The Citron homology domain of MAP4Ks improves outcomes of traumatic brain injury.

The mitogen-activated protein kinase kinase kinase kinases (MAP4Ks) signaling pathway plays a pivotal role in axonal regrowth and neuronal degeneration following insults. Whether targeting this pathway is beneficial to brain injury remains unclear. In this study, we showed that adeno-associated virus-delivery of the Citron homology domain of MAP4Ks effectively reduces traumatic brain injury-induced reactive gliosis, tauopathy, lesion size, and behavioral deficits. Pharmacological inhibition of MAP4Ks replicated the ameliorative effects observed with expression of the Citron homology domain. Mechanistically, the Citron homology domain acted as a dominant-negative mutant, impeding MAP4K-mediated phosphorylation of the dishevelled proteins and thereby controlling the Wnt/ß-catenin pathway. These findings implicate a therapeutic potential of targeting MAP4Ks to alleviate the detrimental effects of traumatic brain injury.

Simple and highly specific targeting of resident microglia with adeno-associated virus.

Microglia, as the immune cells of the central nervous system (CNS), play dynamic roles in both healthy and diseased conditions. The ability to genetically target microglia using viruses is crucial for understanding their functions and advancing microglia-based treatments. We here show that resident microglia can be simply and specifically targeted using adeno-associated virus (AAV) vectors containing a 466-bp DNA fragment from the human IBA1 (hIBA1) promoter. This targeting approach is applicable to both resting and reactive microglia. When combining the short hIBA1 promoter with the target sequence of miR124, up to 98% of transduced cells are identified as microglia. Such a simple and highly specific microglia-targeting strategy may be further optimized for research and therapeutics.

Simple and Highly Specific Targeting of Resident Microglia with Adeno-Associated Virus.

Microglia, as the immune cells of the central nervous system (CNS), play dynamic roles in both health and diseased conditions. The ability to genetically target microglia using viruses is crucial for understanding their functions and advancing microglia-based treatments. We here show that resident microglia can be simply and specifically targeted using adeno-associated virus (AAV) vectors containing a 466-bp DNA fragment from the human IBA1 (hIBA1) promoter. This targeting approach is applicable to both resting and reactive microglia. When combining the short hIBA1 promoter with the target sequence of miR124, up to 95% of transduced cells are identified as microglia. Such a simple and highly specific microglia-targeting strategy may be further optimized for research and therapeutics.

NEK6 is an injury-responsive kinase cooperating with STAT3 in regulation of reactive astrogliosis.

In response to brain injury, resident astrocytes become reactive and play dynamic roles in neural repair and regeneration. The signaling pathways underlying such reactive astrogliosis remain largely unclear. We here show that NEK6, a NIMA-related serine/threonine protein kinase, is rapidly induced following pathological stimulations and plays critical roles in reactive astrogliosis. Enhanced NEK6 expression promotes reactive astrogliosis and exacerbates brain lesions; and conversely, NEK6 downregulation dampens injury-induced astrocyte reactivity and reduces lesion size. Mechanistically, NEK6 associates with and phosphorylates STAT3. Kinase activity of NEK6 is required for induction of GFAP and PCNA, markers of reactive astrogliosis. Interestingly, NEK6 is also localized in the nucleus and binds to STAT3-responsive genomic elements in astrocytes. These results indicate that NEK6 constitutes a molecular target for the regulation of reactive astrogliosis.

H2A.Z.1 crosstalk with H3K56-acetylation controls gliogenesis through the transcription of folate receptor.

Astrocytes play crucial roles in the central nervous system, and defects in astrocyte function are closely related to many neurological disorders. Studying the mechanism of gliogenesis has important implications for understanding and treating brain diseases. Epigenetic regulations have essential roles during mammalian brain development. Here, we demonstrate that histone H2A.Z.1 is necessary for the specification of multiple neural precursor cells (NPCs) and has specialized functions that regulate gliogenesis. Depletion of H2A.Z.1 suppresses gliogenesis and results in reduced astrocyte differentiation. Additionally, H2A.Z.1 regulates the acetylation of H3K56 (H3K56ac) by cooperating with the chaperone of ASF1a. Furthermore, RNA-seq data indicate that folate receptor 1 (FOLR1) participates in gliogenesis through the JAK-STAT signaling pathway. Taken together, our results demonstrate that H2A.Z.1 is a key regulator of gliogenesis because it interacts with ASF1a to regulate H3K56ac and then directly affects the expression of FOLR1, which acts as a signal-transducing component of the JAK-STAT signaling pathway.

Brain-specific deletion of histone variant H2A.z results in cortical neurogenesis defects and neurodevelopmental disorder.

Defects in neurogenesis alter brain circuit formations and may lead to neurodevelopmental disorders such as autism and schizophrenia. Histone H2A.z, a variant of histone H2A, plays critical roles in chromatin structure and epigenetic regulation, but its function and mechanism in brain development remain largely unknown. Here, we find that the deletion of H2A.z results in enhanced proliferation of neural progenitors but reduced neuronal differentiation. In addition, neurons in H2A.z knockout mice exhibit abnormal dendrites during brain development. Furthermore, H2A.zcKO mice exhibit serial behavioral deficits, such as decreased exploratory activity and impaired learning and memory. Mechanistically, H2A.z regulates embryonic neurogenesis by targeting Nkx2-4 through interaction with Setd2, thereby promoting H3K36me3 modification to activate the transcription of Nkx2-4. Furthermore, enforced expression of Nkx2-4 can rescue the defective neurogenesis in the H2A.z-knockdown embryonic brain. Together, our findings implicate the epigenetic regulation by H2A.z in embryonic neurogenesis and provide a framework for understanding how disruption in the H2A.z gene may contribute to neurological disorders.

