Congenital sideroblastic anemia model due to ALAS2 mutation is susceptible to ferroptosis.

X-linked sideroblastic anemia (XLSA), the most common form of congenital sideroblastic anemia, is caused by a germline mutation in the erythroid-specific 5-aminolevulinate synthase (ALAS2) gene. In XLSA, defective heme biosynthesis leads to ring sideroblast formation because of excess mitochondrial iron accumulation. In this study, we introduced ALAS2 missense mutations on human umbilical cord blood-derived erythroblasts; hereafter, we refer to them as XLSA clones. XLSA clones that differentiated into mature erythroblasts showed an increased frequency of ring sideroblast formation with impaired hemoglobin biosynthesis. The expression profiling revealed significant enrichment of genes involved in ferroptosis, which is a form of regulated cell death induced by iron accumulation and lipid peroxidation. Notably, treatment with erastin, a ferroptosis inducer, caused a higher proportion of cell death in XLSA clones. XLSA clones exhibited significantly higher levels of intracellular lipid peroxides and enhanced expression of BACH1, a regulator of iron metabolism and potential accelerator of ferroptosis. In XLSA clones, BACH1 repressed genes involved in iron metabolism and glutathione synthesis. Collectively, defective heme biosynthesis in XLSA clones could confer enhanced BACH1 expression, leading to increased susceptibility to ferroptosis. The results of our study provide important information for the development of novel therapeutic targets for XLSA.

Cryo-EM structure of AMP-PNP-bound human mitochondrial ATP-binding cassette transporter ABCB7.

ATP-binding cassette subfamily B member 7 (ABCB7) is localized in the inner membrane of mitochondria, playing a critical role in iron metabolism. Here, we determined the structure of the nonhydrolyzable ATP analog adenosine-5'-(ß-γ-imido) triphosphate (AMP-PNP) bound human ABCB7 at 3.3 Å by single-particle electron cryo-microscopy (cryo-EM). The AMP-PNP-bound human ABCB7 shows an inverted V-shaped homodimeric architecture with an inward-facing open conformation. One AMP-PNP molecule and Mg2+ were identified in each nucleotide-binding domain (NBD) of the hABCB7 monomer. Moreover, four disease-causing missense mutations of human ABCB7 have been mapped to the structure, creating a hotspot map for X-linked sideroblastic anemia and ataxia disease. Our results provide a structural basis for further understanding the transport mechanism of the mitochondrial ABC transporter.

Mitochondrial Ferritin: Its Role in Physiological and Pathological Conditions.

In 2001, a new type of human ferritin was identified by searching for homologous sequences to H-ferritin in the human genome. After the demonstration that this ferritin is located specifically in the mitochondrion, it was called mitochondrial ferritin. Studies on the properties of this new type of ferritin have been limited by its very high homology with the cytosolic H-ferritin, which is expressed at higher levels in cells. This great similarity made it difficult to obtain specific antibodies against the mitochondrial ferritin devoid of cross-reactivity with cytosolic ferritin. Thus, the knowledge of the physiological role of mitochondrial ferritin is still incomplete despite 20 years of research. In this review, we summarize the literature on mitochondrial ferritin expression regulation and its physical and biochemical properties, with particular attention paid to the differences with cytosolic ferritin and its role in physiological condition. Until now, there has been no evidence that the alteration of the mitochondrial ferritin gene is causative of any disorder; however, the identified association of the mitochondrial ferritin with some disorders is discussed.

Identification of circular RNAs as novel biomarkers and potentially functional competing endogenous RNA network for myelodysplastic syndrome patients.

Circular RNAs (circRNAs) have been identified to exert vital biological functions and can be used as new biomarkers in a number of tumors. However, little is known about the functions of circRNAs in myelodysplastic syndrome (MDS). Here, we aimed to investigate circRNA expression profiles and to investigate the functional and clinical value of circRNAs in MDS. Differential expression of circRNAs between MDS and control subjects was analyzed using circRNA arrays, in which we identified 145 upregulated circRNAs and 224 downregulated circRNAs. Validated by real-time quantitative PCR between 100 MDS patients and 20 controls, three upregulated (hsa_circRNA_100352, hsa_circRNA_104056, and hsa_circRNA_104634) and three downregulated (hsa_circRNA_103846, hsa_circRNA_102817, and hsa_circRNA_102526) circRNAs matched the arrays. The receiver operating characteristic curve analysis of these circRNAs showed that the area under the curve was 0.7266, 0.8676, 0.7349, 0.7091, 0.8806, and 0.7472, respectively. Kaplan-Meier survival analysis showed that only hsa_circRNA_100352, hsa_circRNA_104056, and hsa_circRNA_102817 were significantly associated with overall survival. Furthermore, we generated a competing endogenous RNA network focused on hsa_circRNA_100352, hsa_circRNA_104056, and hsa_circRNA_102817. Analyses using Gene Ontology and Kyoto Encyclopedia of Genes and Genomes showed that the three circRNAs were linked with some important cancer-related functions and pathways.

