Deciphering and disrupting PIEZO1-TMEM16F interplay in hereditary xerocytosis.

ABSTRACT: Cell-surface exposure of phosphatidylserine (PS) is essential for phagocytic clearance and blood clotting. Although a calcium-activated phospholipid scramblase (CaPLSase) has long been proposed to mediate PS exposure in red blood cells (RBCs), its identity, activation mechanism, and role in RBC biology and disease remain elusive. Here, we demonstrate that TMEM16F, the long-sought-after RBC CaPLSase, is activated by calcium influx through the mechanosensitive channel PIEZO1 in RBCs. PIEZO1-TMEM16F functional coupling is enhanced in RBCs from individuals with hereditary xerocytosis (HX), an RBC disorder caused by PIEZO1 gain-of-function channelopathy. Enhanced PIEZO1-TMEM16F coupling leads to an increased propensity to expose PS, which may serve as a key risk factor for HX clinical manifestations including anemia, splenomegaly, and postsplenectomy thrombosis. Spider toxin GsMTx-4 and antigout medication benzbromarone inhibit PIEZO1, preventing force-induced echinocytosis, hemolysis, and PS exposure in HX RBCs. Our study thus reveals an activation mechanism of TMEM16F CaPLSase and its pathophysiological function in HX, providing insights into potential treatment.

A randomized clinical trial of the efficacy and safety of rivipansel for sickle cell vaso-occlusive crisis.

The efficacy and safety of rivipansel, a predominantly E-selectin antagonist, were studied in a phase 3, randomized, controlled trial for vaso-occlusive crisis (VOC) requiring hospitalization (RESET). A total of 345 subjects (204 adults and 141 children) were randomized and 320 were treated (162 with rivipansel, 158 with placebo) with an IV loading dose, followed by up to 14 additional 12-hourly maintenance doses of rivipansel or placebo, in addition to standard care. Rivipansel was similarly administered during subsequent VOCs in the Open-label Extension (OLE) study. In the full analysis population, the median time to readiness for discharge (TTRFD), the primary end point, was not different between rivipansel and placebo (-5.7 hours, P = .79; hazard ratio, 0.97), nor were differences seen in secondary end points of time to discharge (TTD), time to discontinuation of IV opioids (TTDIVO), and cumulative IV opioid use. Mean soluble E-selectin decreased 61% from baseline after the loading dose in the rivipansel group, while remaining unchanged in the placebo group. In a post hoc analysis, early rivipansel treatment within 26.4 hours of VOC pain onset (earliest quartile of time from VOC onset to treatment) reduced median TTRFD by 56.3 hours, reduced median TTD by 41.5 hours, and reduced median TTDIVO by 50.5 hours, compared with placebo (all P < .05). A similar subgroup analysis comparing OLE early-treatment with early-treatment RESET placebo showed a reduction in TTD of 23.1 hours (P = .062) and in TTDIVO of 30.1 hours (P = .087). Timing of rivipansel administration after pain onset may be critical to achieving accelerated resolution of acute VOC. Trial Registration: Clinicaltrials.gov, NCT02187003 (RESET), NCT02433158 (OLE).

Nontargeted Plasma Proteomic Analysis of Renal Disease and Pulmonary Hypertension in Patients with Sickle Cell Disease.

Sickle cell disease (SCD) is characterized by red blood cell sickling, vaso-occlusion, hemolytic anemia, damage to multiple organ systems, and, as a result, shortened life expectancy. Sickle cell disease nephropathy (SCDN) and pulmonary hypertension (pHTN) are common and frequently co-occurring complications of SCD; both are associated with markedly accelerated mortality. To identify candidate circulating biomarkers of SCDN and pHTN, we used mass spectrometry to quantify the relative abundance of >1000 proteins in plasma samples from 189 adults with SCD from the Outcome Modifying Genes in SCD (OMG-SCD) cohort (ProteomeXchange identifier PXD048716). Forty-four proteins were differentially abundant in SCDN, most significantly cystatin-C and collagen α-1(XVIII) chain (COIA1), and 55 proteins were dysregulated in patients with SCDN and pHTN, most significantly insulin-like growth factor-binding protein 6 (IBP6). Network analysis identified a module of 133 coregulated proteins significantly associated with SCDN, that was enriched for extracellular matrix proteins, insulin-like growth factor binding proteins, cell adhesion proteins, EGF-like calcium binding proteins, and several cadherin family members. Collectively, these data provide a comprehensive understanding of plasma protein changes in SCDN and pHTN which validate numerous studies of chronic kidney disease and suggest shared profiles of protein disruption in kidney dysfunction and pHTN among SCD patients.

