Expression of the primate-specific LINC00473 RNA in mouse neurons promotes excitability and CREB-regulated transcription.

The LINC00473 (Lnc473) gene has previously been shown to be associated with cancer and psychiatric disorders. Its expression is elevated in several types of tumors and decreased in the brains of patients diagnosed with schizophrenia or major depression. In neurons, Lnc473 transcription is strongly responsive to synaptic activity, suggesting a role in adaptive, plasticity-related mechanisms. However, the function of Lnc473 is largely unknown. Here, using a recombinant adeno-associated viral vector, we introduced a primate-specific human Lnc473 RNA into mouse primary neurons. We show that this resulted in a transcriptomic shift comprising downregulation of epilepsy-associated genes and a rise in cAMP response element-binding protein (CREB) activity, which was driven by augmented CREB-regulated transcription coactivator 1 nuclear localization. Moreover, we demonstrate that ectopic Lnc473 expression increased neuronal excitability as well as network excitability. These findings suggest that primates may possess a lineage-specific activity-dependent modulator of CREB-regulated neuronal excitability.

Lineage divergence of activity-driven transcription and evolution of cognitive ability.

Excitation-transcription coupling shapes network formation during brain development and controls neuronal survival, synaptic function and cognitive skills in the adult. New studies have uncovered differences in the transcriptional responses to synaptic activity between humans and mice. These differences are caused both by the emergence of lineage-specific activity-regulated genes and by the acquisition of signal-responsive DNA elements in gene regulatory regions that determine whether a gene can be transcriptionally induced by synaptic activity or alter the extent of its inducibility. Such evolutionary divergence may have contributed to lineage-related advancements in cognitive abilities.

Npas4 regulates medium spiny neuron physiology and gates cocaine-induced hyperlocomotion.

We show here that the transcription factor Npas4 is an important regulator of medium spiny neuron spine density and electrophysiological parameters and that it determines the magnitude of cocaine-induced hyperlocomotion in mice. Npas4 is induced by synaptic stimuli that cause calcium influx, but not dopaminergic or PKA-stimulating input, in mouse medium spiny neurons and human iPSC-derived forebrain organoids. This induction is independent of ubiquitous kinase pathways such as PKA and MAPK cascades, and instead depends on calcineurin and nuclear calcium signalling. Npas4 controls a large regulon containing transcripts for synaptic molecules, such as NMDA receptors and VDCC subunits, and determines in vivo MSN spine density, firing rate, I/O gain function and paired-pulse facilitation. These functions at the molecular and cellular levels control the locomotor response to drugs of abuse, as Npas4 knockdown in the nucleus accumbens decreases hyperlocomotion in response to cocaine in male mice while leaving basal locomotor behaviour unchanged.

The Intellectual Disability and Schizophrenia Associated Transcription Factor TCF4 Is Regulated by Neuronal Activity and Protein Kinase A.

Transcription factor 4 (TCF4 also known as ITF2 or E2-2) is a basic helix-loop-helix (bHLH) protein associated with Pitt-Hopkins syndrome, intellectual disability, and schizophrenia (SCZ). Here, we show that TCF4-dependent transcription in cortical neurons cultured from embryonic rats of both sexes is induced by neuronal activity via soluble adenylyl cyclase and protein kinase A (PKA) signaling. PKA phosphorylates TCF4 directly and a PKA phosphorylation site in TCF4 is necessary for its transcriptional activity in cultured neurons and in the developing brain in vivo We also demonstrate that Gadd45g (growth arrest and DNA damage inducible gamma) is a direct target of neuronal-activity-induced, TCF4-dependent transcriptional regulation and that TCF4 missense variations identified in SCZ patients alter the transcriptional activity of TCF4 in neurons. This study identifies a new role for TCF4 as a neuronal-activity-regulated transcription factor, offering a novel perspective on the association of TCF4 with cognitive disorders.SIGNIFICANCE STATEMENT The importance of the basic helix-loop-helix transcription factor transcription factor 4 (TCF4) in the nervous system is underlined by its association with common and rare cognitive disorders. In the current study, we show that TCF4-controlled transcription in primary cortical neurons is induced by neuronal activity and protein kinase A. Our results support the hypotheses that dysregulation of neuronal-activity-dependent signaling plays a significant part in the etiology of neuropsychiatric and neurodevelopmental disorders.

