Involvement of 101F6, a Homologue of Cytochrome b561, in the Reduction of Ferric Ions

Abstract

We have isolated and characterized a small transmembrane protein, called 101F6, showing high sequence homology to cytochrome b(561), a protein containing two binding sites for haem. The newly identified 101F6 contains six membrane spanning domains in which conserved histidine residues are located, and has a molecular mass of 25 kDa. When the haem-binding with bacterial expressed 101F6 was examined, the protein bound haem and the deletion of one histidine residue at 149 caused a loss of the binding. 101F6 mRNA was expressed widely in various tissues, and especially abundant in liver, kidney and lung. It was also expressed in several cultured cell lines. The protein expressed from the 101F6 cDNA in Balb/3T3 cells was about 25 kDa in size and was localized in small vesicles, including endosomes and endoplasmic reticulum of the perinuclear region. Comparison of the location of 101F6 with that of transferrin receptor-1 revealed that the localization of 101F6 in small vesicles was not always the same as the localization of transferrin receptor-1, but was similar to that of haem oxygenase-1. The other homologue to cytochrome b(561), SDR-2 was also expressed in the small vesicles similar to the location of 101F6. Finally, reduction of ferric ions as well as of azo-dye increased with 101F6- or SDR-2-expressing cells. Thus, both 101F6 and SDR-2 were localized in small vesicles of cells and played roles in the reduction of ferric ions.

Functional expression and characterization of human 101F6 protein, a homologue of cytochrome b561 and a candidate tumor suppressor gene product

“…It is prospected as one of the important keys for understanding of the tumorigenesis and the tumor development and can be considered as a promising target for cancer therapy [15]. Based on the multiple sequence alignments, the 101F6 protein was, however, only ∼25% homologous to the bovine adrenal cytochrome b 561 [12,20]. The predicted structure of the 101F6 protein consists of six transmembrane helices and four well-conserved histidine residues at the four inner helices, which might act as binding sites for two heme b prosthetic groups [16].…”

“…Mouse 101F6 protein was recently reported to be capable of reducing extracellular ferric ions when expressed in human embryonic kidney HEK293T cells, though the source of electrons for the reported activity was not identified [12]. In their additional experiment, plasmid containing the mouse 101F6 cDNA gene was infected into cultured CHO cells, and the 101F6 protein expression was found to be located in small vesicles and in endoplasmic reticulum [12]. AsA was considered to be the most likely candidate as an electron donor for the reduction of extracellular ferric ions, although their observation was apparently discrepant from their own proposal to have a transplasmamembrane ferric reductase activity.…”

Duodenal Cytochrome b (DCYTB) in Iron Metabolism: An Update on Function and Regulation

“…In addition to DCYTB, additional members of cytochrome b 561 family of enzymes in mammals include lysosomal cytochrome b 561 (CYB561A3) [ 53 , 67 ], stromal cell-derived receptor 2 (SDR2/FRRS1) [ 68 , 69 ] and the putative tumor suppressor, 101F6 (CYB561D2/TSP10) [ 70 , 71 ]. Intriguingly, all proteins have been shown capable of stimulating cellular ferrireduction, and are expressed at the plasma membrane, as well as at intracellular sites [ 38 , 53 , 68 , 72 , 73 ].…”

Functional Assembly of Caenorhabditis elegans Cytochrome b-2 (Cecytb-2) into Phospholipid Bilayer Nanodisc with Enhanced Iron Reductase Activity

“…(TCytb) has an ascorbate-dependent ferric reductase activity [11], although its role in plant cells is not clarified yet. Other members of the mammalian cytochrome b 561 family were also proposed to have a ferric reductase activity [9,12].…”