A novel long non-coding natural antisense RNA is a negative regulator of Nos1 gene expression

Long non-coding natural antisense transcripts (NATs) are widespread in eukaryotic species. Although recent studies indicate that long NATs are engaged in the regulation of gene expression, the precise functional roles of the vast majority of them are unknown. Here we report that a long NAT (Mm-antiNos1 RNA) complementary to mRNA encoding the neuronal isoform of nitric oxide synthase (Nos1) is expressed in the mouse brain and is transcribed from the non-template strand of the Nos1 locus. Nos1 produces nitric oxide (NO), a major signaling molecule in the CNS implicated in many important functions including neuronal differentiation and memory formation. We show that the newly discovered NAT negatively regulates Nos1 gene expression. Moreover, our quantitative studies of the temporal expression profiles of Mm-antiNos1 RNA in the mouse brain during embryonic development and postnatal life indicate that it may be involved in the regulation of NO-dependent neurogenesis

Modeling the functional genomics of autism using human neurons.

Human neural progenitors from a variety of sources present new opportunities to model aspects of human neuropsychiatric disease in vitro. Such in vitro models provide the advantages of a human genetic background combined with rapid and easy manipulation, making them highly useful adjuncts to animal models. Here, we examined whether a human neuronal culture system could be utilized to assess the transcriptional program involved in human neural differentiation and to model some of the molecular features of a neurodevelopmental disorder, such as autism. Primary normal human neuronal progenitors (NHNPs) were differentiated into a post-mitotic neuronal state through addition of specific growth factors and whole-genome gene expression was examined throughout a time course of neuronal differentiation. After 4 weeks of differentiation, a significant number of genes associated with autism spectrum disorders (ASDs) are either induced or repressed. This includes the ASD susceptibility gene neurexin 1, which showed a distinct pattern from neurexin 3 in vitro, and which we validated in vivo in fetal human brain. Using weighted gene co-expression network analysis, we visualized the network structure of transcriptional regulation, demonstrating via this unbiased analysis that a significant number of ASD candidate genes are coordinately regulated during the differentiation process. As NHNPs are genetically tractable and manipulable, they can be used to study both the effects of mutations in multiple ASD candidate genes on neuronal differentiation and gene expression in combination with the effects of potential therapeutic molecules. These data also provide a step towards better understanding of the signaling pathways disrupted in ASD

Timed and targeted differential regulation of nitric oxide synthase (NOS) and anti-NOS genes by reward conditioning leading to long-term memory formation

In a number of neuronal models of learning, signaling by the neurotransmitter nitric oxide (NO), synthesized by the enzyme neuronal NO synthase (nNOS), is essential for the formation of long-term memory (LTM). Using the molluscan model system Lymnaea, we investigate here whether LTM formation is associated with specific changes in the activity of members of the NOS gene family: Lym-nNOS1, Lym-nNOS2, and the antisense RNA-producing pseudogene (anti-NOS). We show that expression of the Lym-nNOS1 gene is transiently upregulated in cerebral ganglia after conditioning. The activation of the gene is precisely timed and occurs at the end of a critical period during which NO is required for memory consolidation. Moreover, we demonstrate that this induction of the Lym-nNOS1 gene is targeted to an identified modulatory neuron called the cerebral giant cell (CGC). This neuron gates the conditioned feeding response and is an essential part of the neural network involved in LTM formation. We also show that the expression of the anti-NOS gene, which functions as a negative regulator of nNOS expression, is downregulated in the CGC by training at 4 h after conditioning, during the critical period of NO requirement. This appears to be the first report of the timed and targeted differential regulation of the activity of a group of related genes involved in the production of a neurotransmitter that is necessary for learning, measured in an identified neuron of known function. We also provide the first example of the behavioral regulation of a pseudogene

Induction and repression of mammalian achaete-scute homologue (MASH) gene expression during neuronal differentiation of P19 embryonal carcinoma cells

