Inappropriately High Iron Regulatory Protein Activity in Monocytes of Patients With Genetic Hemochromatosis

Abstract In genetic hemochromatosis (GH), excess iron is deposited in parenchymal cells, whereas little iron is found in reticuloendothelial (RE) cells until the later stages of the disease. As iron absorption is inversely related to RE cells stores, a failure of RE to retain iron has been proposed as the basic defect in GH. In RE cells of GH subjects, we examined the activity of iron regulatory protein (IRP), a reliable indicator of the elusive regulatory labile iron pool, which modulates cellular iron homeostasis through control of ferritin (Ft) and transferrin receptor gene expression. RNA-bandshift assays showed a significant increase in IRP activity in monocytes from 16 patients with untreated GH compared with 28 control subjects (1.5-fold) and five patients with secondary hemochromatosis (SH) with similar iron burden (fourfold). In 17 phlebotomy-treated GH patients, IRP activity did not differ from that of control subjects. In both GH and SH monocyte-macrophages, Ft content increased by twofold and the L subunit-rich isoferritin profile was unchanged as compared with controls. IRP activity was still upregulated in vitro in monocyte-derived macrophages of GH subjects but, following manipulations of iron levels, was modulated normally. Therefore, the sustained activity of monocyte IRP found in vivo in monocytes of GH patients is not due to an inherent defect of its control, but is rather the expression of a critical abnormality of iron metabolism, eg, a paradoxical contraction of the regulatory iron pool. By preventing Ft mRNA translation, high IRP activity in monocytes may represent a molecular mechanism contributing to the inadequate Ft accumulation and insufficient RE iron storage in GH.

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Response of Monocyte Iron Regulatory Protein Activity to Inflammation: Abnormal Behavior in Genetic Hemochromatosis

Blood ◽

10.1182/blood.v91.7.2565 ◽

1998 ◽

Vol 91 (7) ◽

pp. 2565-2572 ◽

Cited By ~ 45

Author(s):

Stefania Recalcati ◽

Roberta Pometta ◽

Sonia Levi ◽

Dario Conte ◽

Gaetano Cairo

Keyword(s):

Iron Homeostasis ◽

Iron Status ◽

Regulatory Protein ◽

Abnormal Behavior ◽

Iron Regulatory Protein ◽

Iron Release ◽

Protein Activity ◽

Intracellular Iron ◽

Ifn Γ ◽

Genetic Hemochromatosis

Abstract In genetic hemochromatosis (GH), iron overload affects mainly parenchymal cells, whereas little iron is found in reticuloendothelial (RE) cells. We previously found that RE cells from GH patients had an inappropriately high activity of iron regulatory protein (IRP), the key regulator of intracellular iron homeostasis. Elevated IRP should reflect a reduction of the iron pool, possibly because of a failure to retain iron. A defect in iron handling by RE cells that results in a lack of feedback regulation of intestinal absorption might be the basic abnormality in GH. To further investigate the capacity of iron retention in RE cells of GH patients, we used inflammation as a model system as it is characterized by a block of iron release from macrophages. We analyzed the iron status of RE cells by assaying IRP activity and ferritin content after 4, 8, and 24 hours of incubation with lipopolysaccharide (LPS) and interferon-γ (IFN-γ). RNA-bandshift assays showed that in monocytes and macrophages from 16 control subjects, IRP activity was transiently elevated 4 hours after treatment with LPS and IFN-γ but remarkably downregulated thereafter. Treatment with NO donors produced the same effects whereas an inducible Nitric Oxide Synthase (iNOS) inhibitor prevented them, which suggests that the NO pathway was involved. Decreased IRP activity was also found in monocytes from eight patients with inflammation. Interestingly, no late decrease of IRP activity was detected in cytokine-treated RE cells from 12 GH patients. Ferritin content was increased 24 hours after treatment in monocytes from normal subjects but not in monocytes from GH patients. The lack of downregulation of IRP activity under inflammatory conditions seems to confirm that the control of iron release from RE cells is defective in GH.