A genome-wide association study of alloimmunization in the TOPMed OMG-SCD cohort identifies a locus on chromosome 12.

BACKGROUND: Red cell alloimmunization after exposure to donor red cells is a very common complication of transfusion for patients with sickle cell disease (SCD), resulting frequently in accelerated donor red blood cell destruction. Patients show substantial differences in their predisposition to alloimmunization, and genetic variability is one proposed component. Although several genetic association studies have been conducted for alloimmunization, the results have been inconsistent, and the genetic determinants of alloimmunization remain largely unknown. STUDY DESIGN AND METHODS: We performed a genome-wide association study (GWAS) in 236 African American (AA) SCD patients from the Outcome Modifying Genes in Sickle Cell Disease (OMG-SCD) cohort, which is part of Trans-Omics for Precision Medicine (TOPMed), with whole-genome sequencing data available. We also performed sensitivity analyses adjusting for different sets of covariates and applied different sample grouping strategies based on the number of alloantibodies patients developed. RESULTS: We identified one genome-wide significant locus on chr12 (p = 3.1e-9) with no evidence of genomic inflation (lambda = 1.003). Further leveraging QTL evidence from GTEx whole blood and/or Jackson Heart Study PBMC RNA-Seq data, we identified a number of potential genes, such as ARHGAP9, STAT6, and ATP23, that may be driving the association signal. We also discovered some suggestive loci using different analysis strategies. DISCUSSION: We call for the community to collect additional alloantibody information within SCD cohorts to further the understanding of the genetic basis of alloimmunization in order to improve transfusion outcomes.

Red cell exchange modulates neutrophil degranulation responses in sickle cell disease.

BACKGROUND: Neutrophils in sickle cell disease (SCD) are activated, contributing to disease. Red cell exchange (RCE), with the goal of lowering hemoglobin S (HbS), is an important part of therapy for many SCD patients. Whether RCE impacts neutrophil reactivity is unknown. STUDY DESIGN AND METHODS: To determine the effect of RCE on neutrophil activation, SCD patients undergoing RCE in steady-state were enrolled. Neutrophil degranulation responses were examined before/after RCE. Kinetic studies were completed to determine the duration of the effect of RCE on neutrophil function. Degranulation results were examined in relation to white blood cell count, neutrophil count, and HbS levels. The effect of RCE on RBC phosphatidylserine (PS) exposure was examined as a possible contributor to modulation of neutrophil function by RCE. RESULTS: Twenty-two patients with SCD, genotype SS, who underwent RCE (average pre-RCE HbS 33 ± 14%) were included for the study. RCE significantly decreased neutrophil degranulation responses. The effect of RCE on neutrophil activation was unrelated to cell count and instead directly correlated with HbS. The effect of RCE on neutrophil activation was sustained over several days post-apheresis. Furthermore, while increased RBC PS exposure results in increased neutrophil degranulation, RCE decreases RBC PS exposure. DISCUSSION: To our knowledge, this is the first study demonstrating that RCE significantly decreases neutrophil activation in a sustained HbS-dependent manner. Modulation of PS exposure by RCE may be a contributing mechanism by which RCE modulates neutrophil activation. These studies raise the possibility that modulation of neutrophil activation contributes significantly to the therapeutic effect of RCE.

Variation and impact of polygenic hematologic traits in monogenic sickle cell disease.

Several of the complications observed in sickle cell disease (SCD) are influenced by variation in hematologic traits (HT), such as fetal hemoglobin (HbF) level and neutrophil count. Previous large-scale genome-wide association studies carried out in largely healthy individuals have identified thousands of variants associated with HT, which have then been used to develop multi-ancestry polygenic trait scores (PTS). Here, we tested whether these PTS associate with HT in SCD patients and if they can improve statistical models associated with SCD-related complications. In 2,056 SCD patients, we found that the PTS predicted less HT variance than in non-SCD individuals of African ancestry. This was particularly striking at the Duffy/DARC locus, where we observed an epistatic interaction between the SCD genotype and the Duffy null variant (rs2814778) that led to a two-fold weaker effect on neutrophil count. PTS for these HT which are measured as part of routine practice were not associated with complications in SCD. In contrast, we found that a simple PTS for HbF that includes only six variants explained a large fraction of the phenotypic variation (20.5-27.1%), associated with acute chest syndrome and stroke risk, and improved the statistical modeling of the vaso-occlusive crisis rate. Using Mendelian randomization, we found that increasing HbF by 4.8% reduces stroke risk by 39% (P=0.0006). Taken together, our results highlight the importance of validating PTS in large diseased populations before proposing their implementation in the context of precision medicine initiatives.

