Iron Regulatory Proteins: From Molecular Mechanisms to Drug Development

Recalcati, Stefania; Minotti, Giorgio; Cairo, Gaetano

doi:10.1089/ars.2009.2983

Cited by 106 publications

(105 citation statements)

References 234 publications

(262 reference statements)

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“…Clones and controls were grown for 3 days, and K562 cells treated with 1 mM DFO for 48 h were used as a positive control because DFO treatment has an effect on apoptosis 32 and on iron-dependent TfR1 translation. 33 In the Mt1 clone, the Bcl-xL transcript was not significantly different; however, it was significantly reduced in Mt4 and Mt8 clones (approximately 35%) and in DFO-treated cells (approximately 60%) with respect to the Bcl-xL transcript of the control cells ( Figure 2A Figure 2B). There were not statistically significant differences in IRP1 and IRP2 transcripts between control cells and clones (data not shown).…”

Section: Effect Of Mitochondrial Ferritin Expression On Transcriptionmentioning

confidence: 87%

Over-expression of mitochondrial ferritin affects the JAK2/STAT5 pathway in K562 cells and causes mitochondrial iron accumulation

Santambrogio¹,

Erba²,

Campanella³

et al. 2011

Haematologica

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Mitochondrial ferritin is a nuclear encoded iron-storage protein localized in mitochondria. It has anti-oxidant properties related to its ferroxidase activity, and it is able to sequester iron avidly into the organelle. The protein has a tissue-specific pattern of expression and is also highly expressed in sideroblasts of patients affected by hereditary sideroblastic anemia and by refractory anemia with ringed sideroblasts. The present study examined whether mitochondrial ferritin has a role in the pathogenesis of these diseases. Design and MethodsWe analyzed the effect of mitochondrial ferritin over-expression on the JAK2/STAT5 pathway, on iron metabolism and on heme synthesis in erythroleukemic cell lines. Furthermore its effect on apoptosis was evaluated on human erythroid progenitors. Results Data revealed that a high level of mitochondrial ferritin reduced reactive oxygen species andStat5 phosphorylation while promoting mitochondrial iron loading and cytosolic iron starvation. The decline of Stat5 phosphorylation induced a decrease of the level of anti-apoptotic BclxL transcript compared to that in control cells; however, transferrin receptor 1 transcript increased due to the activation of the iron responsive element/iron regulatory protein machinery. Also, high expression of mitochondrial ferritin increased apoptosis, limited heme synthesis and promoted the formation of Perls-positive granules, identified by electron microscopy as iron granules in mitochondria. ConclusionsOur results provide evidence suggesting that Stat5-dependent transcriptional regulation is displaced by strong cytosolic iron starvation status induced by mitochondrial ferritin. The protein interferes with JAK2/STAT5 pathways and with the mechanism of mitochondrial iron accumulation.Key words: mitochondrial ferritin, iron acculumation, hereditary sideroblastic anemia, refractory anemia with ringed sideblasts, ROS, iron starvation, JAK2/STA5. 2011;96(10):1424-1432 doi:10.3324/haematol.2011 This is an open-access paper. Citation: Santambrogio P, Erba BG, Campanella A, Cozzi A, Causarano V, Cremonesi L, Gallì A, Della Porta MG, Invernizzi R, and Levi S. Over-expression of mitochondrial ferritin affects the JAK2/STAT5 pathway in K562 cells and causes mitochondrial iron accumulation. Haematologica Over-expression of mitochondrial ferritin affects the JAK2/STAT5 pathway in K562 cells and causes mitochondrial iron accumulation

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Section: Effect Of Mitochondrial Ferritin Expression On Transcriptionmentioning

confidence: 87%

Over-expression of mitochondrial ferritin affects the JAK2/STAT5 pathway in K562 cells and causes mitochondrial iron accumulation

