Synergistic binding of transcription factors to cell-specific enhancers programs motor neuron identity

Abstract

Efficient transcriptional programming promises to open new frontiers in regenerative medicine. However, mechanisms by which programming factors transform cell fate are unknown, preventing more rational selection of factors to generate desirable cell types. Three transcription factors, Ngn2, Isl1 and Lhx3, were sufficient to program rapidly and efficiently spinal motor neuron identity when expressed in differentiating mouse embryonic stem cells. Replacement of Lhx3 by Phox2a led to specification of cranial, rather than spinal, motor neurons. Chromatin immunoprecipitation–sequencing analysis of Isl1, Lhx3 and Phox2a binding sites revealed that the two cell fates were programmed by the recruitment of Isl1-Lhx3 and Isl1-Phox2a complexes to distinct genomic locations characterized by a unique grammar of homeodomain binding motifs. Our findings suggest that synergistic interactions among transcription factors determine the specificity of their recruitment to cell type–specific binding sites and illustrate how a single transcription factor can be repurposed to program different cell types.

Islet 1 specifies the identity of hypothalamic melanocortin neurons and is critical for normal food intake and adiposity in adulthood

“…This and our conditional knockout results point to ISL1 being a terminal differentiation gene that is necessary to establish the specific identity of POMC neurons after these cells are born and accumulate in the mantle zone, giving rise to the arcuate nucleus. Recent work has shown that Isl1 plays pivotal roles in determining the cell fate of GABAergic medium spiny striatofugal neurons (40), cholinergic magnocellular striatal interneurons (41), and cholinergic spinal and cranial motor neurons (42). Interestingly, these different ISL1-dependent neuronal types participate in pyramidal and extrapyramidal circuits that coordinate body movement and locomotion.…”

Identification of STAM1 as a novel effector of ventral projection of spinal motor neurons

“…To identify novel regulators of motor axon pathfinding that function downstream of the ISL1-LHX3 complex, we analyzed genome-wide ISL1-LHX3-bound genomic loci mapped by ChIP-seq analyses (Lee et al, 2013;Mazzoni et al, 2013). Bioinformatics analysis of these ChIP-seq datasets led us to identify a series of putative direct MN target genes of the ISL1-LHX3 complex (data not shown).…”

“…To understand better the role of the ISL1-LHX3 complex in spinal MN specification and differentiation, a genome-wide mapping of ISL1-LHX3 complexbinding sites has been recently performed using chromatin immunoprecipitation sequencing (ChIP-seq) (Lee et al, 2013;Mazzoni et al, 2013). These studies, combined with MN transcriptome analyses (Lee et al, 2012;Mazzoni et al, 2013), revealed that the ISL1-LHX3 complex directly regulates the expression of a wide range of downstream effector genes that function not only in cell fate specification but also in subsequent maturation steps of MN differentiation, such as cell migration, axon pathfinding and synapse formation. As a result, the roles of individual target genes of the ISL1-LHX3 complex in MN specification and differentiation/maturation have finally begun to be dissected (Cho et al, 2014;Lee et al, 2012Lee et al, , 2013Mazzoni et al, 2013).…”

“…These studies, combined with MN transcriptome analyses (Lee et al, 2012;Mazzoni et al, 2013), revealed that the ISL1-LHX3 complex directly regulates the expression of a wide range of downstream effector genes that function not only in cell fate specification but also in subsequent maturation steps of MN differentiation, such as cell migration, axon pathfinding and synapse formation. As a result, the roles of individual target genes of the ISL1-LHX3 complex in MN specification and differentiation/maturation have finally begun to be dissected (Cho et al, 2014;Lee et al, 2012Lee et al, , 2013Mazzoni et al, 2013). Interestingly, v-MNs in embryos deficient in both LHX3 and LHX4 reorients their axonal projections dorsally, whereas the misexpression of LHX3 in d-MNs reorients their axonal projections ventrally (Sharma et al, 1998).…”

Zrf1 controls mesoderm lineage genes and cardiomyocyte differentiation

“…ChIPs were carried out as previously described 57 with the following modifications: A) Differentiated EBs were collected, washed with PBS and then dissociated with Accutase (Gibco) enzyme for 3 minutes. After spin down, single-cell suspension were fixed in fixation buffer (11% freshly added formaldehyde / 0.1M NaCl/1mM EDTA/50mM HEPES-KOH pH 7.6) containing 1% formaldehyde for 15 min at room temperature and quenched with 0.125 M Glycine for additional 5 min at room temperature.…”