Design, Synthesis, and Investigation of Novel Nitric Oxide (NO)-Releasing Aromatic Aldehydes as Drug Candidates for the Treatment of Sickle Cell Disease.

Sickle cell disease (SCD) is caused by a single-point mutation, and the ensuing deoxygenation-induced polymerization of sickle hemoglobin (HbS), and reduction in bioavailability of vascular nitric oxide (NO), contribute to the pathogenesis of the disease. In a proof-of-concept study, we successfully incorporated nitrate ester groups onto two previously studied potent antisickling aromatic aldehydes, TD7 and VZHE039, to form TD7-NO and VZHE039-NO hybrids, respectively. These compounds are stable in buffer but demonstrated the expected release of NO in whole blood in vitro and in mice. The more promising VZHE039-NO retained the functional and antisickling activities of the parent VZHE039 molecule. Moreover, VZHE039-NO, unlike VZHE039, significantly attenuated RBC adhesion to laminin, suggesting this compound has potential in vivo RBC anti-adhesion properties relevant to vaso-occlusive events. Crystallographic studies show that, as with VZHE039, VZHE039-NO also binds to liganded Hb to make similar protein interactions. The knowledge gained during these investigations provides a unique opportunity to generate a superior candidate drug in SCD with enhanced benefits.

Potential causal role of l-glutamine in sickle cell disease painful crises: A Mendelian randomization analysis.

In a recent clinical trial, the metabolite l-glutamine was shown to reduce painful crises in sickle cell disease (SCD) patients. To support this observation and identify other metabolites implicated in SCD clinical heterogeneity, we profiled 129 metabolites in the plasma of 705 SCD patients. We tested correlations between metabolite levels and six SCD-related complications (painful crises, cholecystectomy, retinopathy, leg ulcer, priapism, aseptic necrosis) or estimated glomerular filtration rate (eGFR), and used Mendelian randomization (MR) to assess causality. We found a potential causal relationship between l-glutamine levels and painful crises (N = 1278, odds ratio (OR) [95% confidence interval] = 0.68 [0.52-0.89], P = 0.0048). In two smaller SCD cohorts (N = 299 and 406), the protective effect of l-glutamine was observed (OR = 0.82 [0.50-1.34]), although the MR result was not significant (P = 0.44). We identified 66 significant correlations between the levels of other metabolites and SCD-related complications or eGFR. We tested these correlations for causality using MR analyses and found no significant causal relationship. The baseline levels of quinolinic acid were associated with prospectively ascertained survival in SCD patients, and this effect was dependent on eGFR. Metabolomics provide a promising approach to prioritize small molecules that may serve as biomarkers or drug targets in SCD.

Rapid decline in estimated glomerular filtration rate in sickle cell anemia: results of a multicenter pooled analysis.

Not available.

Diversity of variant alleles encoding Kidd, Duffy, and Kell antigens in individuals with sickle cell disease using whole genome sequencing data from the NHLBI TOPMed Program.

