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144 Rps19 and Rpl5 Play Distinct Roles in hematopoietic Stem Cell maintenance and Erythroid Differentiation

Program: Oral and Poster Abstracts
Type: Oral
Session: 101. Red Cells and Erythropoiesis, Excluding Iron: Red Cells: Erythropoiesis in Stress and Disease
Hematology Disease Topics & Pathways:
Research, Fundamental Science, Bone Marrow Failure Syndromes, Inherited Marrow Failure Syndromes, Genetic Disorders, Diseases
Saturday, December 9, 2023: 3:15 PM

Yuefeng Tang, PhD1, Te Ling, PhD2, Mushran Khan3*, Riaz Rao4*, Vincent P Schulz, PhD5*, Julien Papoin, MS4*, Anupama Narla, MD6, Jeffrey Michael Lipton, MD, PhD7, James Palis, MD8, Laurie A. Steiner, MD9, Patrick G Gallagher, MD10, Mohandas Narla, DSc11, John D. Crispino, PhD, MBA12 and Lionel Blanc, PhD4

1The Feinstein Institutes for Medical Research, Northwell Health, Manhasset, NY
2St. Jude Children's Research Hospital, Memphis, TN
3The Feinstein Institute For Medical Research, Manhasset, NY
4Feinstein Institutes for Medical Research, Northwell, Manhasset, NY
5Yale School of Medicine, New Haven, CT
6Stanford University, Stanford, CA
7Cohen Children's Medical Center of NY, New Hyde Park, NY
8University of Rochester, Rochester, NY
9Department of Pediatrics, University of Rochester, Rochester, NY
10Yale University School of Medicine, New Haven, CT
11LFKRI, New York Blood Center Enterprises, New York, NY
12Division of Experimental Hematology, St Jude Children's Research Hospital, Memphis, TN

Mutations in ribosomal proteins (RPs) lead to ribosomopathies. Mechanistic basis underlying the failure of erythropoiesis due to mutations in various RPs is yet to be fully defined. Lack of relevant murine models of ribosomapathies has hindered significant progress. To fill this gap, we generated Rps19 and Rpl5 floxed mice using CRISPR/Cas9 technologies and characterized their hematologic phenotype following crossing them to a Vav-iCremice. Vav-iCre;Rps19fl/+ mice are born at expected Mendelian rates with severe macrocytic anemia and reticulocytopenia. These mice die by post-natal day 10 (P10), due to bone marrow failure and absence of splenic stress erythropoiesis. Analysis of fetal erythropoiesis revealed a 40% reduction in the total number of erythroid cells in Vav-iCre;Rps19fl/+, however erythroblasts at all stages of terminal erythroid differentiation were found. Intriguingly, studies of hematopoietic stem and progenitor cells demonstrated a progressive exhaustion of the stem cell compartment.

scRNAseq studies of Vav-iCre;Rps19fl/+ fetal livers demonstrated increased expression of RUNX1, a transcription factor implicated in the regulation of ribosome biogenesis. Accordingly, we observed that hematopoietic progenitors of Vav-iCre,Rps19fl/+ had enhanced global protein synthesis along with increased RUNX1 protein, despite ribosomal protein haploinsufficiency demonstrated by polysome profiling.

In marked contrast, Vav-iCre;Rpl5fl/+ mice presented with a more severe phenotype and died perinatally from anemia. Further analysis revealed a failure of erythropoiesis at erythroid progenitor stages in Vav-iCre;Rpl5fl/+. In striking contrast with the Rps19 model, Vav-iCre;Rpl5fl/+ mice had an expansion of the hematopoietic stem and progenitor cell compartment that also had an increased protein translation rate. This increased translation occurred in the context of normal expression levels of RUNX1. Interestingly, Vav-iCre;Rpl5fl/+ mice had reduced mTORC1 activity and a dramatic reduction in the expression of other ribosomal proteins. Consistent with previous studies about the pivotal role of mTORC1 in erythropoiesis, Vav-iCre,Raptorfl/fl (mTORC1 conditional knockout) had reduced expressions of RPs in fetal liver.

In summary, we report different requirements for Rps19 and Rpl5 during fetal hematopoiesis in regulating erythroid progenitor biology. While Rps19 plays an essential role at the HSC level to maintain the generation of erythroid progenitors, Rpl5 is critical once progenitors are committed towards an erythroid fate. Along with the findings that translation is increased in RP haploinsufficient models, our study unravels potential compensatory mechanisms during fetal hematopoiesis.

Disclosures: Crispino: SAB of Alethiomics: Other: Member; Cellarity: Consultancy.

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