Identification and characterization of promoter and regulatory regions for mouse Adam2 gene expression.
ADAM2, a member of the 'a disintegrin and metalloprotease' (ADAM) family, is a key protein in mammalian fertilization that is specifically expressed in testicular germ cells. Here, we investigated the transcriptional regulation of the mouse Adam2 gene. An in silico analysis identified two conserved non-coding sequences located upstream of the mouse and human ADAM2 genes. The upstream region of the mouse Adam2 gene was found to lack typical TATA and CAAT boxes, and to have a high GC content. Our in vitro transient transfection-reporter analysis identified a promoter in this region of the mouse Adam2 gene, along with regulatory regions that inhibit the activity of this promoter in somatic cells. Site-directed mutagenesis revealed that the caudal-type homeobox 1 and CCTC-binding factor motifs are responsible for the inhibitory activities of the repressor regions. Finally, electrophoretic mobility shift assays showed putative transcription factor-promoter DNA complexes, and DNA-affinity chromatography and proteomic analyses identified myelin gene regulatory factor as a binding partner of the Adam2 promoter. This provides the first identification and characterization of promoter and repressor regions that regulate the transcription of the mouse Adam2 gene, and offers insights into the regulation of this germ-cell-specific gene.
School of Life Sciences, Gwangju Institute of Science and Technology, Gwangju, 500-712, Korea.
Heejin
H
Choi
Boyeon
B
Lee
Sora
S
Jin
Jun Tae
JT
Kwon
Jihye
J
Kim
Juri
J
Jeong
Seungmin
S
Oh
Byung-Nam
BN
Cho
Zee Yong
ZY
Park
Chunghee
C
Cho
D051722:ADAM Proteins / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D024202:Electrophoretic Mobility Shift Assay; D000072196:Fertilins; D015971:Gene Expression Regulation, Enzymologic; D017930:Genes, Reporter; D057809:HEK293 Cells; D006801:Humans; D049410:Luciferases, Renilla / Q000096:biosynthesis / Q000235:genetics; D008297:Male; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D058977:Molecular Sequence Annotation; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D041681:NIH 3T3 Cells; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017422:Sequence Analysis, DNA; D013737:Testis / Q000166:cytology; D014157:Transcription Factors / Q000378:metabolism
D008562:Membrane Glycoproteins; D014157:Transcription Factors; C571125:myelin gene regulatory factor, mouse; D049410:Luciferases, Renilla; D051722:ADAM Proteins; C000606454:ADAM2 protein, human; C000606458:Adam2 protein, mouse; D000072196:Fertilins