scholarly journals MASL1 Is a Critical Modifier of Hepcidin Expression Via Smad1/5/8 Phosphorylation in the Bone Morphogenetic Protein (BMP) Signaling Pathway

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 1275-1275
Author(s):  
Chutima Kumkhaek ◽  
Christian LaChance ◽  
Wulin Aerbajinai ◽  
Jianqiong Zhu ◽  
Griffin P. Rodgers
Keyword(s):  
Iron Homeostasis ◽  
Conflicts Of Interest ◽  
Lentiviral Vector ◽  
Bmp Signaling ◽  
Down Regulation ◽  
Induced Differentiation ◽  
Hepcidin Expression ◽  
Cd34 Cells ◽  
Huh7 Cells ◽  
Lps Treatment

Abstract Iron is essential for hemoglobin synthesis during terminal erythropoiesis. Hepcidin is the main regulator of iron homeostasis and is repressed by erythropoiesis. Although several candidates have been proposed to act as hepcidin inhibitors and erythroid regulators such as growth differentiation factor 15 (GDF15), twisted gastrulation BMP signaling modulator 1 (TWSG1) or erythroferrone (ERFE), their role in hepcidin repression during erythropoiesis is still unclear. We previously demonstrated that malignant fibrous histiocytoma-amplified sequence 1 (MASL1) is important for terminal erythropoiesis. In addition, down-regulation of MASL1 expression in macrophages strongly enhances IL-6 production following LPS or poly IC stimulation. Therefore, we hypothesized that MASL1 could directly (or indirectly) influence hepcidin expression. We found that endogenous MASL1 expression was significantly decreased in CD34+ cells treated with IL-6 compared with EPO-treated CD34+ cells at day 3 (0.35±0.05 fold vs 2.34±0.13 fold, P=0.002) and day 7 (1.03±0.75 fold vs 205.31±10.83 fold, P=0.001) of differentiation. In contrast, endogenous hepcidin expression was markedly increased in CD34+ cells treated with IL-6 compared with EPO-treated CD34+ cells. Interestingly, an increased hepcidin expression was detected in MASL1-knockdown CD34+ cells at day 3 of EPO-induced differentiation when compared with mock (47.47±23.49 fold vs 2.10±2.46 fold, P=0.029) or control lentiviral vector (47.47±23.49 fold vs 2.54±1.29 fold, P=0.029). Of note, ERFE, GDF15 and TWSG1 expression were decreased in MASL1-knockdown CD34+ cells at day 3 of EPO-induced differentiation. In addition, endogenous MASL1 expression is down-regulation after LPS treatment in PMA-induced THP1 cells but IL-6 is enhanced in MASL1-knockdown-PMA-induced THP1 cells after LPS treatment. In human hepatic cells (Huh-7), we found a significant decrease in hepcidin expression in MASL1-overexpressed Huh-7 cells after BMP2 (5.85±0.32 fold vs 9.98±0.97 fold, P=0.028) or BMP6 (10.07±1.35 fold vs 20.3±0.75 fold, P=0.007) stimulation for 24 hrs. Moreover, up-regulation of MASL1 enhanced the phosphorylation of Erk1/2 proteins while inhibited the phosphorylation of Smad1/5/8 proteins in Huh-7 cells after BMP2 or BMP6 stimulation for 1 hr that consequently affect in down-regulation of hepcidin expression. Strikingly, MASL1 overexpressed-Huh7 cells showed moderately decreased nuclear localization of phospho-Smad1/5/8 after BMP2 or BMP6 treatment. Taken together, these data demonstrate that MASL1 is a critical modifier of hepcidin expression potentially via additional mechanisms related to erythropoiesis and body iron homeostasis. Further clarification of these pathways may be the useful in developing novel treatment of anemias or iron disorders. Disclosures No relevant conflicts of interest to declare.

