Transcriptomic analysis identifies lipid metabolism signature of defective erythropoiesis in diamond blackfan anemia syndrome

Abstract

Abstract Background: Diamond Blackfan anemia syndrome (DBAS) is a heterogeneous genetic disorder mainly caused by de novo heterozygous ribosomal protein variants, including RPL5. Analysis of DBAS cohorts and animal models has not revealed a potential mechanism for the variable anemia phenotype, which is a key feature of this disease. Treatment-independence occurs in approximately 20% of individuals who previously required steroids or red blood cell transfusions. The underlying mechanism of this phenomenon remains unknown. Design/Method We previously characterized Rpl5Skax23-Jus/+mice and demonstrated a severe defect in erythropoiesis at E12.5, which led to early embryonic demise in some mutants while others (within the same litter) survived and had complete resolution of anemia by 3 weeks of age. In order to further explore the mechanism leading to this defect, we performed timed matings (Rpl5Skax23-Jus/+ x wildtype (WT)) to obtain E12.5 fetal liver (FL) cells, which were sorted by flow cytometry to obtain CD71+ Ter119- early erythroid progenitor cells. Total RNA was extracted and we performed bulk RNA-seq analysis. We divided mutants into two groups based on liver cellularity with the hypothesis that mutants with very low cellularity (M-low) were the ones with impending erythroid failure and death, while those with close to normal cellularity (M-high) had a higher chance of spontaneous recovery. Results Analysis of RNA-seq data demonstrated downregulation of erythroid differentiation pathways consistent with the DBAS phenotype. We also identified dysregulation of lipid metabolism genes with significant downregulation of Scd1 in the subset of E12.5 mutant embryos at risk for complete erythroid failure (M-low). SCD1 is a key enzyme found in the endoplasmic reticulum, which catalyzes the conversion of saturated to monounsaturated fatty acids. The role of SCD1 and lipid metabolism in erythropoiesis and in DBA is currently unknown. To test the effect of Scd1 downregulation on erythropoiesis, we pretreated adult mice with a SCD1 inhibitor (SCD1-i) or DMSO daily for 2 weeks then administered phenylhydrazine following pretreatment in order to induce anemia.Mutant mice treated with DMSO showed a significant anemia compared with WT whereas SCD1-i treated mice had no or less significant differences in red blood cell counts. In order to explore the effect of the drug on erythropoiesis, we analyzed hematopoietic stem and progenitor cells by flow cytometry. Mice treated with SCD1-i showed a significant increase in CFU-E and decrease in pre-CFU-E counts in the bone marrow when compared to mice treated with DMSO indicating that downregulation of Scd1 is a compensatory mechanism to improve erythropoiesis in DBA. Conclusions We propose modulation of lipid metabolism and/or SCD1 as a possible mechanism for the variable anemia penetrance in DBAS and as a novel treatment strategy that warrants further study.