CD63 is Regulated by Iron via the IRE-IRP System and is Important for Ferritin Secretion by Extracellular Vesicles

Extracellular vesicles (EVs) transfer functional molecules between cells. CD63 is a widely recognized EV marker that contributes to EV secretion from cells. However, the regulation of its expression remains largely unknown. Ferritin is a cellular iron storage protein that can be also secreted by the exosome pathway (Truman-Rosentsvit M. et al. BLOOD 131 (2018) 342-352), with serum ferritin levels classically reflecting body iron stores. Iron metabolism-associated proteins, such as ferritin, are intricately regulated by cellular iron levels via the iron responsive element (IRE)-iron regulatory protein (IRP) system. Herein, we present a novel mechanism demonstrating that the expression of the EV-associated protein, CD63, is under the regulation of the IRE-IRP system. We discovered a canonical IRE in the 5'-untranslated region (UTR) of CD63 mRNA responsible for regulating its expression in response to increased iron. Cellular iron-loading caused a marked increase in CD63 expression and the secretion from cells of CD63 positive (i.e., CD63(+)) EVs, which were shown to contain ferritin-H (FtH) and -L (FtL). Our results demonstrate that under iron-loading, intracellular ferritin is transferred via nuclear receptor coactivator 4 (NCOA4) to CD63(+) EVs that are then secreted. Such iron-regulated secretion of the major iron storage protein ferritin via CD63(+) EVs, poses significant impact for understanding the local cell-to-cell exchange of ferritin and iron.

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Ferritin — The Structure and Function of an Iron Storage Protein

The Biological Chemistry of Iron ◽

10.1007/978-94-009-7882-9_4 ◽

1982 ◽

pp. 45-61 ◽

Cited By ~ 1

Author(s):

Robert R. Crichton

Keyword(s):

Storage Protein ◽

Structure And Function ◽

Iron Storage ◽

And Function ◽

Iron Storage Protein

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Analysis of Iron in Ferritin, the Iron-Storage Protein: A General Chemistry Experiment

Journal of Chemical Education ◽

10.1021/ed075p437 ◽

1998 ◽

Vol 75 (4) ◽

pp. 437 ◽

Cited By ~ 17

Author(s):

Maureen J. Donlin ◽

Regina F. Frey ◽

Christopher Putnam ◽

Jody Proctor ◽

James K. Bashkin

Keyword(s):

General Chemistry ◽

Storage Protein ◽

Protein A ◽

Iron Storage ◽

Chemistry Experiment ◽

Iron Storage Protein

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Preparation of platinum nanoparticles using iron(ii) as reductant and photosensitized H2 generation on an iron storage protein scaffold

RSC Advances ◽

10.1039/d0ra00341g ◽

2020 ◽

Vol 10 (10) ◽

pp. 5551-5559

Author(s):

Brenda S. Benavides ◽

Silvano Valandro ◽

Donald M. Kurtz

Keyword(s):

Heavy Chain ◽

Platinum Nanoparticles ◽

Storage Protein ◽

Iron Storage ◽

Protein Scaffold ◽

H2 Generation ◽

Iron Storage Protein

An assembly of platinum nanoparticles produced by Fe(ii) reduction of Pt(ii) and stabilized by human heavy chain ferritin's native catalysis of Fe(ii)(aq) autoxidation functions as an efficient photosensitized H2 evolution catalyst.

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Growth of carbon nanotubes on a gold (111) surface using two-dimensional iron oxide nano-particle catalysts derived from iron storage protein

Chemical Communications ◽

10.1039/b402984d ◽

2004 ◽

pp. 1518 ◽

Cited By ~ 27

Author(s):

Masato Tominaga ◽

Akihiro Ohira ◽

Atsushi Kubo ◽

Isao Taniguchi ◽

Masashi Kunitake

Keyword(s):

Carbon Nanotubes ◽

Iron Oxide ◽

Storage Protein ◽

Two Dimensional ◽

Nano Particle ◽

Iron Storage ◽

Iron Storage Protein

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Overexpression of Drosophila mitoferrin in l(2)mbn cells results in dysregulation of Fer1HCH expression

