ABSTRACT
Memory engrams are a subset of learning activated neurons critical for memory recall, consolidation, extinction and separation. While the transcriptional profile of engrams after learning suggests profound neural changes underlying plasticity and memory formation, little is known about how memory engrams are selected and allocated. As epigenetic factors suppress memory formation, we developed a CRISPR screening in the hippocampus to search for factors controlling engram formation. We identified histone lysine-specific demethylase 4a (Kdm4a) as a negative regulator for engram formation. Kdm4a is downregulated after neural activation and controls the volume of mossy fiber boutons. Mechanistically, Kdm4a anchors to the exonic region of Trpm7 gene loci, causing the stalling of nascent RNAs and allowing burst transcription of Trpm7 upon the dismissal of Kdm4a. Furthermore, the YTH domain containing protein 2 (Ythdc2) recruits Kdm4a to the Trpm7 gene and stabilizes nascent RNAs. Reducing the expression of Kdm4a in the hippocampus via genetic manipulation or artificial neural activation facilitated the ability of pattern separation in rodents. Our work indicates that Kdm4a is a negative regulator of engram formation and suggests a priming state to generate a separate memory.
Subject(s)
Hippocampus , Memory , TRPM Cation Channels , Animals , Hippocampus/metabolism , Mice , Memory/physiology , TRPM Cation Channels/metabolism , TRPM Cation Channels/genetics , Jumonji Domain-Containing Histone Demethylases/metabolism , Jumonji Domain-Containing Histone Demethylases/genetics , Humans , Down-Regulation/genetics , Neurons/metabolism , Male , Mice, Inbred C57BL , Rats , CRISPR-Cas Systems , RNA-Binding Proteins/metabolism , RNA-Binding Proteins/genetics , Neuronal Plasticity/genetics , HEK293 Cells , Histone DemethylasesABSTRACT
BACKGROUND AND PURPOSE: Psoriasis is a common autoimmune skin disease that significantly diminishes patients' quality of life. Interactions between primary afferents of the somatosensory system and the cutaneous immune system mediate the pathogenesis of psoriasis. This study aims to elucidate the molecular mechanisms of how primary sensory neurons regulate psoriasis formation. EXPERIMENTAL APPROACH: Skin and total RNA were extracted from wild-type (WT) and ASH1-like histone lysine methyltransferase (Ash1l+/- ) mice in both naive and imiquimod (IMQ)-induced psoriasis models. Immunohistochemistry, quantitative real-time polymerase chain reaction (qRT-PCR) and fluorescence-activated cell sorting (FACS) were then performed. Microfluidic chamber coculture was used to investigate the interaction between somatosensory neurons and bone marrow dendritic cells (BMDCs) ex vivo. Whole-cell patch clamp recordings were used to evaluate neuronal excitability after Ash1L haploinsufficiency in primary sensory neurons. KEY RESULTS: The haploinsufficiency of ASH1L, a histone methyltransferase, in primary sensory neurons causes both neurite hyperinnervation and increased neuronal excitability, which promote miR-let-7b release from primary afferents in the skin in a neuronal activity-dependent manner. With a 'GUUGUGU' core sequence, miR-let-7b functions as an endogenous ligand of toll-like receptor 7 (TLR7) and stimulates the activation of dermal dendritic cells (DCs) and interleukin (IL)-23/IL-17 axis, ultimately exacerbating the symptoms of psoriasis. Thus, by limiting miR-let-7b release from primary afferents, ASH1L prevents dermal DC activation and ameliorates psoriasis. CONCLUSION AND IMPLICATIONS: Somatosensory neuron ASH1L modulates the cutaneous immune system by limiting neuronal activity-dependent release of miR-let-7b, which can directly activate dermal DCs via TLR7 and ultimately lead to aggravated psoriatic lesion.
Subject(s)
MicroRNAs , Psoriasis , Humans , Animals , Mice , Toll-Like Receptor 7/genetics , Quality of Life , Psoriasis/etiology , Psoriasis/pathology , Skin/pathology , MicroRNAs/genetics , Neurons/pathology , Disease Models, Animal , DNA-Binding Proteins , Histone-Lysine N-MethyltransferaseABSTRACT
When the thermoplastic composites reach the service limits during the service, the recovery and utilization are the key concerns. Meanwhile, the improvement of strength, toughness and durability of epoxy resin is the effective method to prolong the service life of materials and structures. In the present paper, three kinds of thermoplastic resins (polypropylene-PP, polyamide 6-PA6 and polyether-ether-ketone-PEEK) and composites (carbon fiber-PEEK, glass fiber-PA6 and glass fiber-PP) were adopted as the fillers to reinforce and toughen the epoxy resin (Ts). The mechanical, thermal and microscopic analysis were conducted to reveal the performance improvement mechanism of Ts. It can be found that adding thermoplastic resin and composite fillers at the low mass ratio of 0.5~1.0% brought about the maximum improvement of tensile strength (7~15%), flexural strength (7~15%) and shear strength (20~30%) of Ts resin. The improvement mechanism was because the addition of thermoplastic fillers can prolong the cracking path and delay the failure process through the load bearing of fiber, energy absorption of thermoplastic resin and superior interface bonding. In addition, the thermoplastic composite had better enhancement effect on the mechanical/thermal properties of Ts resin compared to thermoplastic resin. When the mass ratio was increased to 2.0~3.0%, the agglomeration and stress concentration of thermoplastic filler in Ts resin appeared, leading to the decrease of mechanical and thermal properties. The optimal addition ratios of thermoplastic resin were 0.5~1.0% (PEEK), 1.0~2.0% (PA6) and 0.5~1.0% (PP) to obtain the desirable property improvement. In contrast, the optimal mass ratios of three kinds of composite were determined to be 0.5~1.0%. Application prospect analysis indicated adding the thermoplastic resin and composite fillers to Ts resin can promote the recycling and reutilization of thermoplastic composites and improve the performance of Ts resin, which can be used as the resin matrix, interface adhesive and anti-corrosion coating.
