Hepcidin Bound to alpha(2)-Macroglobulin Reduces Ferroportin-1 Expression and Enhances Its Activity at Reducing Serum Iron Levels
Abstract
Hepcidin regulates iron metabolism by down-regulating ferroportin-1 (Fpn1). We demonstrated that hepcidin is complexedto the blood transport protein, α 2-macroglobulin (Peslova, G., Petrak, J., Kuzelova, K., Hrdy, I., Halada, P., Kuchel, P.
W., Soe-Lin, S., Ponka, P., Sutak, R., Becker, E., Huang, M. L.,Suryo Rahmanto, Y., Richardson, D.
R., and Vyoral, D. (2009) Blood 113, 6225–6236). However, nothing is known about the mechanism of hepcidin binding to a 2M or the effects of the α 2M hepcidin complex in vivo.
We show that decreased Fpn1 expression can be mediated by hepcidin bound to native α 2M and also, for the first time, hepcidin bound to methylamineactivatedα 2M (α M-MA). Passage of high molecular weight α 2M_hepcidin or α 2M-MA_hepcidin complexes (≈725 kDa) through a Sephadex G-25 size exclusion column retained their ability to decrease Fpn1 expression.
Further studies using ultrafiltration indicated that hepcidin binding to α 2Mand α 2M-MA was labile, resulting in some release from the protein, and this may explain its urinary excretion. To determine whether α 2MMA α hepcidin is delivered to cells via the _2M receptor (Lrp1), we assessed α 2Muptake and Fpn1 expression in Lrp1_/_and Lrp1_/_ cells.
Interestingly, α 2M_hepcidin or α 2M-MA_hepcidin demonstrated similar activities at decreasing Fpn1 expression in Lrp1_/_and Lrp1_/_ cells, indicating that Lrp1 is not essential for Fpn1 regulation. In vivo, hepcidin bound to α 2M or α 2M-MA did not affect plasma clearance of _2M/_2M-MA.
However, serum iron levels were reduced to a significantly greater extent in mice treated with α 2M_hepcidin or α 2M-MA_hepcidin relative to unbound hepcidin. This effect could be mediated by the ability of α 2M or α 2M-MA to retard kidney filtration of bound hepcidin, increasing its half-life.
A model is proposed that suggests that unlike proteases, which are irreversibly bound to activated α 2M, hepcidin remains labile and available to down-regulate Fpn1.