p53 activation during ribosome biogenesis regulates normal erythroid differentiation.

The role of ribosome biogenesis in erythroid development is supported by the recognition of erythroid defects in ribosomopathies in both Diamond-Blackfan anemia and 5q- syndrome. Whether ribosome biogenesis exerts a regulatory function on normal erythroid development is still unknown. In the present study, a detailed characterization of ribosome biogenesis dynamics during human and murine erythropoiesis showed that ribosome biogenesis is abruptly interrupted by the decline in ribosomal DNA transcription and the collapse of ribosomal protein neosynthesis. Its premature arrest by the RNA Pol I inhibitor CX-5461 targeted the proliferation of immature erythroblasts. p53 was activated spontaneously or in response to CX-5461, concomitant to ribosome biogenesis arrest, and drove a transcriptional program in which genes involved in cell cycle-arrested, negative regulation of apoptosis, and DNA damage response were upregulated. RNA Pol I transcriptional stress resulted in nucleolar disruption and activation of the ATR-CHK1-p53 pathway. Our results imply that the timing of ribosome biogenesis extinction and p53 activation is crucial for erythroid development. In ribosomopathies in which ribosome availability is altered by unbalanced production of ribosomal proteins, the threshold downregulation of ribosome biogenesis could be prematurely reached and, together with pathological p53 activation, prevents a normal expansion of erythroid progenitors.

Translation defects in ribosomopathies.

PURPOSE OF REVIEW: Congenital or acquired ribosomopathies related to mutations or deletions in ribosomal proteins gene or ribosome-associated proteins exhibit defective ribosome biogenesis that expose the cell to translation defects. The mechanisms leading to low translation rate, loss-of-translation fidelity and translation selectivity are reviewed. RECENT FINDINGS: New quantitative techniques to measure ribosome component stoichiometry reveal that the pool of ribosomes could be heterogeneous and/or decreased with a limited number of translationally competent ribosomes. During development or cell differentiation, the absence of specific ribosome components or their replacement by paralogs generate heterogeneous ribosomes that are specialized in the translation of specific mRNAs. Decreased ribosome content by defective biosynthesis of a subunit results in translation selectivity at the expense of short structured transcripts with high codon adaptation index. Activation of p53, as a witness of nucleolar stress associated with the hematological phenotype of ribosomopathies participates in translational reprogramming of the cell by interfering with cap-dependent translation. SUMMARY: Translation selectivity is a common feature of ribosomopathies. p53 is more selectively activated in ribosomopathies with erythroid phenotype. The discovery of its dual role in regulating transcriptional and translational program supports new therapeutic perspectives.

When refeeding is not enough: severe and prolonged pancytopenia in an adolescent with anorexia nervosa.

A 15-year-old female patient with anorexia nervosa presented an unusually prolonged and severe episode of pancytopenia with severe thrombopenia and severe leucopenia. Despite effective refeeding, active specialized interventions were necessary. Upon admission, the patient presented with severe and symptomatic thrombopenia, severe neutropenia and gelatinous marrow transformation. In addition to refeeding, active interventions such as platelet transfusion and granulocyte-colony stimulating factor were successful to manage the patient's complications. The etiological search for pancytopenia was negative. The patient's prolonged starvation was probably a key factor. Medical history, clinical presentation, evolution, and biological data including bone marrow aspiration results are presented. Management of cytopenia and of their complications in a context of severe starvation is discussed in regard of existing literature. A simple monitoring attitude may prove insufficient in cases of severe pancytopenia in anorexia nervosa.Level of evidence V, descriptive study.

Integrated analyses of translatome and proteome identify the rules of translation selectivity in RPS14-deficient cells.

In ribosomopathies, the Diamond-Blackfan anemia (DBA) or 5q- syndrome, ribosomal protein (RP) genes are affected by mutation or deletion, resulting in bone marrow erythroid hypoplasia. Unbalanced production of ribosomal subunits leading to a limited ribosome cellular content regulates translation at the expense of the master erythroid transcription factor GATA1. In RPS14-deficient cells mimicking 5q- syndrome erythroid defects, we show that the transcript length, codon bias of the coding sequence (CDS) and 3'UTR (untranslated region) structure are the key determinants of translation. In these cells, short transcripts with a structured 3'UTR and high codon adaptation index (CAI) showed a decreased translation efficiency. Quantitative analysis of the whole proteome confirmed that the post-transcriptional changes depended on the transcript characteristics that governed the translation efficiency in conditions of low ribosome availability. In addition, proteins involved in normal erythroid differentiation share most determinants of translation selectivity. Our findings thus indicate that impaired erythroid maturation due to 5q- syndrome may proceed from a translational selectivity at the expense of the erythroid differentiation program, and suggest that an interplay between the CDS and UTR may regulate mRNA translation.