AP-1 Transcription Factors Mediate BDNF-Positive Feedback Loop in Cortical Neurons.

Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family, regulates both survival and differentiation of several neuronal populations in the nervous system during development, as well as synaptic plasticity in the adult brain. BDNF exerts its biological functions through its receptor TrkB. Although the regulation of BDNF transcription by neuronal activity has been widely studied, little is known about TrkB signaling-dependent expression of BDNF. Using rat primary cortical neuron cultures, we show that the BDNF gene is a subject to an extensive autoregulatory loop, where TrkB signaling upregulates the expression of all major BDNF transcripts, mainly through activating MAPK pathways. Investigating the mechanisms behind this autoregulation, we found that AP-1 transcription factors, comprising Jun and Fos family members, participate in the induction of BDNF exon I, III, and VI transcripts. AP-1 transcription factors directly upregulate the expression of exon I transcripts by binding two novel AP-1 cis-elements in promoter I. Moreover, our results show that the effect of AP-1 proteins on the activity of rat BDNF promoters III and VI is indirect, because AP-1 proteins were not detected to bind the respective promoter regions by chromatin immunoprecipitation (ChIP). Collectively, we describe an extensive positive feedback system in BDNF regulation, adding a new layer to the elaborate control of BDNF gene expression. SIGNIFICANCE STATEMENT: Here, we show for the first time that in rat primary cortical neurons the expression of all major BDNF transcripts (exon I, II, III, IV, VI, and IXa transcripts) is upregulated in response to TrkB signaling, and that AP-1 transcription factors participate in the induction of exon I, III, and VI transcripts. Moreover, we have described two novel functional AP-1 cis-elements in BDNF promoter I, responsible for the activation of the promoter in response to TrkB signaling. Our results indicate the existence of a positive feedback loop for obtaining sufficient BDNF levels necessary for various TrkB signaling-dependent physiological outcomes in neurons.

GLI2 cell-specific activity is controlled at the level of transcription and RNA processing: Consequences to cancer metastasis.

High activity of GLI family zinc finger protein 2 (GLI2) promotes tumor progression. Removal of the repressor domain at the N terminus (GLI2∆N) by recombinant methods converts GLI2 into a powerful transcriptional activator. However, molecular mechanisms leading to the formation of GLI2∆N activator proteins have not been established. Herein we report for the first time that the functional activities of GLI2 are parted into different protein isoforms by alternative promoter usage, selection of alternative splicing, transcription initiation and termination sites. Functional studies using melanoma cells revealed that transcriptional regulation of GLI2 is TGFbeta-dependent and supports the predominant production of GLI2∆N and C-terminally truncated GLI2 (GLI2∆C) isoforms in cells with high migratory and invasive phenotype. Taken together, these results highlight the role of transcription and RNA processing as major processes in the regulation of GLI2 activity with severe impacts in cancer development.

Regulation of different human NFAT isoforms by neuronal activity.

