mTOR and MAPK: from localized translation control to epilepsy

Background: Epilepsy is one of the most common neurological diseases characterized by excessive hyperexcitability of neurons. Molecular mechanisms of epilepsy are diverse and not really understood. All in common is the misregulation of proteins that determine excitability such as potassium and sodium channels as well as GABA receptors;which are all known as biomarkers for epilepsy. Two recently identified key pathways involve the kinases mechanistic target of rapamycin (mTOR) and mitogen-activated protein kinases (MAPK). Interestingly, mRNAs coding for those biomarkers are found to be localized at or near synapses indicating a local misregulation of synthesis and activity. Results: Research in the last decade indicates that RNA-binding proteins (RBPs) responsible for mRNA localization, stability and translation mediate local expression control. Among others, they are affected by mTOR and MAPK to guide expression of epileptic factors. These results suggest that mTOR/MAPK act on RBPs to regulate the fate of mRNAs, indicating a misregulation of protein expression at synapses in epilepsy. Conclusion: We propose that mTOR and MAPK regulate RBPs, thereby guiding the local expression of their target-mRNAs encoding for markers of epilepsy. Thus, misregulated mTOR/MAPK-RBP interplay may result in excessive local synthesis of ion channels and receptors thereby leading to hyperexcitability. Continuous stimulation of synapses further activates mTOR/MAPK pathway reinforcing their effect on RBP-mediated expression control establishing the basis for epilepsy. Here, we highlight findings showing the tight interplay between mTOR as well as MAPK with RBPs to control expression for epileptic biomarkers

An expanded CAG repeat in Huntingtin Causes +1 frameshifting

Maintenance of triplet decoding is crucial for the expression of functional protein because deviations either into the −1 or +1 reading frames are often non-functional. We report here that expression of huntingtin (Htt) exon 1 with expanded CAG repeats, implicated in Huntington pathology, undergoes a sporadic +1 frameshift to generate from the CAG repeat a trans-frame AGC repeat-encoded product. This +1 recoding is exclusively detected in pathological Htt variants, i.e. those with expanded repeats with more than 35 consecutive CAG codons. An atypical +1 shift site, UUC C at the 5′ end of CAG repeats, which has some resemblance to the influenza A virus shift site, triggers the +1 frameshifting and is enhanced by the increased propensity of the expanded CAG repeats to form a stem-loop structure. The +1 trans-frame-encoded product can directly influence the aggregation of the parental Htt exon 1

Pumilio2-deficient mice show a predisposition for epilepsy

Epilepsy is a neurological disease that is caused by abnormal hypersynchronous activities of neuronal ensembles leading to recurrent and spontaneous seizures in human patients. Enhanced neuronal excitability and a high level of synchrony between neurons seem to trigger these spontaneous seizures. The molecular mechanisms, however, regarding the development of neuronal hyperexcitability and maintenance of epilepsy are still poorly understood. Here, we show that pumilio RNA-binding family member 2 (Pumilio2;Pum2) plays a role in the regulation of excitability in hippocampal neurons of weaned and 5-month-old male mice. Almost complete deficiency of Pum2 in adult Pum2 gene-trap mice (Pum2 GT) causes misregulation of genes involved in neuronal excitability control. Interestingly, this finding is accompanied by the development of spontaneous epileptic seizures in Pum2 GT mice. Furthermore, we detect an age-dependent increase in Scn1a (Na(v)1.1) and Scn8a (Na(v)1.6) mRNA levels together with a decrease in Scn2a (Na(v)1.2) transcript levels in weaned Pum2 GT that is absent in older mice. Moreover, field recordings of CA1 pyramidal neurons show a tendency towards a reduced paired-pulse inhibition after stimulation of the Schaffer-collateral-commissural pathway in Pum2 GT mice, indicating a predisposition to the development of spontaneous seizures at later stages. With the onset of spontaneous seizures at the age of 5 months, we detect increased protein levels of Na(v)1.1 and Na(v)1.2 as well as decreased protein levels of Na(v)1.6 in those mice. In addition, GABA receptor subunit alpha-2 (Gabra2) mRNA levels are increased in weaned and adult mice. Furthermore, we observe an enhanced GABRA2 protein level in the dendritic field of the CA1 subregion in the Pum2 GT hippocampus. We conclude that altered expression levels of known epileptic risk factors such as Na(v)1.1, Na(v)1.2, Na(v)1.6 and GABRA2 result in enhanced seizure susceptibility and manifestation of epilepsy in the hippocampus. Thus, our results argue for a role of Pum2 in epileptogenesis and the maintenance of epilepsy

