C/EBPα Activates Pre-existing and De Novo Macrophage Enhancers during Induced Pre-B Cell Transdifferentiation and Myelopoiesis

Abstract

SummaryTranscription-factor-induced somatic cell conversions are highly relevant for both basic and clinical research yet their mechanism is not fully understood and it is unclear whether they reflect normal differentiation processes. Here we show that during pre-B-cell-to-macrophage transdifferentiation, C/EBPα binds to two types of myeloid enhancers in B cells: pre-existing enhancers that are bound by PU.1, providing a platform for incoming C/EBPα; and de novo enhancers that are targeted by C/EBPα, acting as a pioneer factor for subsequent binding by PU.1. The order of factor binding dictates the upregulation kinetics of nearby genes. Pre-existing enhancers are broadly active throughout the hematopoietic lineage tree, including B cells. In contrast, de novo enhancers are silent in most cell types except in myeloid cells where they become activated by C/EBP factors. Our data suggest that C/EBPα recapitulates physiological developmental processes by short-circuiting two macrophage enhancer pathways in pre-B cells.

A Transcription Factor Pulse Can Prime Chromatin for Heritable Transcriptional Memory

“…In the B cell-to-macrophage TD model (15), exogenous C/EBP␣ is induced and collaborates with endogenous PU.1 at enhancer elements to activate the macrophage gene expression program (14). Interestingly, a commitment point during the TD process is reached between 18 and 24 h whereby the induced, exogenous C/EBP␣ expression can be removed, but the cells continue to convert toward macrophage specification (15).…”

“…Indeed, binding of these factors can make previously inaccessible chromatin sites transcriptionally competent and aid in the conversion of gene regulatory elements from a latent/off to a primed or active state (12). CCAAT/enhancer-binding protein alpha (C/EBP␣) and AP-1 TF families have been reported to display pioneering factor activity in the myeloid lineage (13,14), while C/EBP␣ overexpression in conjunction with endogenous PU.1 works as a module to mediate complete B cell-to-macrophage transdifferentiation (TD) (15,16). Therefore, we reasoned that the inducible TD system developed previously (15) could be employed to decipher the epigenetic mechanisms contributing to the transcriptional memory phenotype in immune cells.…”

Direct lineage reprogramming via pioneer factors; a detour through developmental gene regulatory networks

“…These low-signal chromatin regions are characterized by their relative lack of histone modifications (Ho et al, 2014;Kharchenko et al, 2011), where transcription is instead likely to be repressed by the presence of linker histones (Iwafuchi-Doi and Zaret, 2016). Pioneer factor binding results in the local opening of this silent chromatin (Soufi et al, 2012;van Oevelen et al, 2015), although in the absence of any other transcription factors this is insufficient to induce changes in gene expression. Rather, pioneer factors recruit activators or repressors that by themselves are unable to engage with silent chromatin (Carroll et al, 2005;Gualdi et al, 1996;Sekiya and Zaret, 2007).…”

“…Gata4 is another well-characterized pioneer factor (Bossard and Zaret, 1998) responsible for driving both cardiac and hepatic lineage conversions. In addition, C/ebpα and PU.1 (Spi1) have been reported to act as pioneer factors (Heinz et al, 2010;van Oevelen et al, 2015), inducing conversions within the blood lineage. In the neural reprogramming field, Ascl1 acts as a pioneer factor and is employed in the majority of conversions to neural fate (Wapinski et al, 2013).…”

C/EBPα overrides epigenetic reprogramming by oncogenic transcription factors in acute myeloid leukemia

“…9,15 This requirement for C/EBPa to activate a myeloid differentiation response reflects the results of previous knockout models showing that this transcription factor is required for hematopoietic cells to transit to granulocytemonocyte precursors (GMPs) 16 and that in its absence, neutrophil development fails. 17 Others have previously shown that C/EBPa overexpression is sufficient to trans-differentiate lymphocytes into myeloid cells [18][19][20] and is able to initiate myeloid differentiation in RUNX1-ETOexpressing cells. 21 Similarly, simultaneous expression of C/EBPa rescued the RUNX1-EVI1-mediated block of a differentiation response in a model cell line.…”