Nuclear factor of activated T-cells (NFAT) is a family of transcription factors comprising four calcium-regulated members: NFATc1, NFATc2, NFATc3, and NFATc4. Upon activation by the calcium-dependent phosphatase calcineurin (CaN), NFATs translocate from cytosol to the nucleus and regulate their target genes, which in the nervous system are involved in axon growth, synaptic plasticity, and neuronal survival. We have shown previously that there are a number of different splice variants of NFAT genes expressed in the brain. Here, we studied the subcellular localizations and transactivation capacities of alternative human NFAT isoforms in rat primary cortical or hippocampal neurons in response to membrane depolarization and compared the induced transactivation levels in neurons to those obtained from HEK293 cells in response to calcium signaling. We confirm that in neurons the translocation to the nucleus of all NFAT isoforms is reliant on the activity of CaN. However, our results suggest that both the regulation of subcellular localization and transcriptional activity of NFAT proteins in neurons is isoform specific. We show that in primary hippocampal neurons NFATc2 isoforms have very fast translocation kinetics, whereas NFATc4 isoforms translocate relatively slowly to the nucleus. Moreover, we demonstrate that the strongest transcriptional activators in HEK293 cells are NFATc1 and NFATc3, but in neurons NFATc3 and NFATc4 lead to the highest induction, and NFATc2 and NFATc1 display isoform-specific transcription activation capacities. Altogether, our results indicate that the effects of calcium signaling on the action of NFAT proteins are isoform-specific and can differ between cell types. We show that the effects of calcium signaling on the action of NFAT proteins are isoform-specific and differ between cell types. Although nuclear localization of all NFAT isoforms in neurons requires calcineurin, the subcellular distributions, neuronal activity-induced nuclear translocation extent and kinetics, and transcription activation capacities of alternative NFAT proteins vary.

Pitt-Hopkins syndrome-associated mutations in TCF4 lead to variable impairment of the transcription factor function ranging from hypomorphic to dominant-negative effects.

Transcription factor TCF4 (alias ITF2, SEF2 or E2-2) is a broadly expressed basic helix-loop-helix (bHLH) protein that functions as a homo- or heterodimer. Missense, nonsense, frame-shift and splice-site mutations as well as translocations and large deletions encompassing TCF4 gene cause Pitt-Hopkins syndrome (PTHS), a rare developmental disorder characterized by severe motor and mental retardation, typical facial features and breathing anomalies. Irrespective of the mutation, TCF4 haploinsufficiency has been proposed as an underlying mechanism for PTHS. We have recently demonstrated that human TCF4 gene is transcribed using numerous 5' exons. Here, we re-evaluated the impact of all the published PTHS-associated mutations, taking into account the diversity of TCF4 isoforms, and assessed how the reading frame elongating and missense mutations affect TCF4 functions. Our analysis revealed that not all deletions and truncating mutations in TCF4 result in complete loss-of-function and the impact of reading frame elongating and missense mutations ranges from subtle deficiencies to dominant-negative effects. We show that (i) missense mutations in TCF4 bHLH domain and the reading frame elongating mutation damage DNA-binding and transactivation ability in a manner dependent on dimer context (homodimer versus heterodimer with ASCL1 or NEUROD2); (ii) the elongating mutation and the missense mutation at the dimer interface of the HLH domain destabilize the protein; and (iii) missense mutations outside of the bHLH domain cause no major functional deficiencies. We conclude that different PTHS-associated mutations impair the functions of TCF4 by diverse mechanisms and to a varying extent, possibly contributing to the phenotypic variability of PTHS patients.

Identification of cis-elements and transcription factors regulating neuronal activity-dependent transcription of human BDNF gene.