Maternal and obstetric outcomes in women with pregnancy-associated haematological malignancies: an observational nationwide cohort study.

BACKGROUND: Pregnancy-associated haematological malignancy is a rare event; therefore, data available to guide the treatment are scarce. We aimed to evaluate the incidence, overall survival, and maternal morbidity and mortality of women with pregnancy-associated haematological malignancies. METHODS: We conducted a nationwide observational cohort study using the French National Healthcare Data System (SNDS), a health-care administrative database covering up to 99% of the French population. We included all pregnancies in France ending between Jan 1, 2012, and Dec 31, 2022. Pregnancies with terminations or miscarriages managed on an outpatient basis, and women with a history of haematological malignancies before pregnancy were excluded. A Cox proportional hazards model was used to assess overall survival, defined as the date of haematological malignancy diagnosis to either death or the end of the study follow-up, in the haematological malignancy during pregnancy group (pregnancies with a diagnosis of haematological malignancy during pregnancy) compared with the haematological malignancy post-pregnancy group (pregnancies with a diagnosis of haematological malignancy in the year following pregnancy). Severe maternal morbidity was compared in the haematological malignancy during pregnancy group versus the reference group (pregnancies without a history of haematological malignancy or a diagnosis of pregnancy-associated haematological malignancy). Births were classified as very preterm (<32 weeks of pregnancy), preterm (32-36 weeks), and term (≥37 weeks) and compared in the haematological malignancy during pregnancy group versus the reference group. Inverse probability weighting (IPW) was used for confounder adjustment, using maternal age (categorised), comorbidities, socioeconomic status, and year of delivery (as a category). FINDINGS: Of 9 996 523 pregnancies in 5 995 235 women, 1366 pregnancy-associated haematological malignancies were identified: 413 during pregnancy (4·13 per 100 000 pregnancies) and 953 (9·53 per 100 000 pregnancies) within 12 months of the end of pregnancy (post-pregnancy). No significant differences in overall survival were observed between the haematological malignancy during and post-pregnancy groups across all types of haematological malignancy (IPW-adjusted hazard ratio 0·91 [95% CI 0·62-1·34], p=0·63), specifically for Hodgkin lymphoma (0·56 [0·07-4·53], p=0·59), aggressive B-cell non-Hodgkin lymphoma (0·52 [0·12-2·38], p=0·40), and acute leukaemia alone (0·84 [0·50-1·41], p=0·51). Severe maternal morbidity was more frequent in the haematological malignancy during pregnancy group than in the reference group (86 [26·2%] of 328 completed pregnancies vs 120 335 [1·5%] of 7 945 909 completed pregnancies; IPW-adjusted odds ratio 22·71 [95% CI 17·72-29·10], p<0·0001). We observed an increase in very preterm birth (32 [9·8%] vs 92 712 [1·2%]; IPW-adjusted odds ratio 11·90 [95% CI 7·91-17·91], p<0·0001) and preterm birth (116 [35·4%] vs 430 472 [5·4%]; 11·76 [9·34-14·81], p<0·0001) in the haematological malignancy during pregnancy group compared with the reference group. INTERPRETATION: This nationwide observational study examines pregnancy-associated haematological malignancies in France, revealing no significant difference in overall survival between women diagnosed during pregnancy and post-pregnancy. Our data highlight an increased frequency of severe maternal morbidity and obstetric complications among women diagnosed during pregnancy. Notably, the study underscores the necessity for specialised care to manage these complex cases effectively. FUNDING: None. TRANSLATION: For the French translation of the abstract see Supplementary Materials section.

Loss of hematopoietic progenitors heterogeneity is an adverse prognostic factor in lower-risk myelodysplastic neoplasms.