MASH1 and MASH2, mammalian homologues of the Drosophila neural determination genes achaete-scute, are members of the basic helix-loop-helix (bHLH) family of transcription factors. We show here that murine P19 embryonal carcinoma cells can be used as a model system to study the regulation and function of these genes. MASH1 and MASH2 display complementary patterns of expression during the retinoic-acid-induced neuronal differentiation of P19 cells. MASH1 mRNA is undetectable in undifferentiated P19 cells but is induced to high levels by retinoic acid coincident with neuronal differentiation. In contrast, MASH2 mRNA is expressed in undifferentiated P19 cells and is repressed by retinoic acid treatment. These complementary expression patterns suggest distinct functions for MASH1 and MASH2 in development, despite their sequence homology. In retinoic-acid-treated P19 cells, MASH1 protein expression precedes and then overlaps expression of neuronal markers. However, MASH1 is expressed by a smaller proportion of cells than expresses such markers. MASH1 immunoreactivity is not detected in differentiated cells displaying a neuronal morphology, suggesting that its expression is transient. These features of MASH1 expression are similar to those observed in vivo, and suggest that P19 cells represent a good model system in which to study the regulation of this gene. Forced expression of MASH1 was achieved in undifferentiated P19 cells by transfection of a cDNA expression construct. The transfected cells expressing exogenous MASH1 protein contained E-box-binding activity that could be super-shifted by an anti-MASH1 antibody, but exhibited no detectable phenotypic changes. Thus, unlike myogenic bHLH genes, such as MyoD, which are sufficient to induce muscle differentiation, expression of MASH1 appears insufficient to promote neurogenesis

Transcriptional and Epigenetic Regulation in Injury-Mediated Neuronal Dendritic Plasticity

Injury to the nervous system induces localized damage in neural structures and neuronal death through the primary insult, as well as delayed atrophy and impaired plasticity of the delicate dendritic fields necessary for interneuronal communication. Excitotoxicity and other secondary biochemical events contribute to morphological changes in neurons following injury. Evidence suggests that various transcription factors are involved in the dendritic response to injury and potential therapies. Transcription factors play critical roles in the intracellular regulation of neuronal morphological plasticity and dendritic growth and patterning. Mounting evidence supports a crucial role for epigenetic modifications via histone deacetylases, histone acetyltransferases, and DNA methyltransferases that modify gene expression in neuronal injury and repair processes. Gene regulation through epigenetic modification is of great interest in neurotrauma research, and an early picture is beginning to emerge concerning how injury triggers intracellular events that modulate such responses. This review provides an overview of injury-mediated influences on transcriptional regulation through epigenetic modification, the intracellular processes involved in the morphological consequences of such changes, and potential approaches to the therapeutic manipulation of neuronal epigenetics for regulating gene expression to facilitate growth and signaling through dendritic arborization following injury

Cellular excitability and the regulation of functional neuronal identity: from gene expression to neuromodulation

The intrinsic properties of a neuron determine the translation of synaptic input to axonal output. It is this input– output relationship that is the heart of all nervous system activity. As such, the overall regulation of the intrinsic excitability of a neuron directly determines the output of that neuron at a given point in time, giving the cell a unique “functional identity.” To maintain this distinct functional output, neurons must adapt to changing patterns of synaptic excitation. These adaptations are essential to prevent neurons from either falling silent as synaptic excitation falls or becoming saturated as excitation increases. In the absence of stabilizing mechanisms, activity-dependent plasticity could drive neural activity to saturation or quiescence. Furthermore, as cells adapt to changing patterns of synaptic input, presumably the overall balance of intrinsic conductances of the cell must be maintained so that reliable output is achieved (Daoudal and Debanne, 2003; Turrigiano and Nelson, 2004; Frick and Johnston, 2005). Although these regulatory phenomena have been well documented, the molecular and physiological mechanisms involved are poorly understood

Estrous behavior in dairy cows: identification of underlying mechanisms and gene functions

