The influence of sex steroid hormones on ferrochelatase gene expression in Harderian gland of hamster (Mesocricetus auratus).
Ferrochelatase (protohaem ferrolyase, EC 4.99.1.1), the terminal enzyme of the haem biosynthetic pathway, catalyses the insertion of ferrous iron into protoporphyrin IX to form protohaem. The Syrian hamster Harderian gland (HG) is known for its ability to produce and accumulate large amounts of protoporphyrins. In this species, the female gland contains up to 120 times more porphyrin than the male gland. Data from biochemical studies suggest that this gland possesses the enzymatic complex for haem biosynthesis but lacks ferrochelatase activity. The abundance of intraglandular haem proteins does not support this idea. To gain more insight into this process, we isolated cDNA for ferrochelatase from hamster liver, using the 5'- and 3'- rapid amplification of complementary DNA ends (RACE), and investigated its expression in HG from males and females. The full-length cDNA comprises an open reading frame of 1269 bp encoding a polypeptide of 422 amino-acid residues. Hamster DNA sequence exhibits 92% identity to mouse and 87% identity to human sequences. The predicted hamster enzyme was shown to have structural features of mammalian ferrochelatase, including a putative NH2- terminal presequence, a central core of about 330 amino-acid residues and an extra 30-50-amino-acid stretch at the carboxyl-terminus. RNA blotting experiments indicated that this cDNA hybridized to a liver mRNA of about 2.1 kb, while a weak hybridization signal was observed with mRNA from HG preparations. RT-PCR assays confirmed the expression of specific transcripts in both tissues. Male glands contained approximately twofold more enzyme mRNA than female glands. Likewise, the intraglandular content of mRNA varied during the oestrous cycle, with the highest levels found in the oestrous phase. These cyclic variations were less evident in liver. Ovariectomy plus treatment with progesterone or 17beta-oestradiol plus progesterone increased ferrochelatase mRNA of the gland. In HG of short- or long-term castrated males, the administration of testosterone did not affect the ferrochelatase mRNA concentration. Based on mRNA expression levels, we conclude that Harderian ferrochelatase may play an active role in maintaining the physiological pool of haem required for processing cytochromes and other glandular haem proteins. Likewise, the sex-steroid hormones appear to have only a modest influence upon Harderian ferrochelatase.
Vilchis
F
F
Department of Reproductive Biology, Instituto Nacional de Ciencias Médicas y Nutrición Salvador Zubirán, Vasco de Quiroga no. 15, Del. Tlalpan C.P. 14000, México DF, Mexico.
Ramos
L
L
Timossi
C
C
Chávez
B
B
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern / Q000379:methods; D002369:Castration; D003001:Cloning, Molecular / Q000379:methods; D006224:Cricetinae; D004270:DNA, Circular / Q000235:genetics; D004958:Estradiol / Q000008:administration & dosage / Q000502:physiology; D004971:Estrus / Q000502:physiology; D005260:Female; D005294:Ferrochelatase / Q000235:genetics*; D015870:Gene Expression / Q000235:genetics; D012739:Gonadal Steroid Hormones / Q000502:physiology*; D006243:Harderian Gland / Q000201:enzymology*; D008099:Liver / Q000201:enzymology; D008297:Male; D008647:Mesocricetus; D011374:Progesterone / Q000008:administration & dosage / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013739:Testosterone / Q000008:administration & dosage / Q000502:physiology
D004270:DNA, Circular; D012739:Gonadal Steroid Hormones; D012333:RNA, Messenger; D013739:Testosterone; D011374:Progesterone; D004958:Estradiol; D005294:Ferrochelatase