Hepcidin inhibition improves iron homeostasis in ferrous sulfate and LPS treatment model in mice

Drug Research ◽  
2021 ◽  
Author(s):  
Vishal Patel ◽  
Amit Joharapurkar ◽  
Samadhan Kshirsagar ◽  
Maulik Patel ◽  
Hiren Patel ◽  
...  
Keyword(s):  
Ferrous Sulfate ◽  
Serum Iron ◽  
Iron Homeostasis ◽  
The Body ◽  
Rapid Screening ◽  
Iron Dextran ◽  
Liver Iron ◽  
Hepcidin Expression ◽  
Serum Hepcidin ◽  
Lps Treatment

Abstract Background Hepcidin, a liver-derived peptide, regulates the absorption, distribution, and circulation of iron in the body. Inflammation or iron overload stimulates hepcidin release, which causes the accumulation of iron in tissues. The inadequate levels of iron in circulation impair erythropoiesis. Inhibition of hepcidin may increase iron in circulation and improve efficient erythropoiesis. Activin-like kinase (ALK) inhibitors decrease hepcidin. Methods In this work, we have investigated an ALK inhibitor LDN193189 for its efficacy in iron homeostasis. The effect of LDN193189 treatment was assessed in C57BL6/J mice, in which hepcidin was induced by either ferrous sulfate or lipopolysaccharide (LPS) injection. Results After two hours of treatment, ferrous sulfate increased serum and liver iron, serum hepcidin, and liver hepcidin expression. On the other hand, LPS reduced serum iron in a dose-related manner after six hours of treatment. LDN193189 treatment increased serum iron, decreased spleen and liver iron, decreased serum hepcidin and liver hepcidin expression in ferrous sulfate-treated mice, and increased serum iron in LPS-induced hypoferremia. We observed that ferrous sulfate caused a significantly higher increase in liver iron, serum hepcidin, and liver hepcidin than turpentine oil or LPS in mice. Iron dextran (intraperitoneal or intravenous) increased serum iron, but LDN193189 did not show hyperferremia with iron dextran stimulus. Conclusion Ferrous sulfate-induced hyperferremia can be a valuable and rapid screening model for assessing the efficacy of hepcidin inhibitors.

scholarly journals Inhibition of bone morphogenetic protein signaling attenuates anemia associated with inflammation

Blood ◽  
2011 ◽  
Vol 117 (18) ◽  
pp. 4915-4923 ◽  
Author(s):  
Andrea U. Steinbicker ◽  
Chetana Sachidanandan ◽  
Ashley J. Vonner ◽  
Rushdia Z. Yusuf ◽  
Donna Y. Deng ◽  
...  
Keyword(s):  
Bone Morphogenetic Protein ◽  
Iron Homeostasis ◽  
Iron Bioavailability ◽  
Bmp Signaling ◽  
Neoplastic Disease ◽  
Type I ◽  
Hepcidin Expression ◽  
Hepatic Expression ◽  
Morphogenetic Protein ◽  
Anemia Of Inflammation

Abstract Anemia of inflammation develops in settings of chronic inflammatory, infectious, or neoplastic disease. In this highly prevalent form of anemia, inflammatory cytokines, including IL-6, stimulate hepatic expression of hepcidin, which negatively regulates iron bioavailability by inactivating ferroportin. Hepcidin is transcriptionally regulated by IL-6 and bone morphogenetic protein (BMP) signaling. We hypothesized that inhibiting BMP signaling can reduce hepcidin expression and ameliorate hypoferremia and anemia associated with inflammation. In human hepatoma cells, IL-6–induced hepcidin expression, an effect that was inhibited by treatment with a BMP type I receptor inhibitor, LDN-193189, or BMP ligand antagonists noggin and ALK3-Fc. In zebrafish, the induction of hepcidin expression by transgenic expression of IL-6 was also reduced by LDN-193189. In mice, treatment with IL-6 or turpentine increased hepcidin expression and reduced serum iron, effects that were inhibited by LDN-193189 or ALK3-Fc. Chronic turpentine treatment led to microcytic anemia, which was prevented by concurrent administration of LDN-193189 or attenuated when LDN-193189 was administered after anemia was established. Our studies support the concept that BMP and IL-6 act together to regulate iron homeostasis and suggest that inhibition of BMP signaling may be an effective strategy for the treatment of anemia of inflammation.