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Impaired Ferritin mRNA Translation in Primary Erythroid Progenitors: Shift to Iron-Dependent Regulation by the v-ErbA Oncoprotein

Blood ◽

10.1182/blood.v94.12.4321 ◽

1999 ◽

Vol 94 (12) ◽

pp. 4321-4332 ◽

Cited By ~ 4

Author(s):

Wolfgang Mikulits ◽

Matthias Schranzhofer ◽

Anton Bauer ◽

Helmut Dolznig ◽

Lioba Lobmayr ◽

...

Keyword(s):

Regulatory Protein ◽

Mrna Translation ◽

Erythroid Differentiation ◽

Binding Activity ◽

Nuclear Hormone Receptor ◽

Erythroid Progenitors ◽

Iron Storage ◽

Iron Administration ◽

Key Enzyme

Abstract In immortalized cells of the erythroid lineage, the iron-regulatory protein (IRP) has been suggested to coregulate biosynthesis of the iron storage protein ferritin and the erythroid delta-aminolevulinate synthase (eALAS), a key enzyme in heme production. Under iron scarcity, IRP binds to an iron-responsive element (IRE) located in ferritin and eALAS mRNA leaders, causing a block of translation. In contrast, IRP-IRE interaction is reduced under high iron conditions, allowing efficient translation. We show here that primary chicken erythroblasts (ebls) proliferating or differentiating in culture use a drastically different regulation of iron metabolism. Independently of iron administration, ferritin H (ferH) chain mRNA translation was massively decreased, whereas eALAS transcripts remained constitutively associated with polyribosomes, indicating efficient translation. Variations in iron supply had minor but significant effects on eALAS mRNA polysome recruitment but failed to modulate IRP-affinity to the ferH-IRE in vitro. However, leukemic ebls transformed by the v-ErbA/v-ErbB–expressing avian erythroblastosis virus showed an iron-dependent reduction of IRP mRNA-binding activity, resulting in mobilization of ferH mRNA into polysomes. Hence, we analyzed a panel of ebls overexpressing v-ErbA and/or v-ErbB oncoproteins as well as the respective normal cellular homologues (c-ErbA/TR, c-ErbB/EGFR). It turned out that v-ErbA, a mutated class II nuclear hormone receptor that arrests erythroid differentiation, caused the change in ferH mRNA translation. Accordingly, inhibition of v-ErbA function in these leukemic ebls led to a switch from iron-responsive to iron-independent ferH expression.

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Response of Monocyte Iron Regulatory Protein Activity to Inflammation: Abnormal Behavior in Genetic Hemochromatosis

Blood ◽

10.1182/blood.v91.7.2565.2565_2565_2572 ◽

1998 ◽

Vol 91 (7) ◽

pp. 2565-2572

Author(s):

Stefania Recalcati ◽

Roberta Pometta ◽

Sonia Levi ◽

Dario Conte ◽

Gaetano Cairo

Keyword(s):

