A molecular switch: Ro5-3335 drives hematopoietic stem cell division and clonality through distinct RUNX1 binding partners

Abstract

A defined number of hematopoietic stem cell (HSC) clones are born during development and expand to form the pool of adult stem cells. An intricate balance between self-renewal and differentiation of these HSCs supports hematopoiesis for life. Increasing the diversity of stem cell clones could have great impact on the hematopoietic stress response, for instance during chemotherapy, gene therapy or transplantation. Chemical screening for modulators of HSC expansion in zebrafish led us to identify Ro5-3335, which is known to bind to the transcription factor RUNX1. Diverse RUNX1 binding partners, including CBFb, allow specific regulation of downstream gene expression. RUNX1 is required for definitive hematopoiesis, and its loss has been shown to reduce clonal diversity. We find that Ro5-3335 expands HSCs in zebrafish by inducing more symmetric cell divisions, leading to an increase in HSC clone number. Ro5-3335 also enhances clonal diversity in mouse HSC cultures. Using human CD34+ HSPCs, our studies reveal that Ro5-3335 remodels the RUNX1 transcription complex by a novel recruitment of ELF1/2, without altering CBFb DNA binding affinity. This results in specific expression of cell cycle and hematopoietic genes that enhance HSC self-renewal and prevent differentiation. Our work provides the first reported evidence of an increase in HSC clone number by chemical induction in vivo, via novel recruitment of ELF1/2 to RUNX1, which could help develop treatments that enhance clonal diversity for blood diseases.