A Modified HIV1-Based Lentiviral Vector Can Transduce Rhesus Hematopoietic Repopulating Cells as Efficiently as An SIV Vector in An Autologous Transplantation Model.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 692-692
Author(s):  
Naoya Uchida ◽  
Phillip W Hargrove ◽  
Kareem Washington ◽  
Coen J. Lap ◽  
Matthew M. Hsieh ◽  
...  
Keyword(s):  
T Cells ◽  
Blood Cells ◽  
Conflicts Of Interest ◽  
Lentiviral Vector ◽  
Autologous Transplantation ◽  
Vector System ◽  
Transduction Efficiency ◽  
Large Animal ◽  
Hematopoietic Stem ◽  
Cd34 Cells

Abstract Abstract 692 HIV1-based vectors transduce rhesus hematopoietic stem cells poorly due to a species specific block by restriction factors, such as TRIM5αa which target HIV1 capsid proteins. The use of simian immunodeficiency virus (SIV)-based vectors can circumvent this restriction, yet use of this system precludes the ability to directly evaluate HIV1-based lentiviral vectors prior to their use in human clinical trials. To address this issue, we previously developed a chimeric HIV1 vector (χHIV vector) system wherein the HIV1-based lentiviral vector genome is packaged in the context of SIV capsid sequences. We found that this allowed χHIV vector particles to escape the intracellular defense mechanisms operative in rhesus hematopoietic cells as judged by the efficient transduction of both rhesus and human CD34+ cells. Following transplantation of rhesus animals with autologous cell transduced with the χHIV vector, high levels of marking were observed in peripheral blood cells (J Virol. 2009 Jul. in press). To evaluate whether χHIV vectors could transduce rhesus blood cells as efficiently as SIV vectors, we performed a competitive repopulation assay in two rhesus macaques for which half of the CD34+ cells were transduced with the standard SIV vector and the other half with the χHIV vector both at a MOI=50 and under identical transduction conditions. The transduction efficiency for rhesus CD34+ cells before transplantation with the χHIV vector showed lower transduction rates in vitro compared to those of the SIV vector (first rhesus: 41.9±0.83% vs. 71.2±0.46%, p<0.01, second rhesus: 65.0±0.51% vs. 77.0±0.18%, p<0.01, respectively). Following transplantation and reconstitution, however, the χHIV vector showed modestly higher gene marking levels in granulocytes (first rhesus: 12.4% vs. 6.1%, second rhesus: 36.1% vs. 27.2%) and equivalent marking levels in lymphocytes, red blood cells (RBC), and platelets, compared to the SIV vector at one month (Figure). Three to four months after transplantation in the first animal, in vivo marking levels plateaued, and the χHIV achieved 2-3 fold higher marking levels when compared to the SIV vector, in granulocytes (6.9% vs. 2.8%) and RBCs (3.3% vs. 0.9%), and equivalent marking levels in lymphocytes (7.1% vs. 5.1%) and platelets (2.8% vs. 2.5)(Figure). Using cell type specific surface marker analysis, the χHIV vector showed 2-7 fold higher marking levels in CD33+ cells (granulocytes: 5.4% vs. 2.7%), CD56+ cells (NK cells: 6.5% vs. 3.2%), CD71+ cells (reticulocyte: 4.5% vs. 0.6%), and RBC+ cells (3.6% vs. 0.9%), and equivalent marking levels in CD3+ cells (T cells: 4.4% vs. 3.3%), CD4+ cells (T cells: 3.9% vs. 4.6%), CD8+ cells (T cells: 4.2% vs. 3.9%), CD20+ cells (B cells: 7.6% vs. 4.8%), and CD41a+ cells (platelets: 3.5% vs. 2.2%) 4 months after transplantation. The second animal showed a similar pattern with higher overall levels (granulocytes: 32.8% vs. 19.1%, lymphocytes: 24.4% vs. 17.6%, RBCs 13.1% vs. 6.8%, and platelets: 14.8% vs. 16.9%) 2 months after transplantation. These data demonstrate that our χHIV vector can efficiently transduce rhesus long-term progenitors at levels comparable to SIV-based vectors. This χHIV vector system should allow preclinical testing of HIV1-based therapeutic vectors in the large animal model, especially for granulocytic or RBC diseases. Disclosures: No relevant conflicts of interest to declare.