Brain-derived neurotrophic factor (BDNF) is an important mediator of activity-dependent functions of the nervous system and its expression is dysregulated in several neuropsychiatric disorders. Regulation of rodent BDNF neuronal activity-dependent transcription has been relatively well characterized. Here, we have studied regulation of human BDNF (hBDNF) transcription by membrane depolarization of cultured mouse or rat primary cortical neurons expressing hBDNF gene or transfected with hBDNF promoter constructs, respectively. We identified an asymmetric E-box-like element, PasRE [basic helix-loop-helix (bHLH)-PAS transcription factor response element], in hBDNF promoter I and demonstrate that binding of this element by bHLH-PAS transcription factors ARNT2 (aryl hydrocarbon receptor nuclear translocator 2) and NPAS4 (neuronal PAS domain protein 4) is crucial for neuronal activity-dependent transcription from promoter I. We show that binding of CREB (cAMP response element-binding protein) to the cAMP/Ca(2+)-response element (CRE) in hBDNF promoter IV is critical for activity-dependent transcription from this promoter and that upstream stimulatory factor (USF) transcription factors also contribute to the activation by binding to the upstream stimulatory factor binding element (UBE) in hBDNF promoter IV. However, we report that full induction of hBDNF exon IV mRNA transcription is dependent on ARNT2 and NPAS4 binding to a PasRE in promoter IV. Finally, we demonstrate that CRE and PasRE elements in hBDNF promoter IX are required for the induction of this promoter by neuronal activity. Together, the results of this study have identified the cis-elements and transcription factors regulating neuronal activity-dependent transcription of human BDNF gene.

Subcellular localization and transcription regulatory potency of KCNIP/Calsenilin/DREAM/KChIP proteins in cultured primary cortical neurons do not provide support for their role in CRE-dependent gene expression.

KCNIP3/KChIP3 (voltage-dependent K+ channel interacting protein 3), alias Calsenilin and downstream regulatory element antagonist modulator (DREAM), is a multifunctional protein that modulates A-type potassium channels, affects processing of amyloid precursor protein and regulates transcription. KCNIP3 has been described to negatively influence the activity of CREB (cAMP/Ca(2+)-response element binding protein), an essential factor in neuronal activity-dependent gene expression regulation. However, reports on intracellular localization of KCNIP3 in neurons are diverse and necessitate additional analyses of distribution of KCNIPs in cells to clarify the potential of KCNIP3 to fulfill its functions in different cell compartments. Here, we examined localization of the entire family of highly similar KCNIP proteins in neuronal cells and show that over-expressed isoforms of KCNIP1/KChIP1, KCNIP2/KChIP2, KCNIP3/KChIP3, and KCNIP4/KChIP4 had varied, yet partially overlapping subcellular localization. In addition, although some of the over-expressed KCNIP isoforms localized to the nucleus, endogenous KCNIPs were not detected in nuclei of rat primary cortical neurons. Moreover, we analyzed the role of KCNIP proteins in cAMP/Ca(2+)-response element (CRE)-dependent transcription by luciferase reporter assay and electrophoretic mobility shift assay and report that our results do not support the role for KCNIPs, including DREAM/Calsenilin/KChIP3, in modulation of CREB-mediated transcription in neurons.

N-methyl-d-aspartate Receptor-mediated Preconditioning Mitigates Excitotoxicity in Human Induced Pluripotent Stem Cell-derived Brain Organoids.

Studies in rodent models of acute and chronic neurodegenerative disorders have uncovered that glutamate-induced excitotoxic cell death is mediated primarily by extrasynaptic N-methyl-d-aspartate receptors (NMDARs). Rodent neurons can also build up in an activity-dependent manner a protective shield against excitotoxicity. This form of acquired neuroprotection is induced by preconditioning with low doses of NMDA or by activation of synaptic NMDARs triggered by bursts of action potentials. Whether NMDARs in human neurons have similar dichotomous actions in cell death and survival is unknown. To investigate this, we established an induced pluripotent stem cell (iPSC)-derived forebrain organoid model for excitotoxic cell death and explored conditions of NMDAR activation that promote neuronal survival when applied prior to a toxic insult. We found that glutamate-induced excitotoxicity in human iPSC-derived neurons is mediated by NMDARs. Treatment of organoids with high concentrations of glutamate or NMDA caused the typical excitotoxicity pathology, comprising structural disintegration, neurite blebbing, shut-off of the transcription factor CRE binding protein (CREB), and cell death. In contrast, bath-applied low doses of NMDA elicited synaptic activity, a robust and sustained increase in CREB phosphorylation as well as function, and upregulation of immediate-early genes, including neuroprotective genes. Moreover, we found that conditions of enhanced synaptic activity increased survival of human iPSC-derived neurons if applied as pre-treatment before toxic NMDA application. These results revealed that both toxic and protective actions of NMDARs are preserved in human neurons. The experimental platform described in this study may prove useful for the validation of neuroprotective gene products and drugs in human neurons.

