Regulation of Hemoglobin Switching

Abstract
One of the oldest and most deeply studied problems in developmental gene expression is the switch from fetal
to adult type hemoglobin in red blood cell precursors. Interest in this question has been fueled by its relevance
to genetic blood disorders such as sickle cell disease and thalassemia, and knowledge about regulatory
processes is being translated into gene therapies and other therapeutic approaches. BCL11A is a critical
player in the globin switch, but how it is regulated developmentally is surprisingly still largely unclear. Our
preliminary data show that BCL11A is controlled predominantly at the transcriptional level. Via a CRISPR-Cas9
genetic screen we identified the transcriptional repressor HIC2 as a novel regulator of hemoglobin switching.
HIC2 is expressed highly in fetal erythroid cells and extinguished in adult erythroid cells. Our preliminary data
further suggest that HIC2 represses BCL11A transcription specifically in fetal type cells by directly
decommissioning a fetal stage-specific BCL11A enhancer element. Together, these observations define the
foundational hypothesis of this application: HIC2 expression is extinguished in adult red cells, allowing for the
activation of a BCL11A enhancer to boost BCL11A expression in adult cells and trigger the silencing of fetal
type globin genes. This places HIC2 upstream of BCL11A in the regulatory circuitry that controls hemoglobin
switching. Additional preliminary data suggest that HIC2 promotes a broader fetal transcriptional program. In
Specific Aim 1 we will examine the biology of HIC2 in gain- and loss-of-function experiments in vivo using a
combination of cell culture models and whole animal studies. Specific Aim 2 is focused on mechanistic
experiments, defining the way by which HIC2 regulates chromatin features and transcription factor binding at
target genes, including BCL11A, and what HIC2 co-factors are involved. Specific Aim 3 is dedicated to
defining the developmental control of HIC2 expression. This will be accomplished by characterizing the
regulatory landscape of the HIC2 locus in fetal and adult erythroblasts in combination with developmental
stage specific perturbative experiments.
In sum, the proposed studies aim to understand the role of HIC2 in hemoglobin switching and in the
establishment of a fetal erythroid state at an organismal and molecular level. This proposal is expected to
produce results with ramifications for a broader understanding of developmental hematopoiesis as well as the
treatment of sickle cell disease and thalassemia.

Grant Number: 5R01HL119479-13
NIH Institute/Center: NIH
Principal Investigator: Gerd Blobel