CD34+CXCR4(CD184)+ Cells Differentiate Into Myeloid Dendritic Cell Progenitors

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4835-4835
Author(s):  
Robert E. Donahue ◽  
Ping Jin ◽  
Naoya Uchida ◽  
Aylin C. Bonifacino ◽  
Yu Tian ◽  
...  
Keyword(s):  
Innate Immunity ◽  
Confocal Microscopy ◽  
Vaccine Development ◽  
Inflammatory Responses ◽  
Conflicts Of Interest ◽  
Lentiviral Vector ◽  
Enzyme Linked Immunosorbent Assay ◽  
Cd34 Cells ◽  
Human Primate

Abstract Dendritic cells (DCs) play a central role in innate immunity and adaptive immunity. DCs are antigen presenting cells capable of inducing primary T cell responses or facilitating self-tolerance. Myeloid DCs (mDC) express CD11c. They are further divided into Type 1 myeloid DCs (mDC1) which are CD1c+CD141+ and Type 2 mDCs (mDC2) which are CD1c-CD141+, and plasmacytoid DC which are CD11c- and express CD303. Plasmacytoid DCs are the main source of type 1 interferon upon infection. CD34+ cells are a heterogeneous population of cells which contain both hematopoietic progenitors and stem cells. The microarray signature of CD34+CXCR4 (CD184)+ cells in both human and non-human primates suggest that this cell population plays a role in innate immunity. The signaling pathway for the Triggering Receptor Expressed on Myeloid Cells 1 (TREM1) is the most up-regulated pathway in both human and non-human primate sorted CD34+CD184+ cells. The TREM family is involved in the amplification and regulation of inflammatory responses. Upon sorting both human and rhesus CD34+ cells into CD34+CD184+ and CD34+CD184-, we observe that CD34+CD184+ cells are both adherent and non-adherent in nature (Figure 1) and while maintained in Flt3-L, IL-3, and SCF form progeny which appear dendritic in nature (Figure 2). In addition, we have found the CD34+CD184+ subpopulation to be resistant to lentiviral vector transduction. Upon culturing in defined, serum free media supplemented with cytokines, the progeny of the CD34+CD184+ population using both flow cytometry and confocal microscopy are CD11c+ and contain both CD1c+ and CD1c- subpopulations (Figure 3) with a predominately CD80(B7-1)+, CD123(IL-3R)+, CD197 (CCR7)+ , and HLA-Dr+ immunophenotype, having variability in CD86(B7-2) +/-, CD141(BDCA-3/Thrombomodulin)+/- , and CD144 (VE-cadherin)+/- expression, and being negative for CD303(BDCA-2)and CD309(VEGFR). Of special interest is that the CD34+CD184+ progeny contain both CD1c-CD16+ and CD1c+CD16- subpopulations of mDC. CD1c-CD16+ mDC have been shown to have strong pro-inflammatory activity, whereas CD1c+CD16- mDC are mainly inducers of chemotaxis. The progeny of the CD34+CD184+ cells can be stimulated by LPS and IFNγ to produce IL-12 based on an enzyme-linked immunosorbent assay. These results demonstrate the importance of the CD34+CXCR4+ progenitor in mDC development and allow one to speculate on how this mDC progenitor might prove of therapeutic benefit in vaccine development and cancer therapy.Figure 1Human CD34+CD184+ Sorted PopulationFigure 1. Human CD34+CD184+ Sorted PopulationFigure 2Cultured Progeny CD34+CD184+Figure 2. Cultured Progeny CD34+CD184+Figure 3Non-Human Primate CD34+CD184+ Confocal MicroscopyFigure 3. Non-Human Primate CD34+CD184+ Confocal Microscopy Disclosures: No relevant conflicts of interest to declare.

scholarly journals Hepcidin at the Nexus of Infection and Anemia

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. SCI-39-SCI-39
Author(s):  
Hal Drakesmith
Keyword(s):  
Animal Models ◽  
Blood Loss ◽  
Iron Homeostasis ◽  
Conflicts Of Interest ◽  
Indian Subcontinent ◽  
Peptide Hormone ◽  
The Body ◽  
Sub Saharan Africa ◽  
Human Populations ◽  
Hepcidin Expression

