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Essential role of endocytosis of the type II transmembrane serine protease TMPRSS6 in regulating its functionality.

ABSTRACT: The type II transmembrane serine protease TMPRSS6 (also known as matriptase-2) controls iron homeostasis through its negative regulation of expression of hepcidin, a key hormone involved in iron metabolism. Upstream of the hepcidin-regulated signaling pathway, TMPRSS6 cleaves its target substrate hemojuvelin (HJV) at the plasma membrane, but the dynamics of the cell-surface expression of the protease have not been addressed. Here, we report that TMPRSS6 undergoes constitutive internalization in transfected HEK293 cells and in two human hepatic cell lines, HepG2 and primary hepatocytes, both of which express TMPRSS6 endogenously. Cell surface-labeled TMPRSS6 was internalized and was detected in clathrin- and AP-2-positive vesicles via a dynamin-dependent pathway. The endocytosed TMPRSS6 next transited in early endosomes and then to lysosomes. Internalization of TMPRSS6 is dependent on specific residues within its N-terminal cytoplasmic domain, as site-directed mutagenesis of these residues abrogated internalization and maintained the enzyme at the cell surface. Cells coexpressing these mutants and HJV produced significantly decreased levels of hepcidin compared with wild-type TMPRSS6 due to the sustained cleavage of HJV at the cell surface by TMPRSS6 mutants. Our results underscore for the first time the importance of TMPRSS6 trafficking at the plasma membrane in the regulation of hepcidin expression, an event that is essential for iron homeostasis.

SUBMITTER: Beliveau F

PROVIDER: S-EPMC3190711 | biostudies-literature | 2011 Aug

REPOSITORIES: biostudies-literature

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Essential role of endocytosis of the type II transmembrane serine protease TMPRSS6 in regulating its functionality.

Béliveau François F Brulé Cédric C Désilets Antoine A Zimmerman Brandon B Laporte Stéphane A SA Lavoie Christine L CL Leduc Richard R

The Journal of biological chemistry 20110701 33

The type II transmembrane serine protease TMPRSS6 (also known as matriptase-2) controls iron homeostasis through its negative regulation of expression of hepcidin, a key hormone involved in iron metabolism. Upstream of the hepcidin-regulated signaling pathway, TMPRSS6 cleaves its target substrate hemojuvelin (HJV) at the plasma membrane, but the dynamics of the cell-surface expression of the protease have not been addressed. Here, we report that TMPRSS6 undergoes constitutive internalization in ...[more]

PMID: 21724843

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Project description:PURPOSE:Matriptase-2 (also known as TMPRSS6) is a critical regulator of the iron-regulatory hormone hepcidin in the liver; matriptase-2 cleaves membrane-bound hemojuvelin and consequently alters bone morphogenetic protein (BMP) signaling. Hemojuvelin and hepcidin are expressed in the retina and play a critical role in retinal iron homeostasis. However, no information on the expression and function of matriptase-2 in the retina is available. The purpose of the present study was to examine the retinal expression of matriptase-2 and its role in retinal iron homeostasis. METHODS:RT-PCR, quantitative PCR (qPCR), and immunofluorescence were used to analyze the expression of matriptase-2 and other iron-regulatory proteins in the mouse retina. Polarized localization of matriptase-2 in the RPE was evaluated using markers for the apical and basolateral membranes. Morphometric analysis of retinas from wild-type and matriptase-2 knockout (Tmprss6(msk/msk) ) mice was also performed. Retinal iron status in Tmprss6(msk/msk) mice was evaluated by comparing the expression levels of ferritin and transferrin receptor 1 between wild-type and knockout mice. BMP signaling was monitored by the phosphorylation status of Smads1/5/8 and expression levels of Id1 while interleukin-6 signaling was monitored by the phosphorylation status of STAT3. RESULTS:Matriptase-2 is expressed in the mouse retina with expression detectable in all retinal cell types. Expression of matriptase-2 is restricted to the apical membrane in the RPE where hemojuvelin, the substrate for matriptase-2, is also present. There is no marked difference in retinal morphology between wild-type mice and Tmprss6(msk/msk) mice, except minor differences in specific retinal layers. The knockout mouse retina is iron-deficient, demonstrable by downregulation of the iron-storage protein ferritin and upregulation of transferrin receptor 1 involved in iron uptake. Hepcidin is upregulated in Tmprss6(msk/msk) mouse retinas, particularly in the neural retina. BMP signaling is downregulated while interleukin-6 signaling is upregulated in Tmprss6(msk/msk) mouse retinas, suggesting that the upregulaton of hepcidin in knockout mouse retinas occurs through interleukin-6 signaling and not through BMP signaling. CONCLUSIONS:The iron-regulatory serine protease matriptase-2 is expressed in the retina, and absence of this enzyme leads to iron deficiency and increased expression of hemojuvelin and hepcidin in the retina. The upregulation of hepcidin expression in Tmprss6(msk/msk) mouse retinas does not occur via BMP signaling but likely via the proinflammatory cytokine interleukin-6. We conclude that matriptase-2 is a critical participant in retinal iron homeostasis.

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Essential role of endocytosis of the type II transmembrane serine protease TMPRSS6 in regulating its functionality.

Publications

Essential role of endocytosis of the type II transmembrane serine protease TMPRSS6 in regulating its functionality.

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