Forebrain-specific, conditional silencing of Staufen2 alters synaptic plasticity, learning, and memory in rats

Background: Dendritic messenger RNA (mRNA) localization and subsequent local translation in dendrites critically contributes to synaptic plasticity and learning and memory. Little is known, however, about the contribution of RNA-binding proteins (RBPs) to these processes in vivo. Results: To delineate the role of the double-stranded RBP Staufen2 (Stau2), we generate a transgenic rat model, in which Stau2 expression is conditionally silenced by Cre-inducible expression of a microRNA (miRNA) targeting Stau2 mRNA in adult forebrain neurons. Known physiological mRNA targets for Stau2, such as RhoA, Complexin 1, and Rgs4 mRNAs, are found to be dysregulated in brains of Stau2-deficient rats. In vivo electrophysiological recordings reveal synaptic strengthening upon stimulation, showing a shift in the frequency-response function of hippocampal synaptic plasticity to favor long-term potentiation and impair long-term depression in Stau2-deficient rats. These observations are accompanied by deficits in hippocampal spatial working memory, spatial novelty detection, and in tasks investigating associative learning and memory. Conclusions: Together, these experiments reveal a critical contribution of Stau2 to various forms of synaptic plasticity including spatial working memory and cognitive management of new environmental information. These findings might contribute to the development of treatments for conditions associated with learning and memory deficits

Pumilio2 Promotes Growth of Mature Neurons

RNA-binding proteins (RBPs) are essential regulators controlling both the cellular transcriptome and translatome. These processes enable cellular plasticity, an important prerequisite for growth. Cellular growth is a complex, tightly controlled process. Using cancer cells as model, we looked for RBPs displaying strong expression in published transcriptome datasets. Interestingly, we found the Pumilio (Pum) protein family to be highly expressed in all these cells. Moreover, we observed that Pum2 is regulated by basic fibroblast growth factor (bFGF). bFGF selectively enhances protein levels of Pum2 and the eukaryotic initiation factor 4E (eIF4E). Exploiting atomic force microscopy and in vitro pulldown assays, we show that Pum2 selects for eIF4E mRNA binding. Loss of Pum2 reduces eIF4E translation. Accordingly, depletion of Pum2 led to decreased soma size and dendritic branching of mature neurons, which was accompanied by a reduction in essential growth factors. In conclusion, we identify Pum2 as an important growth factor for mature neurons. Consequently, it is tempting to speculate that Pum2 may promote cancer growth

Evaluation of a Configurable, Mobile and Modular Floor-Pen System for Group-Housing of Laboratory Rabbits

The major responsibility of researchers and laboratory animal facilities is to ensure animal well-being during the time of acclimatization, experiments, and recovery. In this context, animal housing conditions are of utmost importance. Here, we implemented a mobile and modular floor-pen housing system for laboratory rabbits that combines rabbits’ natural behavioral requirements and the high hygiene standards needed in biomedical science. Twelve female New Zealand White (NZW) rabbits were single- or group-housed for 12 months in mobile and modular floor-pens. Their general health status was evaluated at the end of the experimental setup. Further, we performed behavioral analysis of six additional NZW females group-housed for eight weeks in pens of two different sizes. We show that our improved housing concept supported species-specific behavioral patterns. Taken together, our housing system provides an optimal setup for rabbits in animal facilities that combines strict requirements for animal experiments with animal welfare

Altered Glutamate Receptor Ionotropic Delta Subunit 2 Expression in Stau2-Deficient Cerebellar Purkinje Cells in the Adult Brain

Staufen2 (Stau2) is an RNA-binding protein that is involved in dendritic spine morphogenesis and function. Several studies have recently investigated the role of Stau2 in the regulation of its neuronal target mRNAs, with particular focus on the hippocampus. Here, we provide evidence for Stau2 expression and function in cerebellar Purkinje cells. We show that Stau2 downregulation (Stau2GT) led to an increase of glutamate receptor ionotropic delta subunit 2 (GluD2) in Purkinje cells when animals performed physical activity by voluntary wheel running compared with the age-matched wildtype (WT) mice (C57Bl/6J). Furthermore, Stau2GT mice showed lower performance in motor coordination assays but enhanced motor learning abilities than did WT mice, concomitantly with an increase in dendritic GluD2 expression. Together, our results suggest the novel role of Stau2 in Purkinje cell synaptogenesis in the mouse cerebellum