Bidirectional transcription from human LRRTM2/CTNNA1 and LRRTM1/CTNNA2 gene loci leads to expression of N-terminally truncated CTNNA1 and CTNNA2 isoforms.

α-Catenins (CTNNAs) are essential for the regulation of cell-cell and cell-matrix interactions in tissues. All human CTNNA genes contain antisense oriented leucine rich repeat transmembrane (LRRTM) genes within their seventh introns. Recently, a haplotype upstream of one of the human LRRTM genes, LRRTM1 that resides in CTNNA2, was shown to be associated with handedness and schizophrenia. Here, we show that both CTNNA1 and CTNNA2 contain alternative 5' exons linked to bidirectional promoters that are shared with the antisense oriented LRRTM2 and LRRTM1 genes, respectively. We demonstrate that bidirectional activity of these promoters results in alternative CTNNA1 and CTNNA2 transcripts that are expressed at high levels in the nervous system and show that N-terminally truncated CTNNA1 and CTNNA2 proteins lacking the ß-catenin interaction domain are produced from these alternative CTNNA mRNAs. In addition, our results indicate that the haplotype that affects LRRTM1 expression and is associated with schizophrenia and handedness, could also influence the expression of brain-enriched alternative transcripts of CTNNA2.

Nuclear factor of activated T-cells isoform c4 (NFATc4/NFAT3) as a mediator of antiapoptotic transcription in NMDA receptor-stimulated cortical neurons.

During cortical development, when NR2B subunit is the major component of the NMDA glutamate receptors (NMDARs), moderate NMDAR activity supports neuronal survival at least in part by regulating gene transcription. We report that, in cultured cortical neurons from newborn rats, the NMDARs activated the calcium-responsive transcription regulator nuclear factor of activated T cells (NFAT). Moreover, in developing rat cortex, the NFAT isoforms c3 and c4 (NFATc3 and NFATc4) were expressed at relatively higher levels at postnatal day 7 (P7) than P21, overlapping with the period of NMDAR-dependent survival. In cultured cortical neurons, NFATc3 and NFATc4 were regulated at least in part by the NR2B NMDAR. Conversely, knockdown of NFATc4 but not NFATc3 induced cortical neuron apoptosis. Likewise, NFATc4 inhibition prevented antiapoptotic neuroprotection in response to exogenous NMDA. Expression of the brain-derived neurotrophic factor (BDNF) was reduced by NFATc4 inhibition. NFATc4 regulated transcription by the NMDAR-responsive bdnf promoter IV. In addition, NMDAR blockers including NR2B-selective once reduced BDNF expression in P7 cortex and cultured cortical neurons. Finally, exogenous BDNF rescued from the proapoptotic effects of NFATc4 inhibition. These results identify bdnf as one of the target genes for the antiapoptotic signaling by NMDAR-NFATc4. Thus, the previously unrecognized NMDAR-NFATc4-BDNF pathway contributes to the survival signaling network that supports cortical development.

N-terminally truncated BAF57 isoforms contribute to the diversity of SWI/SNF complexes in neurons.

The SWItch/Sucrose NonFermentable, a nucleosome remodeling complex (SWI/SNF) chromatin-remodelling complexes act upon the nucleosomal structure and regulate transcription, replication, repair of chromatin and splicing. In this study, we present evidence that human, mouse and rat genes encoding one of the SWI/SNF complex subunits, BAF57, undergo neuron-specific splicing of exons II, III and IV. Alternative splicing yields in at least three isoforms of BAF57 protein that have truncated N-termini (N-BAF57s). The transcripts encoding N-BAF57 isoforms are predominantly expressed in the nervous system. The biochemical fractionation data supported by the results of the co-immunoprecipitation analysis show that N-BAF57 isoforms associate into protein complexes together with Brg1, Brm, BAF155 and BAF170. Transient over-expression of N-BAF57 isoforms in non-neural cells affects the level of expression of certain neuron-restrictive silencer element-containing genes. Together these data suggest that neuronal isoforms of BAF57 contribute to functional SWI/SNF complexes regulating neurogenesis.