Mutations in the iron-sulfur cluster biogenesis protein HSCB cause congenital sideroblastic anemia.

The congenital sideroblastic anemias (CSAs) can be caused by primary defects in mitochondrial iron-sulfur (Fe-S) cluster biogenesis. HSCB (heat shock cognate B), which encodes a mitochondrial cochaperone, also known as HSC20 (heat shock cognate protein 20), is the partner of mitochondrial heat shock protein A9 (HSPA9). Together with glutaredoxin 5 (GLRX5), HSCB and HSPA9 facilitate the transfer of nascent 2-iron, 2-sulfur clusters to recipient mitochondrial proteins. Mutations in both HSPA9 and GLRX5 have previously been associated with CSA. Therefore, we hypothesized that mutations in HSCB could also cause CSA. We screened patients with genetically undefined CSA and identified a frameshift mutation and a rare promoter variant in HSCB in a female patient with non-syndromic CSA. We found that HSCB expression was decreased in patient-derived fibroblasts and K562 erythroleukemia cells engineered to have the patient-specific promoter variant. Furthermore, gene knockdown and deletion experiments performed in K562 cells, zebrafish, and mice demonstrate that loss of HSCB results in impaired Fe-S cluster biogenesis, a defect in RBC hemoglobinization, and the development of siderocytes and more broadly perturbs hematopoiesis in vivo. These results further affirm the involvement of Fe-S cluster biogenesis in erythropoiesis and hematopoiesis and define HSCB as a CSA gene.

MDS/MPN-RS-T justified inclusion as a unique disease entity?

Myelodysplastic syndrome/myeloproliferative neoplasm with ring sideroblasts and thrombocytosis (MDS/MPN-RS-T) is a disease entity characterized by anemia, bone marrow dysplasia with ring sideroblasts and persistent thrombocytosis ≥450 × 109/L with proliferation of large and morphologically atypical megakaryocytes. Although initially recognized by the World Health Organization only as a provisional entity, next generation sequencing has identified recurrent somatic mutations in SF3B1, JAK2 and other genes providing further evidence of the clonal nature of this disease and the need to recognize it as a separate entity. Despite its overlapping features with MDS with ring sideroblasts and essential thrombocythemia, MDS/MPN-RS-T is characterized by specific clinical features and distinct survival outcomes. In the current review we will describe the morphological and genomic features of MDS-RS-T and the potential diagnostic challenges and distinction from other possible conditions. We will also review how the current evidence supports its recognition as an independent disorder.

Hereditary Ataxia: A Focus on Heme Metabolism and Fe-S Cluster Biogenesis.

Heme and Fe-S clusters regulate a plethora of essential biological processes ranging from cellular respiration and cell metabolism to the maintenance of genome integrity. Mutations in genes involved in heme metabolism and Fe-S cluster biogenesis cause different forms of ataxia, like posterior column ataxia and retinitis pigmentosa (PCARP), Friedreich's ataxia (FRDA) and X-linked sideroblastic anemia with ataxia (XLSA/A). Despite great efforts in the elucidation of the molecular pathogenesis of these disorders several important questions still remain to be addressed. Starting with an overview of the biology of heme metabolism and Fe-S cluster biogenesis, the review discusses recent progress in the understanding of the molecular pathogenesis of PCARP, FRDA and XLSA/A, and highlights future line of research in the field. A better comprehension of the mechanisms leading to the degeneration of neural circuity responsible for balance and coordinated movement will be crucial for the therapeutic management of these patients.

Myelodysplastic Syndrome/Myeloproliferative Neoplasm with Ring Sideroblasts and Thrombocytosis with Cooccurrent SF3B1 and MPL Gene Mutations: A Case Report and Brief Review of the Literature.