Biochemical Journal ◽

10.1042/bj20082231 ◽

2009 ◽

Vol 421 (3) ◽

pp. 463-471 ◽

Cited By ~ 36

Author(s):

Christoph Metzendorf ◽

Wenlin Wu ◽

Maria I. Lind

Keyword(s):

Cell Lines ◽

Iron Content ◽

Regulatory Protein ◽

Control Cell ◽

Iron Loading ◽

Protein Maturation ◽

Normal Medium ◽

Aconitase Activity ◽

Cellular Iron ◽

Mitochondrial Transporter

Mrs3p and Mrs4p (Mrs3/4p) are yeast mitochondrial iron carrier proteins that play important roles in ISC (iron-sulphur cluster) and haem biosynthesis. At low iron conditions, mitochondrial and cytoplasmic ISC protein maturation is correlated with MRS3/4 expression. Zebrafish mitoferrin1 (mfrn1), one of two MRS3/4 orthologues, is essential for erythropoiesis, but little is known about the ubiquitously expressed paralogue mfrn2. In the present study we identified a single mitoferrin gene (dmfrn) in the genome of Drosophila melanogaster, which is probably an orthologue of mfrn2. Overexpression of dmfrn in the Drosophila l(2)mbn cell line (mbn-dmfrn) resulted in decreased binding between IRP-1A (iron regulatory protein 1A) and stem-loop RNA structures referred to as IREs (iron responsive elements). mbn-dmfrn cell lines also had increased cytoplasmic aconitase activity and slightly decreased iron content. In contrast, iron loading results in decreased IRP-1A–IRE binding, but increased cellular iron content, in experimental mbn-dmfrn and control cell lines. Iron loading also increases cytoplasmic aconitase activity in all cell lines, but with slightly higher activity observed in mbn-dmfrn cells. From this we concluded that dmfrn overexpression stimulates cytoplasmic ISC protein maturation, as has been reported for MRS3/4 overexpression. Compared with control cell lines, mbn-dmfrn cells had higher Fer1HCH (ferritin 1 heavy chain homologue) transcript and protein levels. RNA interference of the putative Drosophila orthologue of human ABCB7, a mitochondrial transporter involved in cytoplasmic ISC protein maturation, restored Fer1HCH transcript levels of iron-treated mbn-dmfrn cells to those of control cells grown in normal medium. These results suggest that dmfrn overexpression in l(2)mbn cells causes an ‘overestimation’ of the cellular iron content, and that regulation of Fer1HCH transcript abundance probably depends on cytoplasmic ISC protein maturation.

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Ferritin does not accumulate iron oxidized by caeruloplasmin

Biochemical Journal ◽

10.1042/bj3050021 ◽

1995 ◽

Vol 305 (1) ◽

pp. 21-23 ◽

Cited By ~ 9

Author(s):

A Treffry ◽

D Gelvan ◽

A M Konijn ◽

P M Harrison

Keyword(s):

Dissolved Oxygen ◽

Storage Protein ◽

Iron Oxidation ◽

Primary Effect ◽

Significant Extent ◽

Iron Storage ◽

Horse Spleen Ferritin ◽

Iron Storage Protein

Ferritin is an iron-storage protein ubiquitous in mammals, plants and bacteria. It can be reconstituted, in vitro, from the apoprotein and Fe(II) salts in the presence of dissolved oxygen. Recently it has been reported that caeruloplasmin can facilitate apoferritin reconstitution and that iron oxidized by caeruloplasmin is sequestered within the ferritin shell. Here we show that the primary effect of adding caeruloplasmin to horse spleen ferritin during reconstitution is the competition between the two molecules for the iron. This competition results in overall increased rates of iron oxidation and a mixture of products, namely iron-containing ferritin and iron hydroxy polymers attached to caeruloplasmin. Iron oxidized by caeruloplasmin is not incorporated, to any significant extent, into horse spleen ferritin.

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