Regulation of murine lactate dehydrogenase C (Ldhc) gene expression.
Expression of Ldhc begins with the onset of meiosis in male germ cells and continues throughout spermatogenesis. Transcriptional regulatory mechanisms, especially in primary spermatocytes, are poorly described because of the lack of a reliable cell culture system. We constructed mouse transgenics and transfected germ cells in situ to study expression of the testis-specific isozyme of lactate dehydrogenase (LDH). From previous work, we determined that a 100-bp Ldhc core promoter contained potential cis regulatory elements, including a palindrome (-21 to +10), GC box (-70 to -65), and cAMP-responsive element (CRE) sites (-53 to -49, -39 to -35). We provide here the demonstration of a functional role for these sequences by expression of mutated transgenes in vivo. Our results reveal for the first time that mutation of the GC box does not abolish promoter activity, which remains testis-specific. Mutation of GC box or CRE sites resulted in a 73% and 74% reduction in promoter activity, respectively, in a transient transfection of germ cells in vivo by electroporation; the combination of GC box and CRE site mutations eliminates promoter activity. Therefore, we conclude that simultaneous occupancy of the GC box and CRE sites in the core promoter is necessary for full expression of Ldhc in the testis.
Tang
HuangHui
H
Department of Biochemistry, Molecular Biology and Cell Biology, Northwestern University, 2205 Tech Drive, Evanston, IL 60208, USA.
Kung
Aisha
A
Goldberg
Erwin
E
D000818:Animals; D001482:Base Composition; D001483:Base Sequence; D001665:Binding Sites; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D015971:Gene Expression Regulation, Enzymologic; D007527:Isoenzymes / Q000235:genetics / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000235:genetics* / Q000378:metabolism; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D009928:Organ Specificity / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D020218:Response Elements; D012689:Sequence Homology, Nucleic Acid; D013737:Testis / Q000378:metabolism; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D017362:Cyclic AMP Response Element-Binding Protein; D007527:Isoenzymes; D011993:Recombinant Fusion Proteins; D007770:L-Lactate Dehydrogenase; C409673:lactate dehydrogenase C4; D001616:beta-Galactosidase