In the developing cerebral cortex, sequential transcriptional programs take neuroepithelial cells from proliferating progenitors to differentiated neurons with unique molecular identities. The regulatory changes that occur in the chromatin of the progenitors are not well understood. During deep layer neurogenesis, we show that transcription factor Lhx2 binds to distal regulatory elements of Fezf2 and Sox11, critical determinants of neuron subtype identity in the mouse neocortex. We demonstrate that Lhx2 binds to the NuRD histone remodeling complex subunits LSD1, HDAC2, and RBBP4, which are proximal regulators of the epigenetic state of chromatin. When Lhx2 is absent, active histone marks at the Fezf2 and Sox11 loci are increased. Loss of Lhx2 produces an increase, and overexpression of Lhx2 causes a decrease, in layer 5 Fezf2 and Ctip2 expressing neurons. Our results provide mechanistic insight into how Lhx2 acts as a necessary and sufficient regulator of genes that control cortical neuronal subtype identity. SIGNIFICANCE STATEMENT: The functional complexity of the cerebral cortex arises from an array of distinct neuronal subtypes with unique connectivity patterns that are produced from common progenitors. This study reveals that transcription factor Lhx2 regulates the numbers of specific cortical output neuron subtypes by controlling the genes that are required to produce them. Loss or increase in Lhx2 during neurogenesis is sufficient to increase or decrease, respectively, a particular sub-cerebrally projecting population. Mechanistically, Lhx2 interacts with chromatin modifying protein complexes to edit the chromatin landscape of its targets Fezf2 and Sox11, which regulates their expression and consequently the identities of the neurons produced. Thus Lhx2 is a key component of the control network for producing neurons that will participate in cortical circuitry.
BCL11A, a repressor of human fetal (γ-)globin expression, is required for immune and hematopoietic stem cell functions and brain development. Regulatory sequences within the gene, which are subject to genetic variation affecting fetal globin expression, display hallmarks of an erythroid enhancer in cell lines and transgenic mice. As such this enhancer is a novel, attractive target for therapeutic gene editing. To explore the roles of such sequences in vivo, we generated mice in which the orthologous 10 kb intronic sequences were removed. Bcl11a-enhancer deleted mice (Bcl11a(Δenh)) phenocopy the BCL11A-null state with respect to alterations of globin expression, yet are viable and exhibit no observable blood, brain, or other abnormalities. These preclinical findings provide strong in vivo support for genetic modification of the enhancer for therapy of hemoglobin disorders.
The neocortex contains hundreds to thousands of distinct subtypes of precisely connected neurons, allowing it to perform remarkably complex tasks of high-level cognition. Callosal projection neurons (CPN) connect the cerebral hemispheres via the corpus callosum, integrating cortical information and playing key roles in associative cognition. CPN are a strikingly diverse set of neuronal subpopulations, and development of this diversity requires precise control by a complex, interactive set of molecular effectors. We have found that the transcriptional coregulator Cited2 regulates and refines two stages of CPN development. Cited2 is expressed broadly by progenitors in the embryonic day 15.5 subventricular zone, during the peak of superficial layer CPN birth, with a progressive postmitotic refinement in expression, becoming restricted to CPN of the somatosensory cortex postnatally. We generated progenitor-stage and postmitotic forebrain-specific Cited2 conditional knock-out mice, using the Emx1-Cre and NEX-Cre mouse lines, respectively. We demonstrate that Cited2 functions in progenitors, but is not necessary postmitotically, to regulate both (1) broad generation of layer II/III CPN and (2) acquisition of precise area-specific molecular identity and axonal/dendritic connectivity of somatosensory CPN. This novel CPN subtype-specific and area-specific control from progenitor action of Cited2 adds yet another layer of complexity to the multistage developmental regulation of neocortical development. SIGNIFICANCE STATEMENT: This study identifies Cited2 as a novel subtype-specific and area-specific control over development of distinct subpopulations within the broad population of callosal projection neurons (CPN), whose axons connect the two cerebral hemispheres via the corpus callosum (CC). Currently, how the remarkable diversity of CPN subtypes is specified, and how they differentiate to form highly precise and specific circuits, are largely unknown. We found that Cited2 functions within subventricular zone progenitors to both broadly regulate generation of superficial layer CPN throughout the neocortex, and to refine precise area-specific development and connectivity of somatosensory CPN. Gaining insight into molecular development and heterogeneity of CPN will advance understanding of both diverse functions of CPN and of the remarkable range of neurodevelopmental deficits correlated with CPN/CC development.
Corticothalamic projection neurons (CThPN) are a diverse set of neurons, critical for function of the neocortex. CThPN development and diversity need to be precisely regulated, but little is known about molecular controls over their differentiation and functional specialization, critically limiting understanding of cortical development and complexity. We report the identification of a set of genes that both define CThPN and likely control their differentiation, diversity, and function. We selected the CThPN-specific transcriptional coregulator Fog2 for functional analysis. We identify that Fog2 controls CThPN molecular differentiation, axonal targeting, and diversity, in part by regulating the expression level of Ctip2 by CThPN, via combinatorial interactions with other molecular controls. Loss of Fog2 specifically disrupts differentiation of subsets of CThPN specialized in motor function, indicating that Fog2 coordinates subtype and functional-area differentiation. These results confirm that we identified key controls over CThPN development and identify Fog2 as a critical control over CThPN diversity.