Abstract Humans have two main reasons for needing to control the amount and distribution of iron in the body. First, iron is used in fundamental physiological processes, including carrying oxygen, generating energy from oxygen, and in utilizing the energy for activities such as macromolecular synthesis and DNA repair. However, iron requirements are not stable over time; blood loss, growth, pregnancy, changes in diet and even altitude all have powerful and sometimes acute effects on our need for iron and how we route it to tissues that need it – for example, in the case of blood loss, iron is released for erythropoiesis to help replace lost red blood cells. The second reason to regulate iron is in response to infection; evidence for a critical influence of iron on the outcome of infections is broad. Sequencing of microbial genomes reveals significant investment into iron acquisition, allowing pathogenic organisms to obtain iron from multiple host sources. Iron is required for pathogen proliferation and increased availability of host iron, either through experimental administration in animal models or, in humans, due to genetic causes or because of nutritive or therapeutic iron supplementation, exacerbates infection or increases its incidence. Part of the innate immune response to infection is to deny iron to the pathogen, slowing its growth and giving more time for other arms of immunity to mobilize. The hypoferremia of infection helps to inhibit growth of microbes in the bloodstream, and may be a critical response to prevent potentially fatal septicemia. Hepcidin is the liver-encoded peptide hormone that allows the body to maintain iron homeostasis, to rapidly release iron for erythropoiesis, and to lock iron away from microorganisms in response to infection. The ability to integrate these activities lies in the unique sensitivity of hepcidin expression to diverse physiological inputs. Transcription of hepcidin is enhanced by signals deriving from iron accumulation (through BMP/SMAD signaling) and from immune mediators (IL-6, IL-22, IFN-a), and is suppressed by expanded erythropoiesis via erythroferrone. In animal models the relative strengths of these signals determine hepcidin synthesis, and thus determine iron absorption, release, and storage. Our lab has been focusing on understanding hepcidin regulation in human populations where infectious disease, anemia, iron deficiency and inherited red cell disorders are all prevalent, for example sub-Saharan Africa and the Indian subcontinent. This lecture will describe how hepcidin analysis has helped us to understand the complex etiology of childhood anemia in malarious regions, suggested methods to improve iron therapeutic strategies, and indicated ways to use hepcidin diagnostically. The latter applications are now beginning to be tested clinically at the University of Oxford. Disclosures No relevant conflicts of interest to declare.

Hepcidin Regulation by the BMP Pathway.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. SCI-25-SCI-25
Author(s):  
Jodie L. Babitt
Keyword(s):  
Signaling Pathway ◽  
Serum Iron ◽  
Iron Homeostasis ◽  
Iron Absorption ◽  
Bmp Signaling ◽  
Smad Pathway ◽  
Hepcidin Expression ◽  
Iron Administration ◽  
Smad Proteins

Abstract Abstract SCI-25 Systemic iron balance is regulated by the key iron regulatory hormone hepcidin. Secreted by the liver, hepcidin inhibits iron absorption from the diet and iron mobilization from body stores by decreasing cell surface expression of the iron export protein ferroportin. Iron administration increases hepcidin expression, thereby providing a feedback mechanism to limit further iron absorption, while anemia and hypoxia inhibit hepcidin expression, thereby increasing iron availability for erythropoiesis. Hepcidin excess is thought to have a role in the anemia of inflammation, while hepcidin deficiency is thought to be the common pathogenic mechanism underlying the iron overload disorder hereditary hemochromatosis, due to mutations in the genes encoding hepcidin itself (HAMP), HFE, transferrin receptor 2 (TFR2), or hemojuvelin (HFE2). Notably the precise molecular mechanisms by which iron levels are “sensed” and how this iron “signal” is transduced to modulate hepcidin expression have remained elusive. We have recently discovered that hemojuvelin is a co-receptor for the bone morphogenetic protein (BMP) signaling pathway, and that hemojuvelin-mediated BMP signals increase hepcidin expression at the transcriptional level. In addition to patients with HFE2 mutations and Hfe2 knockout mice, other genetic mouse models associated with impaired hepatic BMP signaling through a global knockout of the ligand Bmp6, or selective hepatic knockout of an intracellular mediator of BMP signaling, Smad4, also cause inappropriately low hepcidin expression and iron overload. Exogenous BMP6 administration in mice increases hepatic hepcidin expression and reduces serum iron, while BMP6 antagonists inhibit hepatic hepcidin expression, mobilize reticuloendothelial cell iron stores and increase serum iron. Not only does the BMP6-hemojuvelin-SMAD pathway regulate hepcidin expression and thereby systemic iron homeostasis, but also the BMP6-SMAD pathway itself is regulated by iron. Acute iron administration in mice increases phosphorylation of Smad proteins in the liver, and chronic changes in dietary iron modulate hepatic Bmp6 mRNA expression and phosphorylation of Smad proteins concordantly with Hamp mRNA expression. Together, these data support the paramount role of the BMP6-hemojuvelin-SMAD signaling pathway in the iron-mediated regulation of hepcidin expression and systemic iron homeostasis, and suggest that modulators of this pathway may be an alternative therapeutic strategy for treating disorders of iron homeostasis. Recent work elucidating the role of the BMP signaling pathway in hepcidin regulation and systemic iron homeostasis will be presented. Disclosures Babitt: Ferrumax Pharmaceuticals, Inc.: Equity Ownership.