Santambrogio¹,

Erba²,

Campanella³

et al. 2011

Haematologica

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“…Depending on iron concentration, Iron Regulatory Proteins (IRP1 and IRP2) co-ordinately modulate iron protein levels post-transcriptionally by binding to Iron Responsive Elements (IREs) on mRNAs of TfR1, ferritin subunits and DMT1 and Fpn IRE-isoforms. 7 In synthesis, high iron reduces the IRP-IRE binding, promoting ferritin and ferroportin synthesis, and in parallel limits iron uptake by reducing TfR1 and DMT1, while low iron does exactly the opposite. Iron excess promotes IRP inactivation by iron-sulphur cluster insertion in the IRP1 molecule and inducing proteasomal degradation of IRP2 protein.…”

Section: Introductionmentioning

confidence: 99%

Iron increases the susceptibility of multiple myeloma cells to bortezomib

et al. 2012

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ABSTRACTbeen reported following proteasome inhibition in cells treated with H 2 O 2 and nitric oxide generators. 13,14 Ferritin might undergo proteasomal degradation to recycle stored iron in cells pre-loaded with ferric ammonium citrate (FAC) 15 in conditions of decreased cell iron content 16,17 or in the presence of oxidative stress stimuli. 18 Here we report the analysis of iron metabolism and the effects of iron manipulation in MM cell lines and primary cells of patients treated with bortezomib. We observed that the basal iron storage capacity of MM cell lines directly correlates with bortezomib resistance and that bortezomib sensitizes MM cells to iron toxicity. Manipulation of iron homeostasis might be a tool to increase the susceptibility of MM cells to the effect of bortezomib and to overcome bortezomib resistance. Design and Methods Cell culture and cellular extractsCell culture media and reagents were from Invitrogen (Karlsruhe, Germany) and from Sigma-Aldrich (St. Louis, MO, USA). Bortezomib was purchased from LC Laboratories (Wobun, MA, USA). Patient-derived plasma cells were purified from bone marrow aspirates by density gradient centrifugation and then by CD138 immunomagnetic positive selection (Miltenyi Biotec, Bergisch Gladbach, Germany). Samples from Patient 2 were collected both at diagnosis (sample a) and relapse (sample b). Informed consent was obtained in accordance with the Declaration of Helsinki and the approval for use of primary samples was obtained from the Institutional Review Board of the San Raffaele Scientific Institute. MM cell lines and primary cells were cultured in RPMI medium supplemented with 10% fetal bovine serum (FBS), 100 units/mL penicillin, 100 mg/mL streptomycin and 2 mM L-glutamine.Cellular extracts were prepared as described in the Online Supplementary Appendix. Iron manipulationsIron removal was obtained by addition of deferoxamine (DFO; Biofutura Pharma, Milan, Italy). Ferric ammonium citrate (FAC), holo-transferrin (FeTf) (Sigma-Aldrich) and N-acetyl Cysteine (NAC) (Sigma-Aldrich) were used for iron loading experiments. FAC and 55 FeAC were prepared by mixing FeCl 3 -HCl (Merk Millipore, Billerica, MA, USA) or 55 FeCl 3 -HCl (PerkinElmer, Monza, Italy) with citric acid at 1:2 ratios and then by adjusting pH to 7.4 with NH 4 OH.To estimate ferritin content in patients' cells, cellular extracts were incubated in vitro with a molar excess of 55 FeAC plus ascorbic acid. Then extracts were loaded without heating on sodium dodecylsulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and exposed to autoradiography. Cell viabilityCell viability was determined by measuring the reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) (Sigma-Aldrich). Briefly, cells were seeded in 96-well plates and assay performed on at least three wells for every condition. MTT was then added for 3 h at a final concentration of 0.25 mg/mL. Plates were centrifuged at 3200g for 5 min and the soluble fraction decanted. Crystal pellets were suspended in DMSO a...

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“…Moreover, the high expression of ferroportin (Fpn), the only known iron exporter from cells, resulted in elevated iron release activity by M2 cells. In line with the expression of Ft and TfR1, the binding activity of the iron regulatory proteins (IRP), which post-transcriptionally regulate the expression of a number of iron genes, 13 was lower in M1 than in M2 cells. Since IRP-binding activity and the labile iron pool are usually inversely related, 14 this result apparently contrasts with the larger labile iron pool (measured by the calcein method) found in M2 cells.…”

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confidence: 99%