Selection in dairy cattle for a higher milk yield has coincided with declined fertility. One of the factors is reduced expression of estrous behavior. Changes in systems that regulate the estrous behavior could be manifested by altered gene expression. This literature review describes the current knowledge on mechanisms and genes involved in the regulation of estrous behavior. The endocrinological regulation of the estrous cycle in dairy cows is well described. Estradiol (E2) is assumed to be the key regulator that synchronizes endocrine and behavioral events. Other pivotal hormones are, for example, progesterone, gonadotropin releasing hormone and insulin-like growth factor-1. Interactions between the latter and E2 may play a role in the unfavorable effects of milk yield-related metabolic stress on fertility in high milk-producing dairy cows. However, a clear understanding of how endocrine mechanisms are tied to estrous behavior in cows is only starting to emerge. Recent studies on gene expression and signaling pathways in rodents and other animals contribute to our understanding of genes and mechanisms involved in estrous behavior. Studies in rodents, for example, show that estrogen-induced gene expression in specific brain areas such as the hypothalamus play an important role. Through these estrogen-induced gene expressions, E2 alters the functioning of neuronal networks that underlie estrous behavior, by affecting dendritic connections between cells, receptor populations and neurotransmitter releases. To improve the understanding of complex biological networks, like estrus regulation, and to deal with the increasing amount of genomic information that becomes available, mathematical models can be helpful. Systems biology combines physiological and genomic data with mathematical modeling. Possible applications of systems biology approaches in the field of female fertility and estrous behavior are discusse

Aberrant neuronal activity-induced signaling and gene expression in a mouse model of RASopathy

Noonan syndrome (NS) is characterized by reduced growth, craniofacial abnormalities, congenital heart defects, and variable cognitive deficits. NS belongs to the RASopathies, genetic conditions linked to mutations in components and regulators of the Ras signaling pathway. Approximately 50% of NS cases are caused by mutations in PTPN11. However, the molecular mechanisms underlying cognitive impairments in NS patients are still poorly understood. Here, we report the generation and characterization of a new conditional mouse strain that expresses the overactive Ptpn11D61Y allele only in the forebrain. Unlike mice with a global expression of this mutation, this strain is viable and without severe systemic phenotype, but shows lower exploratory activity and reduced memory specificity, which is in line with a causal role of disturbed neuronal Ptpn11 signaling in the development of NS-linked cognitive deficits. To explore the underlying mechanisms we investigated the neuronal activity-regulated Ras signaling in brains and neuronal cultures derived from this model. We observed an altered surface expression and trafficking of synaptic glutamate receptors, which are crucial for hippocampal neuronal plasticity. Furthermore, we show that the neuronal activity-induced ERK signaling, as well as the consecutive regulation of gene expression are strongly perturbed. Microarray-based hippocampal gene expression profiling revealed profound differences in the basal state and upon stimulation of neuronal activity. The neuronal activity-dependent gene regulation was strongly attenuated in Ptpn11D61Y neurons. In silico analysis of functional networks revealed changes in the cellular signaling beyond the dysregulation of Ras/MAPK signaling that is nearly exclusively discussed in the context of NS at present. Importantly, changes in PI3K/AKT/mTOR and JAK/STAT signaling were experimentally confirmed. In summary, this study uncovers aberrant neuronal activity-induced signaling and regulation of gene expression in Ptpn11D61Y mice and suggests that these deficits contribute to the pathophysiology of cognitive impairments in NS

An investigation of herpes simplex virus promoter activity compatible with latency establishment reveals VP16-independent activation of immediate-early promoters in sensory neurones

Herpes simplex virus (HSV) type-1 establishes lifelong latency in sensory neurones and it is widely assumed that latency is the consequence of a failure to initiate virus immediate-early (IE) gene expression. However, using a Ore reporter mouse system in conjunction with Ore-expressing HSV-1 recombinants we have previously shown that activation of the IE ICPO promoter can precede latency establishment in at least 30 % of latently infected cells. During productive infection of non-neuronal cells, IE promoter activation is largely dependent on the transactivator VP16 a late structural component of the virion. Of significance, VP16 has recently been shown to exhibit altered regulation in neurones; where its de novo synthesis is necessary for IE gene expression during both lytic infection and reactivation from latency. In the current study, we utilized the Ore reporter mouse model system to characterize the full extent of viral promoter activity compatible with cell survival and latency establishment. In contrast to the high frequency activation of representative IE promoters prior to latency establishment, cell marking using a virus recombinant expressing Ore under VP16 promoter control was very inefficient. Furthermore, infection of neuronal cultures with VP16 mutants reveals a strong VP16 requirement for IE promoter activity in non-neuronal cells, but not sensory neurones. We conclude that only IE promoter activation can efficiently precede latency establishment and that this activation is likely to occur through a VP16-independent mechanism