Myelodysplastic neoplasms (MDS) are characterized by clonal evolution starting from the compartment of hematopoietic stem and progenitors cells (HSPCs), leading in some cases to leukemic transformation. We hypothesized that deciphering the diversity of the HSPCs compartment may allow for the early detection of an emergent sub-clone that drives disease progression. Deep analysis of HSPCs repartition by multiparametric flow cytometry revealed a strong disorder of the hematopoietic branching system in most patients at diagnosis with different phenotypic signatures closely related to specific MDS features. In two independent cohorts of 131 and 584 MDS, the HSPCs heterogeneity quantified through entropy calculation was decreased in 47% and 46% of cases, reflecting a more advanced state of the disease with deeper cytopenias, higher IPSS-R risk and accumulation of somatic mutations. We demonstrated that patients with lower-risk MDS and low CD34 + CD38+HSPCs entropy had an adverse outcome and that this parameter is as an independent predictive biomarker for progression free survival, leukemia free survival and overall survival. Analysis of HSPCs repartition at diagnosis represents therefore a very powerful tool to identify lower-risk MDS patients with a worse outcome and valuable for clinical decision-making, which could be fully integrated in the MDS diagnostic workflow.

Accelerated DNA replication fork speed due to loss of R-loops in myelodysplastic syndromes with SF3B1 mutation.

Myelodysplastic syndromes (MDS) with mutated SF3B1 gene present features including a favourable outcome distinct from MDS with mutations in other splicing factor genes SRSF2 or U2AF1. Molecular bases of these divergences are poorly understood. Here we find that SF3B1-mutated MDS show reduced R-loop formation predominating in gene bodies associated with intron retention reduction, not found in U2AF1- or SRSF2-mutated MDS. Compared to erythroblasts from SRSF2- or U2AF1-mutated patients, SF3B1-mutated erythroblasts exhibit augmented DNA synthesis, accelerated replication forks, and single-stranded DNA exposure upon differentiation. Importantly, histone deacetylase inhibition using vorinostat restores R-loop formation, slows down DNA replication forks and improves SF3B1-mutated erythroblast differentiation. In conclusion, loss of R-loops with associated DNA replication stress represents a hallmark of SF3B1-mutated MDS ineffective erythropoiesis, which could be used as a therapeutic target.

[Pathophysiology of myelodysplastic syndromes]. / Physiopathologie des syndromes myélodysplasiques.

During aging, the onset of mutations at low frequency in hematopoietic cells or clonal hematopoiesis of indeterminate significance favors the evolution towards hemopathies such as myelodysplastic syndromes or acute leukemias, but also cardiovascular diseases and other pathologies. Acute or chronic inflammation related to age influences the clonal evolution and the immune response. Conversely, mutated hematopoietic cells create an inflammatory bone marrow environment facilitating their expansion. Various pathophysiological mechanisms depending on the type of mutation produce the diversity of phenotypes. Identifying factors affecting clonal selection is mandatory to improve patient care.

C/EBPα Confers Dependence to Fatty Acid Anabolic Pathways and Vulnerability to Lipid Oxidative Stress-Induced Ferroptosis in FLT3-Mutant Leukemia.

Although transcription factor CCAAT-enhancer binding protein α (C/EBPα) is critical for normal and leukemic differentiation, its role in cell and metabolic homeostasis is largely unknown in cancer. Here, multiomics analyses uncovered a coordinated activation of C/EBPα and Fms-like tyrosine kinase 3 (FLT3) that increased lipid anabolism in vivo and in patients with FLT3-mutant acute myeloid leukemia (AML). Mechanistically, C/EBPα regulated the fatty acid synthase (FASN)-stearoyl-CoA desaturase (SCD) axis to promote fatty acid (FA) biosynthesis and desaturation. We further demonstrated that FLT3 or C/EBPα inactivation decreased monounsaturated FA incorporation to membrane phospholipids through SCD downregulation. Consequently, SCD inhibition enhanced susceptibility to lipid redox stress that was exploited by combining FLT3 and glutathione peroxidase 4 inhibition to trigger lipid oxidative stress, enhancing ferroptotic death of FLT3-mutant AML cells. Altogether, our study reveals a C/EBPα function in lipid homeostasis and adaptation to redox stress, and a previously unreported vulnerability of FLT3-mutant AML to ferroptosis with promising therapeutic application. SIGNIFICANCE: FLT3 mutations are found in 30% of AML cases and are actionable by tyrosine kinase inhibitors. Here, we discovered that C/EBPα regulates FA biosynthesis and protection from lipid redox stress downstream mutant-FLT3 signaling, which confers a vulnerability to ferroptosis upon FLT3 inhibition with therapeutic potential in AML. This article is highlighted in the In This Issue feature, p. 1501.

[Identification and characterization of a monoclonal IgM reacting with disialylated gangliosides recognizing the CANOMAD syndrome]. / Identification et caractérisation d'une IgM monoclonale sérique à activité anti-gangliosides disialylés évoquant un Canomad syndrome.