NF-kappaB-dependent regulation of brain-derived neurotrophic factor in hippocampal neurons by X-linked inhibitor of apoptosis protein.

X chromosome-linked inhibitor of apoptosis protein (XIAP) is an anti-apoptotic protein enhancing cell survival. Brain-derived neurotrophic factor (BDNF) also promotes neuronal viability but the links between XIAP and BDNF have remained unclear. We show here that the overexpression of XIAP increases BDNF in transgenic mice and cultured rat hippocampal neurons, whereas downregulation of XIAP by silencing RNA decreased BDNF. XIAP also stimulated BDNF signaling, as shown by increased phosphorylation of the TrkB receptor and the downstream molecule, cAMP response element-binding protein. The mechanism involved nuclear factor-kappaB (NF-kappaB) activation and blocking of NF-kappaB signaling inhibited the increased activities of BDNF promoters I and IV by XIAP. In neuronal cultures XIAP also upregulated interleukin (IL)-6, which is an NF-kappaB-responsive gene. The addition of IL-6 elevated whereas incubation with IL-6-blocking antibodies reduced BDNF in the neurons. BDNF itself activated NF-kappaB in the neurons at higher concentrations. The data show that XIAP has trophic effects on hippocampal neurons by increasing BDNF and TrkB activity. The results reveal a cytokine network in the brain involving BDNF, IL-6 and XIAP interconnected via the NF-kappaB system.

Tissue-specific and neural activity-regulated expression of human BDNF gene in BAC transgenic mice.

BACKGROUND: Brain-derived neurotrophic factor (BDNF) is a small secreted protein that has important roles in the developing and adult nervous system. Altered expression or changes in the regulation of the BDNF gene have been implicated in a variety of human nervous system disorders. Although regulation of the rodent BDNF gene has been extensively investigated, in vivo studies regarding the human BDNF gene are largely limited to postmortem analysis. Bacterial artificial chromosome (BAC) transgenic mice harboring the human BDNF gene and its regulatory flanking sequences constitute a useful tool for studying human BDNF gene regulation and for identification of therapeutic compounds modulating BDNF expression. RESULTS: In this study we have generated and analyzed BAC transgenic mice carrying 168 kb of the human BDNF locus modified such that BDNF coding sequence was replaced with the sequence of a fusion protein consisting of N-terminal BDNF and the enhanced green fluorescent protein (EGFP). The human BDNF-BAC construct containing all BDNF 5' exons preceded by different promoters recapitulated the expression of endogenous BDNF mRNA in the brain and several non-neural tissues of transgenic mice. All different 5' exon-specific BDNF-EGFP alternative transcripts were expressed from the transgenic human BDNF-BAC construct, resembling the expression of endogenous BDNF. Furthermore, BDNF-EGFP mRNA was induced upon treatment with kainic acid in a promotor-specific manner, similarly to that of the endogenous mouse BDNF mRNA. CONCLUSION: Genomic region covering 67 kb of human BDNF gene, 84 kb of upstream and 17 kb of downstream sequences is sufficient to drive tissue-specific and kainic acid-induced expression of the reporter gene in transgenic mice. The pattern of expression of the transgene is highly similar to BDNF gene expression in mouse and human. This is the first study to show that human BDNF gene is regulated by neural activity.

Alternative splicing and expression of human and mouse NFAT genes.