BACKGROUND: Genetic variants in the SLC14A1, ACKR1, and KEL genes, which encode Kidd, Duffy, and Kell red blood cell antigens, respectively, may result in weakened expression of antigens or a null phenotype. These variants are of particular interest to individuals with sickle cell disease (SCD), who frequently undergo chronic transfusion therapy with antigen-matched units. The goal was to describe the diversity and the frequency of variants in SLC14A1, ACKR1, and KEL genes among individuals with SCD using whole genome sequencing (WGS) data. STUDY DESIGN AND METHODS: Two large SCD cohorts were studied: the Recipient Epidemiology and Donor Evaluation Study III (REDS-III) (n = 2634) and the Outcome Modifying Gene in SCD (OMG) (n = 640). Most of the studied individuals were of mixed origin. WGS was performed as part of the National Heart, Lung, and Blood Institute's Trans-Omics for Precision Medicine (TOPMed) program. RESULTS: In SLC14A1, variants included four encoding a weak Jka phenotype and five null alleles (JKnull ). JKA*01N.09 was the most common JKnull . One possible JKnull mutation was novel: c.812G>T. In ACKR1, identified variants included two that predicted Fyx (FY*X) and one corresponding to the c.-67T>C GATA mutation. The c.-67T>C mutation was associated with FY*A (FY*01N.01) in four participants. FY*X was identified in 49 individuals. In KEL, identified variants included three null alleles (KEL*02N.17, KEL*02N.26, and KEL*02N.04) and one allele predicting Kmod phenotype, all in heterozygosity. CONCLUSIONS: We described the diversity and distribution of SLC14A1, ACKR1, and KEL variants in two large SCD cohorts, comprising mostly individuals of mixed ancestry. This information may be useful for planning the transfusion support of patients with SCD.

Feasibility of and barriers to thalassemia screening in migrant populations: a cross-sectional study of Myanmar and Cambodian migrants in Thailand.

BACKGROUND: Thalassemia, an inherited hemoglobin disorder, has become a global public health problem due to population migration. Evidence-based strategies for thalassemia prevention in migrants are lacking. We characterized barriers to thalassemia screening and the burden of thalassemia in migrant workers in Thailand. METHODS: Multilingual demographic and KAP surveys were completed by 197 Thai, 119 Myanmar, and 176 Cambodian adults residing in Thailand. Thalassemia awareness, socio-demographic predictors, and knowledge and attitude scores were compared between migrant and Thai subjects. Comprehensive thalassemia testing was performed for migrants. RESULTS: Migrants had extremely poor thalassemia awareness (4.1%) compared to Thai subjects (79.6%) and had lower thalassemia knowledge scores but similar attitude scores. Surveys identified differing sociodemographic factors predicting awareness in Thai and migrant subjects, as well as key misconceptions likely to hinder thalassemia screening uptake. Nearly all migrants consented to thalassemia testing. We identified abnormal hemoglobin profiles in 52.7% of migrants and a higher projected rate of severe thalassemia births in migrants. CONCLUSIONS: The high burden of thalassemia and tremendous knowledge gap in migrants needs urgent attention. Thalassemia screening was feasible and acceptable in our migrant population. Sociocultural and structural barriers merit further attention when designing thalassemia screening and prevention policies for migrants in Thailand and globally.

Identification of optimal thalassemia screening strategies for migrant populations in Thailand using a qualitative approach.

BACKGROUND: Thalassemia is a common inherited hemoglobin disorder in Southeast Asia. Severe thalassemia can lead to significant morbidity for patients and economic strain for under-resourced health systems. Thailand's thalassemia prevention and control program has successfully utilized prenatal screening and diagnosis to reduce the incidence of severe thalassemia in Thai populations, but migrant populations are excluded despite having high thalassemia prevalence. We sought to identify key barriers to and facilitators of thalassemia screening and to develop tailored recommendations for providing migrants with access to thalassemia prevention and control. METHODS: We conducted 28 in-depth interviews and 4 focus group discussions (FGDs) in Chonburi, Thailand with Myanmar and Cambodian migrants, Thai healthcare providers, Thai parents of children affected by thalassemia, and migrant agents. RESULTS: Participant narratives revealed that migrants' lack of knowledge about the prevalence, manifestations, severity, and inherited nature of thalassemia led to misconceptions, fear, or indifference toward thalassemia and screening. Negative perceptions of pregnancy termination were based in religious beliefs but compounded by other sociocultural factors, presenting a key obstacle to migrant uptake of prenatal screening. Additionally, structural barriers included legal status, competing work demands, lack of health insurance, and language barriers. Participants recommended delivering public thalassemia education in migrants' native languages, implementing carrier screening, and offering thalassemia screening in convenient settings. CONCLUSIONS: An effective thalassemia prevention and control program should offer migrants targeted thalassemia education and outreach, universal coverage for thalassemia screening and prenatal care, and options for carrier screening, providing a comprehensive strategy for reducing the incidence of severe thalassemia in Thailand and establishing an inclusive model for regional thalassemia prevention and control.

Beyond hydroxyurea: new and old drugs in the pipeline for sickle cell disease.

