3175 – HEMATOPOIETIC STEM CELL-DEPENDENT NOTCH TRANSCRIPTION IS MEDIATED BY P53 THROUGH THE HISTONE CHAPERONE SUPT16

Abstract

Hematopoietic stem cells (HSCs) are rare cells that can self-renew and differentiate into all blood cell lineages for life. HSCs have long been the focus of developmental and regenerative studies, yet our understanding of the signaling events regulating their specification remains incomplete. In order to identify novel genes and transcription factors involved in hematopoietic specification, we performed a forward genetic screen to identify zebrafish mutants defective in HSC formation. Through large-scale whole mount in situ hybridization based screens followed by RNA-sequencing-based linkage mapping, we identified that supt16h, a component of the FAcilitates Chromatin Transcription (FACT) complex, is required for HSC formation. Zebrafish supt16h mutants express reduced levels of Notch signaling components, genes essential for HSC development, due to abrogated transcription. Although cellular functions of Supt16h is generally known for regulating transcription and reorganizing nucleosomes to alter chromatin accessibility, global chromatin accessibility in the zebrafish supt16h mutants is unaffected. However, we observe a specific increase in accessibility at the p53 locus leading to an accumulation of P53 protein in the supt16h mutants and abrogation of increased p53 levels in supt16h mutants rescues both loss of Notch and HSC phenotypes. We further demonstrate that P53 levels directly influence expression of the Polycomb Group protein, Phc1, which functions as a transcriptional repressor of Notch genes. Suppression of phc1 or its upstream regulator, p53, rescues both loss of Notch and loss of HSC phenotypes in supt16h mutants. Taken together, our results highlight a previously uncharacterized relationship between supt16h, p53, and phc1 to specify HSCs via modulation of Notch signaling.