Iron Homeostasis ◽

Iron Status ◽

Regulatory Protein ◽

Abnormal Behavior ◽

Iron Regulatory Protein ◽

Iron Release ◽

Protein Activity ◽

Intracellular Iron ◽

Ifn Γ ◽

Genetic Hemochromatosis

In genetic hemochromatosis (GH), iron overload affects mainly parenchymal cells, whereas little iron is found in reticuloendothelial (RE) cells. We previously found that RE cells from GH patients had an inappropriately high activity of iron regulatory protein (IRP), the key regulator of intracellular iron homeostasis. Elevated IRP should reflect a reduction of the iron pool, possibly because of a failure to retain iron. A defect in iron handling by RE cells that results in a lack of feedback regulation of intestinal absorption might be the basic abnormality in GH. To further investigate the capacity of iron retention in RE cells of GH patients, we used inflammation as a model system as it is characterized by a block of iron release from macrophages. We analyzed the iron status of RE cells by assaying IRP activity and ferritin content after 4, 8, and 24 hours of incubation with lipopolysaccharide (LPS) and interferon-γ (IFN-γ). RNA-bandshift assays showed that in monocytes and macrophages from 16 control subjects, IRP activity was transiently elevated 4 hours after treatment with LPS and IFN-γ but remarkably downregulated thereafter. Treatment with NO donors produced the same effects whereas an inducible Nitric Oxide Synthase (iNOS) inhibitor prevented them, which suggests that the NO pathway was involved. Decreased IRP activity was also found in monocytes from eight patients with inflammation. Interestingly, no late decrease of IRP activity was detected in cytokine-treated RE cells from 12 GH patients. Ferritin content was increased 24 hours after treatment in monocytes from normal subjects but not in monocytes from GH patients. The lack of downregulation of IRP activity under inflammatory conditions seems to confirm that the control of iron release from RE cells is defective in GH.

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Impaired Ferritin mRNA Translation in Primary Erythroid Progenitors: Shift to Iron-Dependent Regulation by the v-ErbA Oncoprotein

Blood ◽

10.1182/blood.v94.12.4321.424k15_4321_4332 ◽

1999 ◽

Vol 94 (12) ◽

pp. 4321-4332

Author(s):

Wolfgang Mikulits ◽

Matthias Schranzhofer ◽

Anton Bauer ◽

Helmut Dolznig ◽

Lioba Lobmayr ◽

...

Keyword(s):

Regulatory Protein ◽

Mrna Translation ◽

Erythroid Differentiation ◽

Binding Activity ◽

Nuclear Hormone Receptor ◽

Erythroid Progenitors ◽

Iron Storage ◽

Iron Administration ◽

Key Enzyme

In immortalized cells of the erythroid lineage, the iron-regulatory protein (IRP) has been suggested to coregulate biosynthesis of the iron storage protein ferritin and the erythroid delta-aminolevulinate synthase (eALAS), a key enzyme in heme production. Under iron scarcity, IRP binds to an iron-responsive element (IRE) located in ferritin and eALAS mRNA leaders, causing a block of translation. In contrast, IRP-IRE interaction is reduced under high iron conditions, allowing efficient translation. We show here that primary chicken erythroblasts (ebls) proliferating or differentiating in culture use a drastically different regulation of iron metabolism. Independently of iron administration, ferritin H (ferH) chain mRNA translation was massively decreased, whereas eALAS transcripts remained constitutively associated with polyribosomes, indicating efficient translation. Variations in iron supply had minor but significant effects on eALAS mRNA polysome recruitment but failed to modulate IRP-affinity to the ferH-IRE in vitro. However, leukemic ebls transformed by the v-ErbA/v-ErbB–expressing avian erythroblastosis virus showed an iron-dependent reduction of IRP mRNA-binding activity, resulting in mobilization of ferH mRNA into polysomes. Hence, we analyzed a panel of ebls overexpressing v-ErbA and/or v-ErbB oncoproteins as well as the respective normal cellular homologues (c-ErbA/TR, c-ErbB/EGFR). It turned out that v-ErbA, a mutated class II nuclear hormone receptor that arrests erythroid differentiation, caused the change in ferH mRNA translation. Accordingly, inhibition of v-ErbA function in these leukemic ebls led to a switch from iron-responsive to iron-independent ferH expression.

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ENO1 suppresses cancer cell ferroptosis by degrading the mRNA of iron regulatory protein 1

10.21203/rs.3.rs-79778/v1 ◽

2020 ◽

Author(s):

Huafeng Zhang ◽

Tong Zhang ◽

Linchong Sun ◽

Yijie Hao ◽

Wenhao Ma ◽

...