BACKGROUND: Myelodysplastic syndrome/myeloproliferative neoplasm with ring sideroblasts and thrombocytosis (MDS/MPN-RS-T) is a new disease entity in the current WHO classification. Genetically, 60%-90% of cases have mutations in SF3B1, strongly associated with RS, and more than half of them cooccur with JAK2 V617F. This report describes the rare case of MDS/MPN-RS-T with SF3B1 mutation cooccurring with an MPL mutation. METHODS: We report a 79-year-old man who was referred because of generalized edema. Peripheral blood testing showed macrocytic anemia and thrombocytosis, and bone marrow analysis demonstrated dyserythropoiesis with RS and increased megakaryocytes. A molecular study was performed to detect SF3B1 mutations and recurrent mutations in MPN disease (JAK2 V617F/exon 12, CALR gene exon 9, and MPL gene exon 10 mutations). RESULTS: The molecular study revealed SF3B1 K666T and MPL W515R mutations, while BCR-ABL1 or JAK2 V617F/exon 12 and CALR mutations were all negative. CONCLUSION: This is a rare case of concomitant SF3B1 and MPL mutations in MDS/MPN-RS-T.

Generation and Molecular Characterization of Human Ring Sideroblasts: a Key Role of Ferrous Iron in Terminal Erythroid Differentiation and Ring Sideroblast Formation.

Ring sideroblasts are a hallmark of sideroblastic anemia, although little is known about their characteristics. Here, we first generated mutant mice by disrupting the GATA-1 binding motif at the intron 1 enhancer of the ALAS2 gene, a gene responsible for X-linked sideroblastic anemia (XLSA). Although heterozygous female mice showed an anemic phenotype, ring sideroblasts were not observed in their bone marrow. We next established human induced pluripotent stem cell-derived proerythroblast clones harboring the same ALAS2 gene mutation. Through coculture with sodium ferrous citrate, mutant clones differentiated into mature erythroblasts and became ring sideroblasts with upregulation of metal transporters (MFRN1, ZIP8, and DMT1), suggesting a key role for ferrous iron in erythroid differentiation. Interestingly, holo-transferrin (holo-Tf) did not induce erythroid differentiation as well as ring sideroblast formation, and mutant cells underwent apoptosis. Despite massive iron granule content, ring sideroblasts were less apoptotic than holo-Tf-treated undifferentiated cells. Microarray analysis revealed upregulation of antiapoptotic genes in ring sideroblasts, a profile partly shared with erythroblasts from a patient with XLSA. These results suggest that ring sideroblasts exert a reaction to avoid cell death by activating antiapoptotic programs. Our model may become an important tool to clarify the pathophysiology of sideroblastic anemia.

Molecular pathophysiology and genetic mutations in congenital sideroblastic anemia.

Sideroblastic anemia is a heterogeneous congenital and acquired disorder characterized by anemia and the presence of ring sideroblasts in the bone marrow. Congenital sideroblastic anemia (CSA) is a rare disease caused by mutations in genes involved in the heme biosynthesis, iron-sulfur [Fe-S] cluster biosynthesis, and mitochondrial protein synthesis. The most prevalent form of CSA is X-linked sideroblastic anemia, caused by mutations in the erythroid-specific δ-aminolevulinate synthase (ALAS2), which is the first enzyme of the heme biosynthesis pathway in erythroid cells. To date, a remarkable number of genetically undefined CSA cases remain, but a recent application of the next-generation sequencing technology has recognized novel causative genes for CSA. However, in most instances, the detailed molecular mechanisms of how defects of each gene result in the abnormal mitochondrial iron accumulation remain unclear. This review aims to cover the current understanding of the molecular pathophysiology of CSA.

The molecular genetics of sideroblastic anemia.

The sideroblastic anemias (SAs) are a group of inherited and acquired bone marrow disorders defined by pathological iron accumulation in the mitochondria of erythroid precursors. Like most hematological diseases, the molecular genetic basis of the SAs has ridden the wave of technology advancement. Within the last 30 years, with the advent of positional cloning, the human genome project, solid-state genotyping technologies, and next-generation sequencing have evolved to the point where more than two-thirds of congenital SA cases, and an even greater proportion of cases of acquired clonal disease, can be attributed to mutations in a specific gene or genes. This review focuses on an analysis of the genetics of these diseases and how understanding these defects may contribute to the design and implementation of rational therapies.

Sideroblastic anemia associated with multisystem mitochondrial disorders.

