miR-744 and miR-224 Downregulate Npas4 and Affect Lineage Differentiation Potential and Neurite Development During Neural Differentiation of Mouse Embryonic Stem Cells

Abstract

Neuronal PAS domain protein 4 (Npas4) is a brain-specific transcription factor whose expression is enriched in neurogenic regions of the brain. In addition, it was demonstrated that Npas4 expression is dynamic and highly regulated during neural differentiation of embryonic stem cells (ESCs). While these findings implicate a role for Npas4 in neurogenesis, the underlying mechanisms of regulation remain unknown. Given that growing evidence suggests that microRNAs (miRNAs) play important roles in both embryonic and adult neurogenesis, we reasoned that miRNAs are good candidates for regulating Npas4 expression during neural differentiation of ESCs. In this study, we utilized the small RNA sequencing method to profile miRNA expression during neural differentiation of mouse ESCs. Two differentially expressed miRNAs were identified to be able to significantly reduce reporter gene activity by targeting the Npas4 3'UTR, namely miR-744 and miR-224. More importantly, ectopic expression of these miRNAs during neural differentiation resulted in downregulation of endogenous Npas4 expression. Subsequent functional analysis revealed that overexpression of either miR-744 or miR-224 delayed early neural differentiation, reduced GABAergic neuron production and inhibited neurite outgrowth. Collectively, our findings indicate that Npas4 not only functions at the early stages of neural differentiation but may also, in part, contribute to neuronal subtype specification and neurite development.

MicroRNA-744 Inhibits Proliferation of Bronchial Epithelial Cells by Regulating Smad3 Pathway via Targeting Transforming Growth Factor-β1 (TGF-β1) in Severe Asthma

“…Here, we found miR-744 was lower in severe asthma, showing great promise for treatment of severe asthma. miR-744 has been reported to play essential roles in renal inflammation [27], neural differentiation [28], and cell proliferation [29]. Importantly, we showed that miR-744 limited the proliferation rate of bronchial epithelial cells in severe asthma.…”

GABAergic neurons maturation is regulated by a delicate network

“…Due to their extraordinary neurochemical and functional diversity, GABAergic neurons are involved in various neuropsychiatric disorders, such as epilepsy, anxiety, autism, and other pathological processes, such as cerebral ischemia injury and drug addiction. Neurogenesis requires precisely regulated patterns of gene expression in which positive and negative signals must be balanced to generate the correct cell types at the appropriate developmental period (Choy et al, 2017). The development and differentiation of GABAergic neurons are complex processes that involve the interaction of a series of genes that regulate each other in an orderly manner.…”

“…Npas4 is an inducible transcription factor (ITF) that is expressed exclusively in neurons and is selectively activated by neuronal activity rather than by extracellular stimuli, such as growth factors and neurotrophic factors. Npas4 belongs to the bHLH PAS protein family and is an important target for regulating stress responses (Choy et al, 2017; Moser et al, 2004). As a transcription factor, Npas4 exerts its biological function by controlling the expression of downstream transcription targets.…”

“…Brain‐derived neurotrophic factor (BDNF) is the main downstream target of Npas4 in regulating the synaptic development of GABA neurons. In excitatory neurons, Npas4 activates BDNF transcription, thereby promoting an increase in the number of inhibitory synapses (Choy et al, 2017; Lin et al, 2008). IQ motif and SEC7 domain‐containing protein 3 (IQSEC3) is an ADP ribosylation factor‐guanine nucleotide exchange factor (ARF‐GEF) that directly interacts with gephyrin and is restricted to GABAergic synapses.…”

Transcriptome sequencing of ceRNA network constructing in status epilepticus mice treated by low-frequency repetitive transcranial magnetic stimulation

“…Furthermore, mir-186 controls the surface expression of GluA2 (ionic glutamate receptor A2) in hippocampal neurons (Silva MM et al, 2019). Down-regulated miR-744 could downregulate Npas4 as an intrinsic regulator of seizures and epilepsy (Choy FC et al, 2017;Shan W et al, 2018). Together, these alterations represent an intrinsic therapeutic mechanism of LF-rTMS for epilepsy.…”