008156:Luciferases / Q000235:genetics; D008533:Megakaryocytes / Q000378:metabolism*; D008969:Molecular Sequence Data; D010978:Platelet Factor 4 / Q000235:genetics*; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins; D012045:Regulatory Sequences, Nucleic Acid; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection; D014407:Tumor Cells, Cultured
D011993:Recombinant Fusion Proteins; D010978:Platelet Factor 4; D004247:DNA; D008156:Luciferases; D013755:Tetradecanoylphorbol Acetate