Background Publicly available RNA-seq datasets are often underused although being helpful to improve functional annotation of eukaryotic genomes. This is especially true for filamentous fungi genomes which structure differs from most well annotated yeast genomes. Podospora anserina is a filamentous fungal model, which genome has been sequenced and annotated in 2008. Still, the current annotation lacks information about cis-regulatory elements, including promoters, transcription starting sites and terminators, which are instrumental to integrate epigenomic features into global gene regulation strategies. Results Here we took advantage of 37 RNA-seq experiments that were obtained in contrasted developmental and physiological conditions, to complete the functional annotation of P. anserina genome. Out of the 10,800 previously annotated genes, 5’UTR and 3’UTR were defined for 7554, among which, 3328 showed differential transcriptional signal starts and/or transcriptional end sites. In addition, alternative splicing events were detected for 2350 genes, mostly due alternative 3’splice sites and 1732 novel transcriptionally active regions (nTARs) in unannotated regions were identified. Conclusions Our study provides a comprehensive genome-wide functional annotation of P. anserina genome, including chromatin features, cis-acting elements such as UTRs, alternative splicing events and transcription of non-coding regions. These new findings will likely improve our understanding of gene regulation strategies in compact genomes, such as those of filamentous fungi. Characterization of alternative transcripts and nTARs paves the way to the discovery of putative new genes, alternative peptides or regulatory non-coding RNAs.
During the embryonic period, neuronal communication starts before the establishment of the synapses with alternative forms of neuronal excitability, called here Embryonic Neural Excitability (ENE). ENE has been shown to modulate the unfolding of development transcriptional programs, but the global consequences for developing organisms are not all understood. Here we monitored calcium transients in the telencephalon of zebrafish embryos as a proxy for ENE to assess the efficacy of transient pharmacological treatments to either increase or decrease ENE. Increasing or decreasing ENE at the end of the embryonic period promoted an increase or a decrease in the numbers of dopamine (DA) neurons, respectively. This plasticity of dopaminergic specification occurs in the subpallium of zebrafish larvæ at 6 dpf, within a relatively stable population of vMAT2-positive cells. Non-dopaminergic vMAT2-positive cells hence constitute an unanticipated biological marker for a reserve pool of DA neurons that can be recruited by ENE.Modulating ENE also affected larval locomotion several days after the end of the treatments. In particular, the increase of ENE from 2 to 3 dpf promoted hyperlocomotion of larvæ at 6 dpf, reminiscent of zebrafish endophenotypes reported for Attention Deficit with Hyperactivity Disorders. These results provide a convenient framework for identifying environmental factors that could disturb ENE as well as to study the molecular mechanisms linking ENE to neurotransmitter specification.Significance Statement- Spontaneous calcium transients, used as a proxy for Embryonic Neural Excitability (ENE), are detected in the forebrain of embryonic zebrafish.- Short-term pharmacological treatments by bath application could increase or decrease ENE.- The post-mitotic differentiation of the dopaminergic phenotype is modulated by ENE in the zebrafish forebrain.- The plasticity of the dopaminergic specification occurs within a reserve pool of vMAT2-positive cells.- Transient increase of ENE at the end of the embryonic period induces hyperlocomotion, a phenotype associated with ADHD in this model.- Our results provide a convenient framework to study the molecular mechanisms linking ENE to neurotransmitter specification.
Neuronal communication starts before the establishment of the synapses with forms of neuronal excitability occurring during the embryonic period, we called here Embryonic Neuronal Excitability (ENE). ENE has been shown to modulate the correct unfolding of development transcriptional programs but the global consequences for the developing organisms are not all understood. Here we monitored calcium transients as a proxy for ENE in zebrafish to assess the efficacy of transient pharmacological treatments applied by balneation during the embryonic period to modulate ENE. We also report lasting effects of 24h treatments, performed at the end of the embryonic development, on morphology and behavior of larval zebrafish. The post-mitotic differentiation of the dopaminergic phenotype is modulated by ENE in the forebrain. The plasticity of the dopaminergic specification occurs within a stable population of vMAT2 immuno-reactive cells, hence identifying an unanticipated biological marker for this reserve pool. We also report an effect of ENE on locomotion several days after the end of the treatments. In particular, the increase of ENE from 2 to 3 dpf promoted an hyperlocomotion in 6dpf zebrafish larvae which is an endophenotype for Attention Deficit with Hyperactivity Disorders and schizophrenia in zebrafish. These results provide a convenient framework to identify environmental factors that could regulate ENE and to study further the molecular mechanisms linking ENE to the neurotransmitters specification, with clinical relevance for the pathogenesis of neurodevelopmental disorders.
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