ABSTRACT
ASD-associated genes are enriched for synaptic proteins and epigenetic regulators. How those chromatin modulators establish ASD traits have remained unknown. We find haploinsufficiency of Ash1l causally induces anxiety and autistic-like behavior, including repetitive behavior, and alters social behavior. Specific depletion of Ash1l in forebrain induces similar ASD-associated behavioral defects. While the learning ability remains intact, the discrimination ability of Ash1l mutant mice is reduced. Mechanistically, deletion of Ash1l in neurons induces excessive synapses due to the synapse pruning deficits, especially during the post-learning period. Dysregulation of synaptic genes is detected in Ash1l mutant brain. Specifically, Eph receptor A7 is downregulated in Ash1l+/- mice through accumulating EZH2-mediated H3K27me3 in its gene body. Importantly, increasing activation of EphA7 in Ash1l+/- mice by supplying its ligand, ephrin-A5, strongly promotes synapse pruning and rescues discrimination deficits. Our results suggest that Ash1l haploinsufficiency is a highly penetrant risk factor for ASD, resulting from synapse pruning deficits.
Subject(s)
Autism Spectrum Disorder , Autistic Disorder , Animals , Autism Spectrum Disorder/genetics , Autism Spectrum Disorder/metabolism , Autistic Disorder/genetics , DNA-Binding Proteins/genetics , Disease Models, Animal , Haploinsufficiency , Histone-Lysine N-Methyltransferase/genetics , Mice , Mice, Knockout , Phenotype , Receptor, EphA1ABSTRACT
BACKGROUND: Immediate-early genes (IEGs) have been serving as markers of active neurons for their rapid responses to stimulation. With the development of IEG-EGFP reporters by the GENSAT project, application of the IEGs have been greatly expanded. However, detailed validations for these systems are still lacking, causing trouble in the interpretation of the fluorescence signals. NEW METHOD: In this work, taken Egr1-EGFP transgenic mice as an example, we proposed an improvement for the usage of the Egr1-EGFP reporter system based on detailed validation of its fluorescence signals. RESULTS: Firstly, the exogenous EGFP mRNA levels were linearly correlated with the endogenous Egr1 mRNA levels in neurons. Secondly, the 3-hr-changes of the Egr1-EGFP signals before and after the stimulus were positively correlated with the stimulus-induced neuronal activities. Interestingly, persistent neuronal activity patterns in the post-stimulus phase also showed correlation with the stimulus-induced Egr1-EGFP signal changes. Furthermore, enriched environments engaged dramatic neuronal activations, allowing detailed characterization of Egr1-EGFP expression dynamics. COMPARISON WITH EXISTING METHOD(S): People used to infer the neuronal activities based on the raw fluorescence signals of IEG-EGFP reporter system, which was strongly obstructed by distinct protein regulation or dynamic properties between the EGFP and the IEGs. We demonstrated a better way for data analysis and experimental design. CONCLUSIONS: Taken together, this work proves that Egr1-EGFP signal is weakly but significantly correlated to task-induced neural activity and gives detailed characterization of the signal dynamics. It not only provides basis for the understanding of the IEG-EGFP fluorescence signals but also offers instructions for proper experimental design with IEG-EGFP reporter systems.
Subject(s)
Genes, Immediate-Early , Neurons , Animals , Early Growth Response Protein 1/genetics , Green Fluorescent Proteins , Mice , Mice, Transgenic , RNA, MessengerABSTRACT
Tourette syndrome (TS) is a childhood-onset neuropsychiatric disorder characterized by repetitive motor movements and vocal tics. The clinical manifestations of TS are complex and often overlap with other neuropsychiatric disorders. TS is highly heritable; however, the underlying genetic basis and molecular and neuronal mechanisms of TS remain largely unknown. We performed whole-exome sequencing of a hundred trios (probands and their parents) with detailed records of their clinical presentations and identified a risk gene, ASH1L, that was both de novo mutated and associated with TS based on a transmission disequilibrium test. As a replication, we performed follow-up targeted sequencing of ASH1L in additional 524 unrelated TS samples and replicated the association (P value = 0.001). The point mutations in ASH1L cause defects in its enzymatic activity. Therefore, we established a transgenic mouse line and performed an array of anatomical, behavioral, and functional assays to investigate ASH1L function. The Ash1l+/- mice manifested tic-like behaviors and compulsive behaviors that could be rescued by the tic-relieving drug haloperidol. We also found that Ash1l disruption leads to hyper-activation and elevated dopamine-releasing events in the dorsal striatum, all of which could explain the neural mechanisms for the behavioral abnormalities in mice. Taken together, our results provide compelling evidence that ASH1L is a TS risk gene.