Despite Food and Drug Administration (FDA) approval of hydroxyurea to reduce the frequency of vaso-occlusive episodes, sickle cell disease (SCD) has continued to be treated primarily with analgesics for pain relief. However, elucidation of the multiple pathophysiologic mechanisms leading to vaso-occlusion and tissue injury in SCD has now resulted in a burgeoning effort to identify new treatment modalities to prevent or ameliorate the consequences of the disease. Development of new drugs as well as investigation of drugs previously used in other settings have targeted cell adhesion, inflammatory pathways, upregulation of hemoglobin F, hemoglobin polymerization and sickling, coagulation, and platelet activation. Although these efforts have not yet yielded drugs ready for FDA approval, several early studies have been extremely encouraging. Moreover, the marked increase in clinical pharmaceutical research addressing SCD and the new and old drugs in the pipeline make it reasonable to expect that we will soon have new treatments for SCD.

Clinical and metabolomic risk factors associated with rapid renal function decline in sickle cell disease.

Sickle cell disease (SCD) nephropathy and lower estimated glomerular filtration rate (eGFR) are risk factors for early mortality. Furthermore, rate of eGFR decline predicts progression to end-stage renal disease in many clinical settings. However, factors predicting renal function decline in SCD are poorly documented. Using clinical, laboratory, genetic, and metabolomic data, we evaluated predictors of renal function decline in a longitudinal cohort of 288 adults (mean age 33.0 years). In 193 subjects with 5-year follow-up data, mean rate of eGFR decline was 2.35 mL/min/1.73 m2 /year, nearly twice that of African American adults overall. Hyperfiltration was prevalent at baseline (61.1%), and 36.8% of subjects experienced rapid eGFR decline (≥3 mL/min/1.73 m2 /year). Severe Hb genotype; proteinuria; higher platelet and reticulocyte counts, and systolic BP; and lower Hb level and BMI were associated with rapid decline. A risk scoring system was created using these 7 variables and was highly predictive of rapid eGFR decline, with odds of rapid decline increasing 1.635-fold for every point increment (P < 0.0001). Rapid eGFR decline was also associated with higher organ system severity score and peak creatinine. Additionally, two metabolites (asymmetric dimethylarginine and quinolinic acid) were associated with rapid decline. Further investigation into longitudinal SCD nephropathy (SCDN) trajectory, early markers of SCDN, and tools for risk stratification should inform interventional studies targeted to slowing GFR decline and improving SCD outcomes.

In vivo Modeling Implicates APOL1 in Nephropathy: Evidence for Dominant Negative Effects and Epistasis under Anemic Stress.

African Americans have a disproportionate risk for developing nephropathy. This disparity has been attributed to coding variants (G1 and G2) in apolipoprotein L1 (APOL1); however, there is little functional evidence supporting the role of this protein in renal function. Here, we combined genetics and in vivo modeling to examine the role of apol1 in glomerular development and pronephric filtration and to test the pathogenic potential of APOL1 G1 and G2. Translational suppression or CRISPR/Cas9 genome editing of apol1 in zebrafish embryos results in podocyte loss and glomerular filtration defects. Complementation of apol1 morphants with wild-type human APOL1 mRNA rescues these defects. However, the APOL1 G1 risk allele does not ameliorate defects caused by apol1 suppression and the pathogenicity is conferred by the cis effect of both individual variants of the G1 risk haplotype (I384M/S342G). In vivo complementation studies of the G2 risk allele also indicate that the variant is deleterious to protein function. Moreover, APOL1 G2, but not G1, expression alone promotes developmental kidney defects, suggesting a possible dominant-negative effect of the altered protein. In sickle cell disease (SCD) patients, we reported previously a genetic interaction between APOL1 and MYH9. Testing this interaction in vivo by co-suppressing both transcripts yielded no additive effects. However, upon genetic or chemical induction of anemia, we observed a significantly exacerbated nephropathy phenotype. Furthermore, concordant with the genetic interaction observed in SCD patients, APOL1 G2 reduces myh9 expression in vivo, suggesting a possible interaction between the altered APOL1 and myh9. Our data indicate a critical role for APOL1 in renal function that is compromised by nephropathy-risk encoding variants. Moreover, our interaction studies indicate that the MYH9 locus is also relevant to the phenotype in a stressed microenvironment and suggest that consideration of the context-dependent functions of both proteins will be required to develop therapeutic paradigms.