scholarly journals Molecular mechanisms of normal iron homeostasis

Hematology ◽  
2009 ◽  
Vol 2009 (1) ◽  
pp. 207-214 ◽  
Author(s):  
An-Sheng Zhang ◽  
Caroline A. Enns
Keyword(s):  
Iron Homeostasis ◽  
Molecular Mechanisms ◽  
Iron Absorption ◽  
Hypoxia Inducible Factor ◽  
Bmp Signaling ◽  
Control Mechanisms ◽  
Iron Regulatory Proteins ◽  
Hepcidin Expression ◽  
Cellular Iron ◽  
Morphogenetic Protein

Abstract Humans possess elegant control mechanisms to maintain iron homeostasis by coordinately regulating iron absorption, iron recycling, and mobilization of stored iron. Dietary iron absorption is regulated locally by hypoxia inducible factor (HIF) signaling and iron-regulatory proteins (IRPs) in enterocytes and systematically by hepatic hepcidin, the central iron regulatory hormone. Hepcidin not only controls the rate of iron absorption but also determines iron mobilization from stores through negatively modulating the function of ferroportin, the only identified cellular iron exporter to date. The regulation of hepatic hepcidin is accomplished by the coordinated activity of multiple proteins through different signaling pathways. Recent studies have greatly expanded the knowledge in the understanding of hepcidin expression and regulation by the bone morphogenetic protein (BMP) signaling, the erythroid factors, and inflammation. In this review, we mainly focus on the roles of recently identified proteins in the regulation of iron homeostasis.

scholarly journals Hepatocyte Neogenin Is Required for Hemojuvelin-Mediated Hepcidin Expression and Iron Homeostasis in Mice

Blood ◽  
2021 ◽  
Author(s):  
Caroline A. Enns ◽  
Shall Jue ◽  
An-Sheng Zhang
Keyword(s):  
Plasma Membrane ◽  
Iron Overload ◽  
Serum Iron ◽  
Iron Homeostasis ◽  
Transmembrane Protein ◽  
High Serum ◽  
Bmp Signaling ◽  
Hepcidin Expression ◽  
Iron Utilization

Neogenin (NEO1) is a ubiquitously expressed multi-functional transmembrane protein. It interacts with hemojuvelin (HJV), a BMP co-receptor that plays a pivotal role in hepatic hepcidin expression. Earlier studies suggest that the function of HJV relies on its interaction with NEO1. However, the role of NEO1 in iron homeostasis remains controversial because of the lack of an appropriate animal model. Here, we generated a hepatocyte-specific Neo1 knockout (Neo1fl/fl;Alb-Cre+) mouse model that circumvented the developmental and lethality issues of the global Neo1 mutant. Results show that ablation of hepatocyte Neo1 decreased hepcidin expression and caused iron overload. This iron overload did not result from altered iron utilization by erythropoiesis. Replacement studies revealed that expression of the Neo1L1046E mutant that does not interact with Hjv, was unable to correct the decreased hepcidin expression and high serum iron in Neo1fl/fl;Alb-Cre+ mice. In Hjv-/- mice, expression of HjvA183R mutant that has reduced interaction with Neo1, also displayed a blunted induction of hepcidin expression. These observations indicate that Neo1-Hjv interaction is essential for hepcidin expression. Further analyses suggest that the Hjv binding triggered the cleavage of the Neo1 cytoplasmic domain by a protease, which resulted in accumulation of truncated Neo1 on the plasma membrane. Additional studies did not support that Neo1 functions by inhibiting Hjv shedding as previously proposed. Together, our data favor a model in which Neo1 interaction with Hjv leads to accumulation of cleaved Neo1 on the plasma membrane, where Neo1 acts as a scaffold to induce the Bmp signaling and hepcidin expression.

