Iron has a variety of functions in cellular organisms ranging from electron transport and DNA synthesis to adenosine triphosphate (ATP) and neurotransmitter synthesis. Failure to regulate the homeostasis of iron can lead to cognition and demyelination disorders when iron levels are deficient, and to neurodegenerative disorders when iron is in excess. In this study we show that three members of the b561 family of predicted ferric reductases, namely mouse cytochrome b561 and mouse and fly stromal cell-derived receptor 2 (SDR2), have ferric reductase activity. Given that a fourth member, duodenal cytochrome b (Dcytb), has previously been shown to be a ferric reductase, it is likely that all remaining members of this family also exhibit this activity. Furthermore, we show that the rat sdr2 message is predominantly expressed in the liver and kidney, with low expression in the duodenum. In hypotransferrinaemic (hpx) mice, sdr2 expression in the liver and kidney is reduced, suggesting that it may be regulated by iron. Moreover, we demonstrate the presence of mouse sdr2 in the choroid plexus and in the ependymal cells lining the four ventricles, through in situ hybridization analysis.

Cytochrome b561 is a major transmembrane protein of catecholamine and neuropeptide secretory vesicles. In this report, we describe the cloning and properties of a full-length cDNA encoding human neuroendocrine cytochrome b561 from a human caudate cDNA library and a human peripheral blood genomic library. The human cDNA contains two major transcription start sites and only one translation start site that codes for an apocytochrome b561, which is 22 amino acid residues smaller than the previously deduced amino acid sequence from bovine cDNA. This smaller version of cytochrome b561 may contain only five transmembrane segments rather than the previously proposed six segments. The new model is in agreement with our previous results on transmembrane topology of the gene product. Northern-blot analysis shows an expanded tissue distribution of cytochrome mRNA expression where previous immunological assays were negative. These results support the hypothesis that cytochrome b561 is a marker for peptidergic and adrenergic tissues.

Cytochrome b561 is an electron transfer protein unique to neuroendocrine secretory vesicles. The Southern blot hybridization shows that it is a single copy gene highly conserved throughout phylogeny. The transcription unit spans approximately 11 kilobases, and heterologous transcription sites are located 404 bases 5' to the translation initiation codon. The sequence of the 5'-flanking region is GC-rich and lacks a typical TATA box at the usual position. However, it has a CAAT sequence at -132 and potential recognition sequences for several transcription factors including SP1, GR-PR-MMTV, AP4, gERE, JCV repeat, AP2, and NF-kappa B. Each of the five transmembrane segments are encoded by five consecutive exons. This corroborates the five-transmembrane model proposed for human, mouse, and Xenopus rather than six proposed for bovine. The cytochrome was found to be highly expressed in colon cancer cell lines, T cell lymphomas, and K-562 cell lines. However, in B-cell lymphomas such as Burkitt's and Daudi, the cytochrome b561 expression was completely shut down. The results in this report are the first to demonstrate the structural organization and regulatory sequences of the cytochrome b561 gene encoding an integral membrane protein of neuroendocrine storage vesicles of neurotransmitters and peptide hormones. Unexpected results on cytochrome b561 expression in cells of lymphocytic origin and its complex regulation in tumor cells provide new insights into cytochrome b561 gene regulation.

Cytochrome b561 is an electron transfer protein unique to neuroendocrine secretory vesicles. The Southern blot hybridization shows that it is a single copy gene highly conserved throughout phylogeny. The transcription unit spans approximately 11 kilobases, and heterologous transcription sites are located 404 bases 5' to the translation initiation codon. The sequence of the 5'-flanking region is GC-rich and lacks a typical TATA box at the usual position. However, it has a CAAT sequence at -132 and potential recognition sequences for several transcription factors including SP1, GR-PR-MMTV, AP4, gERE, JCV repeat, AP2, and NF-kappa B. Each of the five transmembrane segments are encoded by five consecutive exons. This corroborates the five-transmembrane model proposed for human, mouse, and Xenopus rather than six proposed for bovine. The cytochrome was found to be highly expressed in colon cancer cell lines, T cell lymphomas, and K-562 cell lines. However, in B-cell lymphomas such as Burkitt's and Daudi, the cytochrome b561 expression was completely shut down. The results in this report are the first to demonstrate the structural organization and regulatory sequences of the cytochrome b561 gene encoding an integral membrane protein of neuroendocrine storage vesicles of neurotransmitters and peptide hormones. Unexpected results on cytochrome b561 expression in cells of lymphocytic origin and its complex regulation in tumor cells provide new insights into cytochrome b561 gene regulation.

Cytochrome b561 is a major transmembrane protein of catecholamine and neuropeptide secretory vesicles. In this report, we describe the cloning and properties of a full-length cDNA encoding human neuroendocrine cytochrome b561 from a human caudate cDNA library and a human peripheral blood genomic library. The human cDNA contains two major transcription start sites and only one translation start site that codes for an apocytochrome b561, which is 22 amino acid residues smaller than the previously deduced amino acid sequence from bovine cDNA. This smaller version of cytochrome b561 may contain only five transmembrane segments rather than the previously proposed six segments. The new model is in agreement with our previous results on transmembrane topology of the gene product. Northern-blot analysis shows an expanded tissue distribution of cytochrome mRNA expression where previous immunological assays were negative. These results support the hypothesis that cytochrome b561 is a marker for peptidergic and adrenergic tissues.