We reported the laboratory phenotype of a monoclonal IgM-lambda against disialylated gangliosides, in a 81-year-old man admitted to a neurological department because of the progressive development of distal paresthesias, gait unsteadiness, difficulty to walk and having falls. Serological studies revealed an IgM monoclonal protein with lambda light chain component of MGUS type. IgM level was 4 g/L. The positive laboratory studies showed high titers of IgM antibodies in excess of 1/10(5) against specific disialylated gangliosides including GD1b, GD3, GT1b and GQ1b. There was no serum IgM binding to MAG and SGPG/SGLPG. Clonality by in-house immunodot of ganglioside antibodies was demonstrated using kappa and lambda light chain specific antibodies. Light chain subtype of the anti-ganglioside antibody activity and monoclonal IgM was lambda subtype. The reactivity at high titers was against gangliosides containing the disialosyl epitope. The clinical and laboratory features have been described under the acronym CANOMAD: Chronic Ataxic Neuropathy with Ophthalmoplegia, M proteins, cold Agglutinins and Disialosyl antibodies. Administration of IVIg produced a significant neurological improvement during six years. Then the neuropathy became refractory in the IVIg and worsened in severity, a cure by Rituximab® was established. The patient died from a pneumopathy only two months later. Monoclonal IgM binding to disialylated gangliosides have high level of specificity for diagnosis of the CANOMAD syndrome.

Comprehensive proteomic analysis of murine terminal erythroid differentiation.

Murine-based cellular models have provided and continue to provide many useful insights into the fundamental mechanisms of erythropoiesis, as well as insights into the pathophysiology of inherited and acquired red cell disorders. Although detailed information on many aspects of these cell models is available, comprehensive proteomic data are lacking. This is a critical knowledge gap, as proteins are effectors of most biologic processes. To address this critical unmet need, proteomes of the murine cell lines Friend erythroleukemia (MEL), GATA1 erythroid (G1ER), and embryonic stem cell-derived erythroid progenitor (MEDEP) and proteomes of cultured murine marrow-derived erythroblasts at different stages of terminal erythroid differentiation were analyzed. The proteomes of MEDEP cells and primary murine erythroid cells were most similar, whereas those of MEL and G1ER cells were more distantly related. We demonstrated that the overall cellular content of histones does not decrease during terminal differentiation, despite strong chromatin condensation. Comparison of murine and human proteomes throughout terminal erythroid differentiation revealed that many noted transcriptomic changes were significantly dampened at the proteome level, especially at the end of the terminal differentiation process. Analysis of the early events associated with induction of terminal differentiation in MEDEP cells revealed divergent alterations in associated transcriptomes and proteomes. These proteomic data are powerful and valuable tools for the study of fundamental mechanisms of normal and disordered erythropoiesis and will be of broad interest to a wide range of investigators for making the appropriate choice of various cell lines to study inherited and acquired diseases of the erythrocyte.

A variant erythroferrone disrupts iron homeostasis in SF3B1-mutated myelodysplastic syndrome.

Myelodysplastic syndromes (MDS) with ring sideroblasts are hematopoietic stem cell disorders with erythroid dysplasia and mutations in the SF3B1 splicing factor gene. Patients with MDS with SF3B1 mutations often accumulate excessive tissue iron, even in the absence of transfusions, but the mechanisms that are responsible for their parenchymal iron overload are unknown. Body iron content, tissue distribution, and the supply of iron for erythropoiesis are controlled by the hormone hepcidin, which is regulated by erythroblasts through secretion of the erythroid hormone erythroferrone (ERFE). Here, we identified an alternative ERFE transcript in patients with MDS with the SF3B1 mutation. Induction of this ERFE transcript in primary SF3B1-mutated bone marrow erythroblasts generated a variant protein that maintained the capacity to suppress hepcidin transcription. Plasma concentrations of ERFE were higher in patients with MDS with an SF3B1 gene mutation than in patients with SF3B1 wild-type MDS. Thus, hepcidin suppression by a variant ERFE is likely responsible for the increased iron loading in patients with SF3B1-mutated MDS, suggesting that ERFE could be targeted to prevent iron-mediated toxicity. The expression of the variant ERFE transcript that was restricted to SF3B1-mutated erythroblasts decreased in lenalidomide-responsive anemic patients, identifying variant ERFE as a specific biomarker of clonal erythropoiesis.