New Insights Into the Intricacies of Proneural Gene Regulation in the Embryonic and Adult Cerebral Cortex

Abstract

Historically, the mammalian brain was thought to lack stem cells as no new neurons were found to be made in adulthood. That dogma changed ∼25 years ago with the identification of neural stem cells (NSCs) in the adult rodent forebrain. However, unlike rapidly self-renewing mature tissues (e.g., blood, intestinal crypts, skin), the majority of adult NSCs are quiescent, and those that become ‘activated’ are restricted to a few neurogenic zones that repopulate specific brain regions. Conversely, embryonic NSCs are actively proliferating and neurogenic. Investigations into the molecular control of the quiescence-to-proliferation-to-differentiation continuum in the embryonic and adult brain have identified proneural genes encoding basic-helix-loop-helix (bHLH) transcription factors (TFs) as critical regulators. These bHLH TFs initiate genetic programs that remove NSCs from quiescence and drive daughter neural progenitor cells (NPCs) to differentiate into specific neural cell subtypes, thereby contributing to the enormous cellular diversity of the adult brain. However, new insights have revealed that proneural gene activities are context-dependent and tightly regulated. Here we review how proneural bHLH TFs are regulated, with a focus on the murine cerebral cortex, drawing parallels where appropriate to other organisms and neural tissues. We discuss upstream regulatory events, post-translational modifications (phosphorylation, ubiquitinylation), protein–protein interactions, epigenetic and metabolic mechanisms that govern bHLH TF expression, stability, localization, and consequent transactivation of downstream target genes. These tight regulatory controls help to explain paradoxical findings of changes to bHLH activity in different cellular contexts.

The Role of Neurod Genes in Brain Development, Function, and Disease

“…It initiates with the formation of the neural tube and the differentiation and specification of neural progenitor cells that, subsequently, lead to the genesis of differentiated neurons in a process called neurogenesis that culminates in the early postnatal life in humans, but can span throughout the adult life in other species, such as rodents (Altman and Das, 1965 ; Johnson, 2001 ; Bayer and Altman, 2007 ; Stiles and Jernigan, 2010 ; Silbereis et al, 2016 ; Sorrells et al, 2018 ; Buffalo et al, 2019 ; Petrik and Encinas, 2019 ). The specification of neural progenitor cells and their activation to self-renew and/or to differentiate in more committed progenitors and neurons is mediated by extrinsic and intrinsic molecular mechanisms (Götz and Sommer, 2005 ; Urbán and Guillemot, 2014 ; Götz et al, 2016 ; Oproescu et al, 2021 ). The intrinsic mechanisms that direct neural progenitor cell progression and differentiation rely on the coordinated function of multiple transcription factors that determine their identity and, simultaneously, the suppression of their progenitor cell programs (Schuurmans et al, 2004 ; Britz et al, 2006 ; Hevner et al, 2006 ; Davidson, 2010 ; Hodge and Hevner, 2011 ; Busskamp et al, 2014 ; Ware et al, 2016 ; Mall et al, 2017 ; Lee et al, 2019 ).…”

Direct Neuronal Reprogramming: Bridging the Gap Between Basic Science and Clinical Application

“…Nor-adrenergic neurons 41.8% Electrophysiologically active, generate action potentials Li et al, 2019 In 2007, iNs were generated from murine P5-7 cortical astrocytes by expressing one of two proneural bHLH genes: Neurogenin 2 (Neurog2) or Ascl1 (previous gene name, Mash1), and were shown to be functional as they could fire action potentials (Berninger et al, 2007a). In the embryonic forebrain, Neurog2 and Ascl1 promote neurogenesis and specify neuronal subtype identities: Neurog2 specifies an excitatory, glutamatergic neuronal phenotype and Ascl1 specifies an inhibitory, GABAergic neuronal identity (reviewed in Oproescu et al, 2021). Neurog2 and Ascl1 also specify distinct glutamatergic and GABAergic neuronal fates, respectively, when they are overexpressed in multipotent NSCs from the adult mouse brain (Berninger et al, 2007b), although this is an example of induced differentiation and not trans-differentiation, the focus of this review.…”

“…While both proneural TFs rapidly induce neuronal differentiation and alter gene expression, there is only a ∼3% overlap in induced genes, consistent with the observation that these TFs specify distinct neuronal phenotypes in the embryo ( Masserdotti et al, 2015 ). One of the common genes activated by both Neurog2 and Ascl1 is Neurod4 , a proneural bHLH gene with a role in early neural lineage development, which functions downstream of Neurog2 in embryonic cortical progenitors (reviewed in Oproescu et al, 2021 ). Notably, misexpression of Neurod4 in P6-P7 astrocytes could similarly induce neuronal marker expression.…”

“…The most common growth factor added to cell culture media are fibroblast growth factors (FGF2/4/8), epidermal growth factor (EGF) or insulin growth factor (IGF), which support general cell proliferation, along with N2 and/or B27 defined growth supplements that supports neuronal growth and survival. Other growth factors are added because they promote neuronal differentiation, such as Wnts, retinoic acid, sonic hedgehog (SHH), and the sex steroid, progesterone (reviewed in Rivetti di Val Cervo et al, 2017 ; Oproescu et al, 2021 ).…”

“…Notch binds Jag/Dll ligands on adjacent cells, ultimately leading to the downstream transcription of Hes1/Hes5 , bHLH TFs that inhibit proneural gene expression and prevent neuronal differentiation. Repression of Notch signaling is achieved using DAPT (N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester), a gamma secretase inhibitor that prevents cleavage of the Notch receptor so that the Notch intracellular domain (NICD) does not translocate to the nucleus to assist in the transcriptional activation of the downstream Hes1/Hes5 bHLH effectors (reviewed in Oproescu et al, 2021 ).…”