Rico Schieweck

Ribosomal RNA (rRNA) transcription rates vary during development, and their dysregulation is linked to diseases such as cancer and ribosomopathies. Owing to their high abundance and genomic redundancy, the functional significance of rRNA-levels remains unclear. Here, we developed TAPIR (Targeted Activation of Protein Translation), a CRISPR-based approach to elevate rRNA-levels by inducing 47S rDNA transcription. TAPIR increased nucleolar size and enhanced protein synthesis, even in rapidly proliferating cells. In neural stem cells, elevated translation promoted self-renewal and proliferation in vitro and in vivo. Furthermore, TAPIR enabled the modeling and partial rescue of associated disease phenotypes. Our findings revealed that rRNA-levels directly regulate translational output and that protein synthesis capacity can act as a key determinant of mammalian stem cell behavior.

Mitochondria are essential organelles of eukaryotic cells and are characterized by their unique and complex membrane system. They are confined from the cytosol by an envelope consisting of two membranes. Signals, metabolites, proteins and lipids have to be transferred across these membranes via proteinaceous contact sites to keep mitochondria functional. In the present study, we identified a novel mitochondrial contact site in Saccharomyces cerevisiae that is formed by the inner membrane protein Cqd1 and the outer membrane proteins Por1 and Om14. Similar to what is found for the mitochondrial porin Por1, Cqd1 is highly conserved, suggesting that this complex is conserved in form and function from yeast to human. Cqd1 is a member of the UbiB protein kinase-like family (also called aarF domain-containing kinases). It was recently shown that Cqd1, in cooperation with Cqd2, controls the cellular distribution of coenzyme Q by a yet unknown mechanism. Our data suggest that Cqd1 is additionally involved in phospholipid homeostasis. Moreover, overexpression of CQD1 and CQD2 causes tethering of mitochondria to the endoplasmic reticulum, which might explain the ability of Cqd2 to rescue ERMES deletion phenotypes.

Protein homeostasis (proteostasis) is a prerequisite for cellular viability and plasticity. In particular, post-mitotic cells such as neurons rely on a tightly regulated safeguard system that allows for regulated protein expression. Previous investigations have identified RNA-binding proteins (RBPs) as crucial regulators of protein expression in nerve cells. However, during neurodegeneration, their ability to control the proteome is progressively disrupted. In this review, we examine the malfunction of key RBPs such as TAR DNA-binding protein 43 (TDP-43), Fused in Sarcoma (FUS), Staufen, Pumilio and fragile-X mental retardation protein (FMRP). Therefore, we focus on two key aspects of RBP dysfunctions in neurodegeneration: protein aggregation and dysregulation of their target RNAs. Moreover, we discuss how the chaperone system responds to changes in the RBP-controlled transcriptome. Based on recent findings, we propose a two-hit model in which both, harmful RBP deposits and target mRNA mistranslation contribute to neurodegeneration observed in RBPathologies.

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.

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.

The hippocampus is central for higher cognition and emotions. In patients suffering from neuropsychiatric or neurodegenerative diseases, hippocampal signaling is altered causing cognitive defects. Thus, therapeutic approaches aim at improving cognition by targeting the hippocampus. Enhanced physical activity (EPA) improves cognition in rodents and humans. A systematic screen, however, for expression changes in the hippocampus along the dorso-ventral axis is missing, which is a prerequisite for understanding molecular mechanisms. Here, we exploited label free mass spectrometry to detect proteomic changes in the hippocampus of male mice upon voluntary wheel running. To identify regional differences, we examined dorsal and ventral CA1, CA3 and dentate gyrus hippocampal subregions. We found metabolic enzymes and actin binding proteins, such as RhoA, being upregulated in the hippocampus upon EPA suggesting a coordination between metabolism and cytoskeleton remodeling; two pathways essential for synaptic plasticity. Strikingly, dorsal and ventral hippocampal subregions respond differentially to EPA. Together, our results provide new insight into proteomic adaptations driven by physical activity in mice. In addition, our results suggest that dorsal and ventral hippocampus, as well as hippocampal subregions themselves, contribute differently to this process. Our study therefore provides an important resource for studying hippocampal subregion diversity in response to EPA.

Behavior and higher cognition rely on the transfer of information between neurons through specialized contact sites termed synapses. Plasticity of neuronal circuits, a prerequisite to respond to environmental changes, is intrinsically coupled with the nerve cell’s ability to form, structurally modulate or remove synapses. Consequently, the synaptic proteome undergoes dynamic alteration on demand in a spatiotemporally restricted manner. Therefore, proper protein localization at synapses is essential for synaptic function. This process is regulated by: (i) protein transport and recruitment; (ii) local protein synthesis; and (iii) synaptic protein degradation. These processes shape the transmission efficiency of excitatory synapses. Whether and how these processes influence synaptic inhibition is, however, widely unknown. Here, we summarize findings on fundamental regulatory processes that can be extrapolated to inhibitory synapses. In particular, we focus on known aspects of posttranscriptional regulation and protein dynamics of the GABA receptor (GABAR). Finally, we propose that local (co)-translational control mechanism might control transmission of inhibitory synapses.

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 (Stau2^GT) 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, Stau2^GT 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.