BACKGROUND: Sideroblastic anemia represents a heterogeneous group of inherited or acquired diseases with disrupted erythroblast iron utilization, ineffective erythropoiesis, and variable systemic iron overload. In a cohort of 421 patients with multisystem mitochondrial diseases, refractory anemia was found in 8 children. RESULTS: Five children had sideroblastic anemia with increased numbers of ring sideroblasts >15%. Two of the children had a fatal course of MLASA1 syndrome (mitochondrial myopathy, lactic acidosis, and sideroblastic anemia [SA]) due to a homozygous, 6-kb deletion in the PUS1 gene, part of the six-member family of pseudouridine synthases (pseudouridylases). Large homozygous deletions represent a novel cause of presumed PUS1-loss-of-function phenotype. The other three children with SA had Pearson syndrome (PS) due to mtDNA deletions of 4 to 8 kb; two of these children showed early onset of PS and died due to repeated sepsis; the other child had later onset of PS and survived as the hematological parameters normalized and the disease transitioned to Kearns-Sayre syndrome. In addition, anemia without ring sideroblasts was found in three other patients with mitochondrial disorders, including two children with later onset of PS and one child with failure to thrive, microcephaly, developmental delay, hypertrophic cardiomyopathy, and renal tubular acidosis due to the heterozygous mutations c.610A>G (p.Asn204Asp) and c.674C>T (p.Pro225Leu) in the COX10 gene encoding the cytochrome c oxidase assembly factor. CONCLUSIONS: Sideroblastic anemia was found in fewer than 1.2% of patients with multisystem mitochondrial disease, and it was usually associated with an unfavorable prognosis.

Congenital sideroblastic anemia: Advances in gene mutations and pathophysiology.

Congenital sideroblastic anemia (CSA) is a series of rare, heterogeneous disorders, characterized by iron overload in the mitochondria of erythroblasts and ringed sideroblasts in bone marrow. In recent years, rapid development of next-generation sequencing technology brings great advance in understanding of genetic and pathophysiologic features of CSA. Based on the pathophysiology of mitochondrial iron metabolism, causative genes of CSA can be divided into three subtypes: heme biosynthesis related; iron­sulfur cluster biosynthesis and transportation related; and mitochondrial respiratory chain synthesis related. Patients with CSA present various clinical manifestation due to relevant mutation gene and require different treatment strategies. The recognition of the causative genes and evolution of pathogenicity is critical. In this review, we summarize the recent progress in mutation genes of CSA, and its potential role in the pathogenesis, diagnosis and treatment.

Pearson syndrome.

INTRODUCTION: Pearson syndrome (PS) is a sporadic and very rare syndrome classically associated with single large-scale deletions of mitochondrial DNA and characterized by refractory sideroblastic anemia during infancy. Areas covered: This review presents an analysis and interpretation of the published data that forms the basis for our understanding of PS. PubMed, Google Scholarand Thompson ISI Web of Knowledge were searched for relevant data. Expert commentary: PS is a very rare mitochodrial disease that involves different organs and systems. Clinical phenotype is extremely variable and may change over the course of disease itself with the possibility both of worsenings and improvements. Outcome is invariably lethal and at the moment no cure is available. Accurate supportive treatment and follow up program in centres with experience in mitochondrial diseases and marrow failure may positively influence quality and duration of life.

Iron metabolism in erythroid cells and patients with congenital sideroblastic anemia.

Sideroblastic anemias are anemic disorders characterized by the presence of ring sideroblasts in a patient's bone marrow. These disorders are typically divided into two types, congenital or acquired sideroblastic anemia. Recently, several genes were reported as responsible for congenital sideroblastic anemia; however, the relationship between the function of the gene products and ring sideroblasts is largely unclear. In this review article, we will focus on the iron metabolism in erythroid cells as well as in patients with congenital sideroblastic anemia.

Splicing factor mutations in MDS RARS and MDS/MPN-RS-T.

Spliceosomal mutations, especially mutations in SF3B1, are frequently (>80%) identified in patients with refractory anemia with ringed sideroblasts (RARS) and myelodysplastic/myeloproliferative neoplasms with ringed sideroblasts and thrombocytosis (MDS/MPN-RS-T; previously known as RARS-T), and SF3B1 mutations have a high positive predictive value for disease phenotype with ringed sideroblasts. These observations suggest that SF3B1 mutations play important roles in the pathogenesis of these disorders and formation of ringed sideroblasts. Here we will review recent insights into the molecular mechanisms of mis-splicing caused by mutant SF3B1 and the pathogenesis of RSs in the context of congenital sideroblastic anemia as well as RARS with SF3B1 mutations. We will also discuss therapy of SF3B1 mutant MDS, including novel approaches.