Molecular cloning and characterization of the promoter of mouse heme-regulated eIF2alpha kinase.
The heme-regulated eIF2alpha kinase (HRI) phosphorylates the alpha subunit of the translation initiation factor 2, which plays an important role in translational regulation during heme deficiency. HRI is expressed mainly in erythroid cells. To elucidate the mechanisms that regulate the tissue-specific expression of the mouse HRI gene, we have cloned and characterized its 5' flanking region. Primer extension and 5' RACE analyses identified a major transcription initiation site 84 nucleotides upstream from the ATG start codon. Sequence analysis of the 5' flanking region revealed that HRI promoter lacked a TATA element, but contained an initiator core element and three SP1 consensus binding motifs. Transient transfection analysis using luciferase reporter gene constructs containing a series of deletions and mutations in the 5' region identified a 159 bp sequence which is necessary and sufficient for basal HRI promoter activities in NIH 3T3 and mouse erythroleukemic cells. Both the initiator and one of the SP1 binding sites in the 159 bp are essential for HRI promoter activity.
Lu
Linrong
L
Harvard-MIT Division of Health Science and Technology, Massachusetts Institute of Technology, E25-545, 77 Massachusetts Avenue, Cambridge, MA 02139, USA. linrong_lu@dfci.harvard.edu
Chen
Jane Jane
JJ
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D018983:DNA Footprinting; D018076:DNA, Complementary / Q000096:biosynthesis / Q000737:chemistry; D024202:Electrophoretic Mobility Shift Assay; D051379:Mice; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D015183:Restriction Mapping; D016415:Sequence Alignment; D014407:Tumor Cells, Cultured; D019892:eIF-2 Kinase / Q000737:chemistry / Q000235:genetics*
D018076:DNA, Complementary; D019892:eIF-2 Kinase