We seek to identify new regulators of red blood cell production (erythropoiesis) by taking advantage of the robust statistical findings from genome-wide association studies (GWAS). While GWAS have identified thousands of loci across the human genome that are associated with various diseases or traits, the biological underpinnings for the vast majority of these loci remain unclear. This has given rise to skepticism about the value of GWAS for gaining a deeper understanding of clinical medicine or functional biology. We have previously utilized the results of erythroid trait GWAS to identify important regulators of erythropoiesis and globin gene regulation. This included work uncovering the first specific regulator of the fetal-to-adult hemoglobin switch, BCL11A, and work that revealed a critical role for cyclin D3 in terminal erythropoiesis. In this project proposal, we aim to extend these findings to over 50 loci associated with erythroid traits in humans in an attempt to uncover new regulators of erythropoiesis. We initially plan to map and identify all candidate genes in the loci revealed from erythroid trait GWAS. This will be accomplished by taking advantage of resources that identify common human variation and allow us to infer linkage disequilibrium patterns. We then plan to functionally screen all candidate genes in these regions using a loss-of-function pooled short hairpin RNA (shRNA) screen in primary human erythroid cells. This will allow us to identify candidate regulators of erythropoiesis from these regions. We then plan to perform in-depth analysis of these candidate regulators in human cells and in mouse models to define the exact role of these candidates in erythropoiesis. This will be accomplished through the use of both loss and gain-of-function perturbation of these candidate genes. Finally, we aim to examine whether the regulators of erythropoiesis identified through our studies can be manipulated to improve the current methods of ex vivo red blood cell production, which is a problem of immediate clinical relevance. Limitations in the blood supply of certain antigen-containing red blood cells pose a considerable clinical problem. Such ex vivo-derived red cells could serve as an alternative to standard transfusions in cases where such problems arise. This project has the potential to substantially advance two distinct areas of biomedical research: (1) The findings from these studies will identify new regulators of erythropoiesis that could be involved in, modifiers of, or help identify therapeutic avenues to treat various forms of anemia. Our prior work on BCL11A illustrates how studying a fundamental biological problem (fetal hemoglobin regulation) can result in promising therapeutic insight. (2) The functional approaches undertaken in this project could provide resources and serve as a paradigm for future research aimed at identifying the biological underpinnings of other GWAS.

Public Health Relevance

Anemia is estimated to affect 25 percent of the world population and the majority of cases are attributable to defective production of red blood cells (erythropoiesis). This project utilizes the robust statistical findings from prior genome-wide association studies to identify new regulators of erythropoiesis in humans through the use of a functional screen. Identification and characterization of such regulators should pave the way for an improved understanding of why certain forms of anemia occur and may lead to new treatment approaches. (End of Abstract)

National Institute of Health (NIH)
National Heart, Lung, and Blood Institute (NHLBI)
Exploratory/Developmental Grants (R21)
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Special Emphasis Panel (ZHL1-CSR-H (S1))
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Qasba, Pankaj
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Children's Hospital Boston
United States
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Polfus, Linda M; Khajuria, Rajiv K; Schick, Ursula M et al. (2016) Whole-Exome Sequencing Identifies Loci Associated with Blood Cell Traits and Reveals a Role for Alternative GFI1B Splice Variants in Human Hematopoiesis. Am J Hum Genet 99:481-8
Wakabayashi, Aoi; Ulirsch, Jacob C; Ludwig, Leif S et al. (2016) Insight into GATA1 transcriptional activity through interrogation of cis elements disrupted in human erythroid disorders. Proc Natl Acad Sci U S A 113:4434-9
Ulirsch, Jacob C; Nandakumar, Satish K; Wang, Li et al. (2016) Systematic Functional Dissection of Common Genetic Variation Affecting Red Blood Cell Traits. Cell 165:1530-1545
Giani, Felix C; Fiorini, Claudia; Wakabayashi, Aoi et al. (2016) Targeted Application of Human Genetic Variation Can Improve Red Blood Cell Production from Stem Cells. Cell Stem Cell 18:73-78
Nandakumar, Satish K; Ulirsch, Jacob C; Sankaran, Vijay G (2016) Advances in understanding erythropoiesis: evolving perspectives. Br J Haematol 173:206-18
Basak, Anindita; Sankaran, Vijay G (2016) Regulation of the fetal hemoglobin silencing factor BCL11A. Ann N Y Acad Sci 1368:25-30
Wakabayashi, Aoi; Sankaran, Vijay G (2016) Society for Pediatric Research 2015 Young Investigator Award: genetics of human hematopoiesis-what patients can teach us about blood cell production. Pediatr Res 79:366-70
Sankaran, Vijay G; Ulirsch, Jacob C; Tchaikovskii, Vassili et al. (2015) X-linked macrocytic dyserythropoietic anemia in females with an ALAS2 mutation. J Clin Invest 125:1665-9
Ludwig, Leif S; Cho, Hyunjii; Wakabayashi, Aoi et al. (2015) Genome-wide association study follow-up identifies cyclin A2 as a regulator of the transition through cytokinesis during terminal erythropoiesis. Am J Hematol 90:386-91
Basak, Anindita; Hancarova, Miroslava; Ulirsch, Jacob C et al. (2015) BCL11A deletions result in fetal hemoglobin persistence and neurodevelopmental alterations. J Clin Invest 125:2363-8

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