Identification of Zyklopen, a New Member of the Vertebrate Multicopper Ferroxidase Family, and Characterization in Rodents and Human Cells

Abstract

We previously detected a membrane-bound, copper-containing oxidase that may be involved in iron efflux in BeWo cells, a human placental cell line. We have now identified a gene encoding a predicted multicopper ferroxidase (MCF) with a putative C-terminal membrane-spanning sequence and high sequence identity to hephaestin (Heph) and ceruloplasmin (Cp), the other known vertebrate MCF. Molecular modeling revealed conservation of all type I, II, and III copper-binding sites as well as a putative iron-binding site. Protein expression was observed in multiple diverse mouse tissues, including placenta and mammary gland, and the expression pattern was distinct from that of Cp and Heph. The protein possessed ferroxidase activity, and protein levels decreased in cellular copper deficiency. Knockdown with small interfering RNA in BeWo cells indicates that this gene represents the previously detected oxidase. We propose calling this new member of the MCF family "zyklopen."

Proteínas relacionadas con el metabolismo del hierro corporal

“…La deficiencia de la proteína en humanos, se denomina aceruloplasminemia y conduce a una degeneración tardía de la retina y de los ganglios basales, por la acumulación patológica de hierro en estos tejidos al igual que en el hígado, bazo, páncreas y células del cerebro (9,28,85,87).…”

“…La hefestina (Hph), por su nombre en inglés hephaestin: es una proteína transmembranal con un solo dominio; homóloga de la Cp, la conforman 1158 aa (87)(88), conserva los átomos de cobre y los enlaces disulfuro de la Cp. Se expresa principalmente, en las microvellosidades del enterocito y se colocaliza con la Fpn en la membrana basolateral y de manera intracelular, en un sitio supranuclear de la célula absortiva del intestino, pero en cantidades inapreciables en las células de la cripta intestinal; estos hechos son consistentes con la función del transporte de hierro de la proteína, en el enterocito diferenciado; sin embargo, pequeñas cantidades de la proteína también se encuentran en el bazo, pulmón, placenta y cerebro (52,64,(88)(89)(90).…”

“…La conforman 891 aa (93), presenta un sitio de unión de alta afinidad para el hierro y al igual que la Hph, tiene un segmento transmembrana cerca de la región C terminal; similar a la CP, la proteína zyklopen tiene 6 dominios organizados en un arreglo triangular. La expresión se ha descrito en placenta, en las células del sincitiotrofoblasto, participando en la oxidación del hierro ferroso a hierro férrico en el transporte materno fetal del mineral (66,87). Resultados de inmunohistoquímica en tejidos de ratón han identificado esta proteína en la retina, los testículos, el riñón y el cerebro, pero no en el hígado ni en el enterocito (94).…”

The placenta: the forgotten essential organ of iron transport

“…Zyklopen contains a transmembrane domain and an extracellular ferroxidase domain with appropriate topology to interact with FPN1. 79 Absent in liver and intestine, zyklopen is abundantly expressed in the placenta and has been localized to the labyrinth, spongiotrophoblasts, and yolk sac of mouse placenta. 79 Thus, there is evidence that all 3 mammalian multicopper ferroxidases are expressed in the placenta, but little is known definitively regarding their functions in placental iron transport.…”

“…A third ferroxidase in the placenta has been described by Danzeisen et al, 78 who detected an intracellular, membrane-bound protein in BeWo cells that exhibited ceruloplasmin-like oxidase activity and reacted with a ceruloplasmin antibody. 72 Recently, this group identified zyklopen as a placenta-specific ferroxidase 79 that has approximately 50% protein identity with ceruloplasmin and hephaestin. Zyklopen contains a transmembrane domain and an extracellular ferroxidase domain with appropriate topology to interact with FPN1.…”