Four members of the nuclear factor of activated T cells (NFAT) family (NFATC1, NFATC2, NFATC3, and NFATC4) are Ca(2+)-regulated transcription factors that regulate several processes in vertebrates, including the development and function of the immune, cardiovascular, musculoskeletal, and nervous systems. Here we describe the structures and alternative splicing of the human and mouse NFAT genes, including novel splice variants for NFATC1, NFATC2, NFATC3, and NFATC4, and show the expression of different NFAT mRNAs in various mouse and human tissues and brain regions by RT-PCR. Our results show that alternatively spliced NFAT mRNAs are expressed differentially and could contribute to the diversity of functions of the NFAT proteins. Since NFAT family members are Ca(2+)-regulated and have critical roles in neuronal gene transcription in response to electrical activity, we describe the expression of NFATC1, NFATC2, NFATC3, and NFATC4 mRNAs in the adult mouse brain and in the adult human hippocampus using in situ hybridization and show that all NFAT mRNAs are expressed in the neurons of the mouse brain with specific patterns for each NFAT.

Shaping the human brain: evolutionary cis-regulatory plasticity drives changes in synaptic activity-controlled adaptive gene expression.

Neuronal activity-induced gene expression programs involved in synaptic structure- and plasticity-related functions are similar in mice and humans, yet bear distinct features. These include gains or losses of activity-responsiveness of certain genes and differences in gene induction profiles. Here, we discuss a possible origin of dissimilarities in activity-regulated transcription between species. We highlight that while synapse-to-nucleus signalling pathways are evolutionarily conserved, cis-regulatory plasticity has been driving species-specific remodelling of the activity-controlled enhancer landscape, thereby affecting gene regulation. In particular, evolutionary rearrangements of transcription factor binding site placements together with potential species-dependent developmental stage- and/or cell type-specific epigenetic and other trans-acting mechanisms are most likely at least in part accountable for between-species diversity in activity-regulated transcription. It is conceivable that cis-regulatory plasticity may have equipped the synaptic activity-driven adaptive gene program in human neurons with unique, species-specific qualities.

Networks of Cultured iPSC-Derived Neurons Reveal the Human Synaptic Activity-Regulated Adaptive Gene Program.

Long-term adaptive responses in the brain, such as learning and memory, require synaptic activity-regulated gene expression, which has been thoroughly investigated in rodents. Using human iPSC-derived neuronal networks, we show that the human and the mouse synaptic activity-induced transcriptional programs share many genes and both require Ca2+-regulated synapse-to-nucleus signaling. Species-specific differences include the noncoding RNA genes BRE-AS1 and LINC00473 and the protein-coding gene ZNF331, which are absent in the mouse genome, as well as several human genes whose orthologs are either not induced by activity or are induced with different kinetics in mice. These results indicate that lineage-specific gain of genes and DNA regulatory elements affects the synaptic activity-regulated gene program, providing a mechanism driving the evolution of human cognitive abilities.

Ultrastructural Characterization of Zika Virus Replication Factories.

A global concern has emerged with the pandemic spread of Zika virus (ZIKV) infections that can cause severe neurological symptoms in adults and newborns. ZIKV is a positive-strand RNA virus replicating in virus-induced membranous replication factories (RFs). Here we used various imaging techniques to investigate the ultrastructural details of ZIKV RFs and their relationship with host cell organelles. Analyses of human hepatic cells and neural progenitor cells infected with ZIKV revealed endoplasmic reticulum (ER) membrane invaginations containing pore-like openings toward the cytosol, reminiscent to RFs in Dengue virus-infected cells. Both the MR766 African strain and the H/PF/2013 Asian strain, the latter linked to neurological diseases, induce RFs of similar architecture. Importantly, ZIKV infection causes a drastic reorganization of microtubules and intermediate filaments forming cage-like structures surrounding the viral RF. Consistently, ZIKV replication is suppressed by cytoskeleton-targeting drugs. Thus, ZIKV RFs are tightly linked to rearrangements of the host cell cytoskeleton.