Keyword(s):

Iron Homeostasis ◽

Rna Binding ◽

Clinical Sample ◽

Regulatory Protein ◽

Iron Regulatory Protein ◽

Aberrant Expression ◽

Hcc Cells ◽

Iron Regulatory Protein 1

Abstract α-Enolase 1 (ENO1) is a critical glycolytic enzyme whose aberrant expression drives the pathogenesis of various cancers. ENO1 has been indicated to have additional roles beyond its conventional metabolic activity, but the underlying mechanisms and biological consequences remain elusive. Here, we show that ENO1 suppresses iron regulatory protein 1 (IRP1) expression to regulate iron homeostasis and survival of hepatocellular carcinoma (HCC) cells. Mechanistically, we unprecedentedly uncover that ENO1, as an RNA-binding protein, recruits CNOT6 to accelerate the mRNA decay of IRP1 in cancer cells, leading to inhibition of mtioferin-1 (Mfrn1) expression and subsequent repression of mitochondrial iron-induced ferroptosis. Moreover, through in vitro and in vivo experiments and clinical sample analysis, we identified IRP1 and Mfrn1 as tumor suppressors by inducing ferroptosis in HCC cells. Taken together, this study establishes a novel role for the ENO1/IRP1/Mfrn1 pathway in the pathogenesis of HCC and reveals a previously unknown connection between the ENO1/IRP1/Mfrn1 pathway and ferroptosis, suggesting a potential innovative cancer therapy.

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Glycogen branching enzyme controls cellular iron homeostasis via Iron Regulatory Protein 1 and mitoNEET

Nature Communications ◽

10.1038/s41467-019-13237-8 ◽

2019 ◽

Vol 10 (1) ◽

Cited By ~ 10

Author(s):

Nhan Huynh ◽

Qiuxiang Ou ◽

Pendleton Cox ◽

Roland Lill ◽

Kirst King-Jones

Keyword(s):

Iron Homeostasis ◽

Nuclear Translocation ◽

Regulatory Protein ◽

Glycogen Metabolism ◽

Glycogen Storage Disease Type ◽

Iron Regulatory Protein ◽

Messenger Rnas ◽

Branching Enzyme ◽

Cellular Iron ◽

Iron Regulatory Protein 1

AbstractIron Regulatory Protein 1 (IRP1) is a bifunctional cytosolic iron sensor. When iron levels are normal, IRP1 harbours an iron-sulphur cluster (holo-IRP1), an enzyme with aconitase activity. When iron levels fall, IRP1 loses the cluster (apo-IRP1) and binds to iron-responsive elements (IREs) in messenger RNAs (mRNAs) encoding proteins involved in cellular iron uptake, distribution, and storage. Here we show that mutations in the Drosophila 1,4-Alpha-Glucan Branching Enzyme (AGBE) gene cause porphyria. AGBE was hitherto only linked to glycogen metabolism and a fatal human disorder known as glycogen storage disease type IV. AGBE binds specifically to holo-IRP1 and to mitoNEET, a protein capable of repairing IRP1 iron-sulphur clusters. This interaction ensures nuclear translocation of holo-IRP1 and downregulation of iron-dependent processes, demonstrating that holo-IRP1 functions not just as an aconitase, but throttles target gene expression in anticipation of declining iron requirements.

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Induction of iron regulatory protein 1 RNA-binding activity by nitric oxide is associated with a concomitant increase in the labile iron pool: implications for DNA damage

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2004.12.012 ◽

2005 ◽

Vol 327 (1) ◽

pp. 349-355 ◽

Cited By ~ 20

Author(s):

Pawel Lipinski ◽

Rafal R. Starzynski ◽

Jean-Claude Drapier ◽

Cecile Bouton ◽

Teresa Bartlomiejczyk ◽

...

Keyword(s):

Nitric Oxide ◽

Dna Damage ◽

Rna Binding ◽

Regulatory Protein ◽

Binding Activity ◽

Iron Regulatory Protein ◽

Concomitant Increase ◽

Labile Iron Pool ◽

Labile Iron ◽

Iron Pool

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Dysregulated monocyte iron homeostasis and erythropoietin formation in patients with anemia of chronic disease

Blood ◽

10.1182/blood-2005-08-3364 ◽

2006 ◽

Vol 107 (10) ◽

pp. 4142-4148 ◽

Cited By ~ 107

Author(s):

Igor Theurl ◽

Verena Mattle ◽

Markus Seifert ◽

Mariagabriella Mariani ◽

Christian Marth ◽

...