Hepcidin Expression Is Regulated by a Complex of Hemochromatosis-Associated Proteins.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 267-267 ◽  
Author(s):  
Paul J. Schmidt ◽  
Franklin W. Huang ◽  
Diedra M. Wrighting ◽  
Paul T. Toran ◽  
Nancy C. Andrews
Keyword(s):  
Transferrin Receptor ◽  
Iron Homeostasis ◽  
Hepatoma Cell ◽  
Bmp Signaling ◽  
Hepatoma Cell Line ◽  
Type I ◽  
Mutant Form ◽  
Iron Release ◽  
Hepcidin Expression ◽  
Bmp Receptors

Abstract Hemochromatosis is a common genetic disease resulting from increased dietary iron absorption and tissue iron deposition. Mutations in five unrelated genes are known to cause hemochromatosis in humans and mice. These encode the classic hemochromatosis protein (HFE), transferrin receptor 2 (TFR2), the iron exporter ferroportin (FPN), hemojuvelin (HJV), and the circulating anti-microbial peptide hepcidin (HAMP). Hepcidin binds to FPN, causing its internalization and degradation, thus decreasing cellular iron release. A basic understanding of the pathophysiology of FPN and hepcidin mutations has recently been elucidated; however, it was still unclear how mutations in HFE, TFR2, and HJV cause hemochromatosis. All are associated with decreased hepcidin production and inappropriately high levels of ferroportin activity. HFE, TFR2 and HJV are normally expressed in the hepatic cells that produce hepcidin. With collaborators, we showed that HJV acts as a bone morphogenetic protein (BMP) co-receptor. HJV binds to the BMP ligands and forms a complex with Type I BMP receptors, resulting in signaling through a SMAD pathway and induction of hepcidin expression. Disease causing mutations in HJV abrogate BMP co-receptor activity, and hepatocytes from Hjv−/ − mice have a blunted response to BMP2. HFE was known to form a complex with the classical transferrin receptor, TFR1. Several models have been proposed implicating this complex in the regulation of normal iron homeostasis, but they have not taken the role of hepcidin into account. To examine the HFE/TFR1 interaction in vivo, we developed mice expressing a mutant form of TFR1 that should constitutively interact with HFE. We found that these transgenic animals have a phenotype similar to Hfe−/ − mice, suggesting that TFR1 serves to sequester HFE to silence its activity. We next asked whether HFE might also participate in BMP signaling. We found that forced expression of HFE in a hepatoma cell line induces transcription of a reporter gene linked to the hepcidin promoter. It also induces transcription from a heterologous promoter containing BMP-responsive elements, suggesting that HFE works through the BMP pathway. In contrast, forced expression of TFR2 did not amplify expression of either reporter, but it prevented cellular release of a soluble cleavage product of HJV. Furthermore, we showed that both HFE and TFR2 are associated with HJV in a stable protein complex that can be isolated by co-immunoprecipitation or Ni-affinity chromatography. TFR2 appears to aid in the recruitment of HFE to this complex. We conclude that HFE and TFR2 thus serve to amplify BMP signaling through an HJV/BMP receptor pathway. Our findings provide a compelling explanation for the similar clinical hemochromatosis phenotypes resulting from mutations in these genes.

Processing of Hemojuvelin Requires Retrograde Trafficking to the Golgi in HepG2 Cells.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3845-3845
Author(s):  
An-Sheng Zhang ◽  
Julia Julia Maxson ◽  
Caroline A Enns
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Hepg2 Cells ◽  
Iron Homeostasis ◽  
Bmp Signaling ◽  
Soluble Form ◽  
Gpi Anchor ◽  
Retrograde Trafficking ◽  
Hepcidin Expression ◽  
Bone Morphogenic Proteins

Abstract Hemojuvelin (HJV) was recently identified as a critical regulator of iron homeostasis. It is either associated with the cells through a GPI-anchor or released as a soluble form. The cellular form acts as a co-receptor for bone morphogenic proteins (BMPs) and activates the transcription of hepcidin, a hormone that regulates iron efflux from cells. Soluble HJV antagonizes BMP signaling and suppresses hepcidin expression. Secretion of HJV requires binding to the transmembrane receptor neogenin. In this study we examined the trafficking and processing of HJV. Cellular HJV reached the plasma membrane without obtaining complex oligosaccharides, indicating that HJV avoided Golgi processing. Secreted HJV, in contrast, had complex oligosaccharides and could be derived from the pool of HJV at the plasma membrane. Neogenin did not play a role in HJV trafficking to the cell surface but was necessary for secretion of HJV, suggesting that it could be involved in either retrograde trafficking of HJV or in cleavage leading to secretion.

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