UCP2 Regulates Embryonic Neurogenesis via ROS-Mediated Yap Alternation in the Developing Neocortex.

Mitochondrial metabolism is a fundamental process in tissue development. How this process play functions in embryonic neurogenesis remains largely unknown. Here, we show that mitochondrial uncoupling protein 2 (UCP2) regulates the embryonic neurogenesis by inhibiting the production of reactive oxygen species (ROS), which affect the proliferation of progenitors. In the embryonic brains of UCP2 knockdown or condition knockout mice, the proliferation of progenitors is significantly increased, while the differentiation of progenitors is reduced. Furthermore, we identify that Yap is the response protein of UCP2-mediated ROS production. When UCP2 is inactive, the production of ROS is increased. The amount of Yap protein is increased as Yap degradation through ubiquitin-proteasome proteolytic pathway is decreased. The defect caused by UCP2 depression can be rescued by Yap downregulation. Collectively, our results demonstrate that UCP2 regulates embryonic neurogenesis through ROS-mediated Yap alternation, thus shedding new sight on mitochondrial metabolism involved in embryonic neurogenesis. Stem Cells 2017;35:1479-1492.

DISC1 regulates astrogenesis in the embryonic brain via modulation of RAS/MEK/ERK signaling through RASSF7.

Disrupted in schizophrenia 1 (DISC1) is known as a high susceptibility gene for schizophrenia. Recent studies have indicated that schizophrenia might be caused by glia defects and dysfunction. However, there is no direct evidence of a link between the schizophrenia gene DISC1 and gliogenesis defects. Thus, an investigation into the involvement of DISC1 (a ubiquitously expressed brain protein) in astrogenesis during the late stage of mouse embryonic brain development is warranted. Here, we show that suppression of DISC1 expression represses astrogenesis in vitro and in vivo, and that DISC1 overexpression substantially enhances the process. Furthermore, mouse and human DISC1 overexpression rescued the astrogenesis defects caused by DISC1 knockdown. Mechanistically, DISC1 activates the RAS/MEK/ERK signaling pathway via direct association with RASSF7. Also, the pERK complex undergoes nuclear translocation and influences the expression of genes related to astrogenesis. In summary, our results demonstrate that DISC1 regulates astrogenesis by modulating RAS/MEK/ERK signaling via RASSF7 and provide a framework for understanding how DISC1 dysfunction might lead to neuropsychiatric diseases.

CHD2 is Required for Embryonic Neurogenesis in the Developing Cerebral Cortex.

Chromodomain helicase DNA-binding protein 2 (CHD2) has been associated with a broad spectrum of neurodevelopmental disorders, such as autism spectrum disorders and intellectual disability. However, it is largely unknown whether and how CHD2 is involved in brain development. Here, we demonstrate that CHD2 is predominantly expressed in Pax6(+) radial glial cells (RGs) but rarely expressed in Tbr2(+) intermediate progenitors (IPs). Importantly, the suppression of CHD2 expression inhibits the self-renewal of RGs and increases the generation of IPs and the production of neurons. CHD2 mediates these functions by directly binding to the genomic region of repressor element 1-silencing transcription factor (REST), thereby regulating the expression of REST. Furthermore, the overexpression of REST rescues the defect in neurogenesis caused by CHD2 knockdown. Taken together, these findings demonstrate an essential role of CHD2 in the maintenance of the RGs self-renewal levels, the subsequent generation of IPs, and neuronal output during neurogenesis in cerebral cortical development, suggesting that inactivation of CHD2 during neurogenesis might contribute to abnormal neurodevelopment.

Retinoic acid receptor γ (Rarg) and nuclear receptor subfamily 5, group A, member 2 (Nr5a2) promote conversion of fibroblasts to functional neurons.

Somatic cells can be reprogrammed to neurons and various other cell types with retrovirus or lentivirus. The limitation of this technology is that these genome-integration viruses may increase the risk of gene mutation and cause insertional mutagenesis. We recently found that non-integration adenovirus carrying neuronal transcription factors can induce fibroblasts to neurons. However, the conversion efficiency by the adenovirus is lower than that of the retrovirus or lentivirus. Therefore, it is crucial to identify other factors or chemical compounds to obtain neurons with high efficiency. In this study we show that the combination of Rarg (retinoic acid receptor γ) and Nr5a2 (nuclear receptor subfamily 5, group A, member 2; also known as Lrh-1 (liver receptor homologue 1)) rapidly promote the iN cell maturation within 1 week and greatly facilitate the conversion with neuronal purities of ∼50% and yields of >130%. They also improve neuronal pattern formation, electrophysiological characteristics, and functional integration in vivo. Moreover, the chemical compound agonists to Rarg and Nr5a2 function effectively as well. This approach may be used for the generation and application of iN cells in regenerative medicine.