Transcription of the catalytic 180-kDa subunit gene of mouse DNA polymerase alpha is controlled by E2F, an Ets-related transcription factor, and Sp1.
We have isolated a genomic DNA fragment spanning the 5'-end of the gene encoding the catalytic subunit of mouse DNA polymerase alpha. The nucleotide sequence of the upstream region was G/C-rich and lacked a TATA box. Transient expression assays in cycling NIH 3T3 cells demonstrated that the GC box of 20 bp (at nucleotides -112/-93 with respect to the transcription initiation site) and the palindromic sequence of 14 bp (at nucleotides -71/-58) were essential for basal promoter activity. Electrophoretic mobility shift assays showed that Sp1 binds to the GC box. We also purified a protein capable of binding to the palindrome and identified it as GA-binding protein (GABP), an Ets- and Notch-related transcription factor. Transient expression assays in synchronized NIH 3T3 cells revealed that three variant E2F sites near the transcription initiation site (at nucleotides -23/-16, -1/+7 and +17/+29) had no basal promoter activity by themselves, but were essential for growth-dependent stimulation of the gene expression. These data indicate that E2F, GABP and Sp1 regulate the gene expression of this principal replication enzyme.
Izumi
M
M
Division of Radioisotope Technology, RIKEN (The Institute of Physical and CHemical Research), Saitama, Japan. mizumi@postman.riken.go.jp
Yokoi
M
M
Nishikawa
N S
NS
Miyazawa
H
H
Sugino
A
A
Yamagishi
M
M
Yamaguchi
M
M
Matsukage
A
A
Yatagai
F
F
Hanaoka
F
F
D016475:3T3 Cells; D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins; D002384:Catalysis; D002453:Cell Cycle / Q000235:genetics; D018797:Cell Cycle Proteins; D004256:DNA Polymerase I / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; D015971:Gene Expression Regulation, Enzymologic; D005809:Genes, Regulator; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D015513:Oncogene Proteins; D011401:Promoter Regions, Genetic / Q000235:genetics; D050783:Proto-Oncogene Proteins c-ets; D056465:Retinoblastoma-Binding Protein 1; D012689:Sequence Homology, Nucleic Acid; D016329:Sp1 Transcription Factor / Q000502:physiology*; D050680:Transcription Factor DP1; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology*; D014158:Transcription, Genetic
C533431:Arid4a protein, mouse; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D050684:E2F Transcription Factors; C086924:Elk3 protein, mouse; D015513:Oncogene Proteins; D050783:Proto-Oncogene Proteins c-ets; D056465:Retinoblastoma-Binding Protein 1; D016329:Sp1 Transcription Factor; D050680:Transcription Factor DP1; D014157:Transcription Factors; D004256:DNA Polymerase I