Sexual dimorphism of testosterone 15 alpha-hydroxylase mRNA levels in mouse liver. cDNA cloning and regulation.
A cDNA library was constructed from 129/J female mouse liver poly(A)+ RNA immunoenriched for testosterone 15 alpha-hydroxylase (P-450(15)alpha) in the expression vector pUC9. Fifteen clones coding for P-450(15)alpha were identified by duplicate colony hybridization to nick-translated cDNAs synthesized from immunoenriched or depleted poly(A)+ RNA. Recombinant plasmid p 15 alpha-29 contained the largest cDNA (1.6 kilobases) and cross-hybridized strongly with all other clones. cDNAs representing two different transcripts were identified based on restriction map analysis. Clones p 15 alpha-29 and p 15 alpha-15 have identical internal restriction fragments generated by ClaI, BamHI, and StuI digests, but p15 alpha-15 has unique PstI and HindIII sites located within 150 base pairs of each other. This suggests P-450(15)alpha is transcribed from two genes. Northern hybridization of poly(A)+ RNA with nick-translated p15 alpha-29 or p15 alpha-15 showed a single size mRNA of 2.1 kilobases. The level of P-450(15)alpha mRNA was 6.6 times higher in 129/J female than in male mice. The difference between P-450(15)alpha mRNA levels in females and males was positively correlated with P-450(15)alpha protein detected by anti-P-450(15)alpha and testosterone 15 alpha-hydroxylase activity in microsomes. Sexual dimorphism in hepatic testosterone 15 alpha-hydroxylase activity is shown to result from differential mRNA regulation in females and males.
B A
BA
Burkhart
N
N
Harada
M
M
Negishi
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D003001:Cloning, Molecular; D000072467:Cytochrome P450 Family 2; D004247:DNA / Q000032:analysis*; D004262:DNA Restriction Enzymes / Q000378:metabolism; D005260:Female; D008099:Liver / Q000201:enzymology*; D008297:Male; D051379:Mice; D009693:Nucleic Acid Hybridization; D010957:Plasmids; D011061:Poly A / Q000032:analysis; D012313:RNA / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis*; D011817:Rabbits; D012727:Sex Characteristics; D013250:Steroid Hydroxylases / Q000235:genetics*
D012333:RNA, Messenger; D011061:Poly A; D012313:RNA; D004247:DNA; D013250:Steroid Hydroxylases; D001189:Aryl Hydrocarbon Hydroxylases; C460223:Cyp2a4 protein, mouse; D000072467:Cytochrome P450 Family 2; C022724:testosterone 7-alpha-hydroxylase, hamster; D004262:DNA Restriction Enzymes