Keyword(s):

Chronic Disease ◽

Iron Homeostasis ◽

Hereditary Spherocytosis ◽

Regulatory Protein ◽

Significant Negative Correlation ◽

Deficiency Anemia ◽

Anemia Of Chronic Disease ◽

Iron Storage ◽

Iron Retention ◽

Reduced Activity

Anemia of chronic disease (ACD) is frequently found in patients with chronic immune activation. Since most studies on ACD pathophysiology were performed with cell culture or animal models but not in humans, we examined 37 ACD patients suffering from autoimmune diseases or infections, 10 subjects with iron-deficiency anemia (IDA), 10 anemic patients with hereditary spherocytosis (HS), and 27 age-matched controls. Although hemoglobin concentrations were comparable between ACD and IDA patients, the latter presented with significantly higher serum erythropoietin concentrations than ACD patients. The significant negative correlation between erythropoietin and hemoglobin levels observed in IDA patients was also found in a group of anemic but not hypoferremic hereditary spherocytosis subjects, but not in ACD patients. Increased serum concentrations of the hepcidin precursor prohepcidin were paralleled by a decreased expression of the iron exporter ferroportin in circulating monocytes of ACD patients. In the latter cells, increased amounts of the iron storage protein ferritin and a reduced activity of iron-regulatory protein indicated monocyte iron accumulation. Our data indicate that hypoferremia in ACD may result from downregulation of ferroportin expression by hepcidin and cytokines with subsequent iron retention in monocytes. Together with a diminished erythropoietin formation, the impaired iron recirculation from monocytes may be central in the pathophysiology of ACD in humans.

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Essential Role of Ferritin Pfr in Helicobacter pylori Iron Metabolism and Gastric Colonization

Infection and Immunity ◽

10.1128/iai.70.7.3923-3929.2002 ◽

2002 ◽

Vol 70 (7) ◽

pp. 3923-3929 ◽

Cited By ~ 80

Author(s):

Barbara Waidner ◽

Stefan Greiner ◽

Stefan Odenbreit ◽

Holger Kavermann ◽

Jyoti Velayudhan ◽

...

Keyword(s):

Helicobacter Pylori ◽

Iron Metabolism ◽

Essential Element ◽

Iron Starvation ◽

Iron Storage ◽

Gastric Colonization ◽

H Pylori ◽

Iron Pool

ABSTRACT The reactivity of the essential element iron necessitates a concerted expression of ferritins, which mediate iron storage in a nonreactive state. Here we have further established the role of the Helicobacter pylori ferritin Pfr in iron metabolism and gastric colonization. Iron stored in Pfr enabled H. pylori to multiply under severe iron starvation and protected the bacteria from acid-amplified iron toxicity, as inactivation of the pfr gene restricted growth of H. pylori under these conditions. The lowered total iron content in the pfr mutant, which is probably caused by decreased iron uptake rates, was also reflected by an increased resistance to superoxide stress. Iron induction of Pfr synthesis was clearly diminished in an H. pylori feoB mutant, which lacked high-affinity ferrous iron transport, confirming that Pfr expression is mediated by changes in the cytoplasmic iron pool and not by extracellular iron. This is well in agreement with the recent discovery that iron induces Pfr synthesis by abolishing Fur-mediated repression of pfr transcription, which was further confirmed here by the observation that iron inhibited the in vitro binding of recombinant H. pylori Fur to the pfr promoter region. The functions of H. pylori Pfr in iron metabolism are essential for survival in the gastric mucosa, as the pfr mutant was unable to colonize in a Mongolian gerbil-based animal model. In summary, the pfr phenotypes observed give new insights into prokaryotic ferritin functions and indicate that iron storage and homeostasis are of extraordinary importance for H. pylori to survive in its hostile natural environment.

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