Genomic analysis of the molecular neuropathology of tuberous sclerosis using a human stem cell model

Abstract

BackgroundTuberous sclerosis complex (TSC) is a genetic disease characterized by benign tumor growths in multiple organs and neurological symptoms induced by mTOR hyperfunction. Because the molecular pathology is highly complex and the etiology poorly understood, we employed a defined human neuronal model with a single mTOR activating mutation to dissect the disease-relevant molecular responses driving the neuropathology and suggest new targets for treatment.MethodsWe investigate the disease phenotype of TSC by neural differentiation of a human stem cell model that had been deleted for TSC2 by genome editing. Comprehensive genomic analysis was performed by RNA sequencing and ribosome profiling to obtain a detailed genome-wide description of alterations on both the transcriptional and translational level. The molecular effect of mTOR inhibitors used in the clinic was monitored and comparison to published data from patient biopsies and mouse models highlights key pathogenic processes.ResultsTSC2-deficient neural stem cells showed severely reduced neuronal maturation and characteristics of astrogliosis instead. Transcriptome analysis indicated an active inflammatory response and increased metabolic activity, whereas at the level of translation ribosomal transcripts showed a 5’UTR motif-mediated increase in ribosome occupancy. Further, we observed enhanced protein synthesis rates of angiogenic growth factors. Treatment with mTOR inhibitors corrected translational alterations but transcriptional dysfunction persisted.ConclusionsOur results extend the understanding of the molecular pathophysiology of TSC brain lesions, and suggest phenotype-tailored pharmacological treatment strategies.Electronic supplementary materialThe online version of this article (doi:10.1186/s13073-016-0347-3) contains supplementary material, which is available to authorized users.

NMD is required for timely cell fate transitions by fine-tuning gene expression and controlling translation

“…6a). The increase in translation rates observed in NMD KO ESCs was similar in magnitude to that observed for Tsc2 KOs, in which deregulated mTORC1 activity leads to an increase in translation 32 . Full Eif4a2 KOs, but also specific depletion of the PTC isoform resulted in mild reductions of translation rates.…”

Recent advances in human stem cell-based modeling of Tuberous Sclerosis Complex

“…Discrepancies between the findings may reflect gene dosedependent effects of cell lines and culture variability. Moreover, transcriptomic analysis of isogenic, geneedited TSC2 heterozygous and homozygous cultures showed significant differences between TSC2 −/− and TSC2 +/+ neurons but not between TSC2 +/− and TSC2 +/+ neurons [26,35]. These phenotypic differences need to be further investigated with additional iPSC-derived TSC patients and control cell lines to determine the link to the TSC2 mutation, cell line variability, or genetic background ( Fig.…”

Progress of Induced Pluripotent Stem Cell Technologies to Understand Genetic Epilepsy

“…Biallelic mutant cells display transcriptome alterations, closely related to those observed for human cortical tubers, and show an active inflammatory state and increased metabolic activity. In addition, neuroprogenitors, neuronal and glial cells show somatic hypertrophy and neuronal delayed maturation [44,45]. Electrophysiological analyses have been undertaken to explore those mechanisms underlying hyperexcitability.…”