Randomized phase 2 study of GMI-1070 in SCD: reduction in time to resolution of vaso-occlusive events and decreased opioid use.

Treatment of vaso-occlusive crises (VOC) or events in sickle cell disease (SCD) remains limited to symptom relief with opioids. Animal models support the effectiveness of the pan-selectin inhibitor GMI-1070 in reducing selectin-mediated cell adhesion and abrogating VOC. We studied GMI-1070 in a prospective multicenter, randomized, placebo-controlled, double-blind, phase 2 study of 76 SCD patients with VOC. Study drug (GMI-1070 or placebo) was given every 12 hours for up to 15 doses. Other treatment was per institutional standard of care. All subjects reached the composite primary end point of resolution of VOC. Although time to reach the composite primary end point was not statistically different between the groups, clinically meaningful reductions in mean and median times to VOC resolution of 41 and 63 hours (28% and 48%, P = .19 for both) were observed in the active treatment group vs the placebo group. As a secondary end point, GMI-1070 appeared safe in acute vaso-occlusion, and adverse events were not different in the two arms. Also in secondary analyses, mean cumulative IV opioid analgesic use was reduced by 83% with GMI-1070 vs placebo (P = .010). These results support a phase 3 study of GMI-1070 (now rivipansel) for SCD VOC. This trial was registered at www.clinicaltrials.gov as #NCT01119833.

Sevuparin binds to multiple adhesive ligands and reduces sickle red blood cell-induced vaso-occlusion.

Sevuparin is a novel drug candidate in phase II development as a treatment for vaso-occlusive crises (VOC) in patients with sickle cell disease (SCD). As a heparin-derived polysaccharide, sevuparin has been designed to retain anti-adhesive properties, while the antithrombin-binding domains have been eliminated, substantially diminishing its anticoagulant activity. Here, we demonstrate that sevuparin inhibits the adhesion of human sickle red blood cells (SS-RBCs) to stimulated cultured endothelial cells in vitro. Importantly, sevuparin prevents vaso-occlusion and normalizes blood flow in an in vivo mouse model of SCD vaso-occlusion. Analyses by surface plasmon resonance (SPR) and fluorescence correlation spectroscopy (FCS) demonstrate that sevuparin binds to P- and L-selectins, thrombospondin, fibronectin and von Willebrand factor, all of which are thought to contribute to vaso-occlusion in SCD. Despite low anticoagulation activity, sevuparin has anti-adhesive efficacy similar to the low molecular weight heparin tinzaparin both in vitro and in vivo. These results suggest that the anti-adhesive properties rather than the anticoagulant effects of heparinoids are critical for the treatment of vaso-occlusion in SCD. Therefore, sevuparin is now being evaluated in SCD patients hospitalized for treatment of VOC.

A comprehensive joint analysis of the long and short RNA transcriptomes of human erythrocytes.

BACKGROUND: Human erythrocytes are terminally differentiated, anucleate cells long thought to lack RNAs. However, previous studies have shown the persistence of many small-sized RNAs in erythrocytes. To comprehensively define the erythrocyte transcriptome, we used high-throughput sequencing to identify both short (18-24 nt) and long (>200 nt) RNAs in mature erythrocytes. RESULTS: Analysis of the short RNA transcriptome with miRDeep identified 287 known and 72 putative novel microRNAs. Unexpectedly, we also uncover an extensive repertoire of long erythrocyte RNAs that encode many proteins critical for erythrocyte differentiation and function. Additionally, the erythrocyte long RNA transcriptome is significantly enriched in the erythroid progenitor transcriptome. Joint analysis of both short and long RNAs identified several loci with co-expression of both microRNAs and long RNAs spanning microRNA precursor regions. Within the miR-144/451 locus previously implicated in erythroid development, we observed unique co-expression of several primate-specific noncoding RNAs, including a lncRNA, and miR-4732-5p/-3p. We show that miR-4732-3p targets both SMAD2 and SMAD4, two critical components of the TGF-ß pathway implicated in erythropoiesis. Furthermore, miR-4732-3p represses SMAD2/4-dependent TGF-ß signaling, thereby promoting cell proliferation during erythroid differentiation. CONCLUSIONS: Our study presents the most extensive profiling of erythrocyte RNAs to date, and describes primate-specific interactions between the key modulator miR-4732-3p and TGF-ß signaling during human erythropoiesis.