Funktionelle Charakterisierung der nicht-kodierenden RNA Pantr1 in der FOXG1-abhängigen Vorderhirnentwicklung und im Rett-Syndrom

Rett-syndrome is an autism-spectrum disorder. Impaired neurodevelopment leads e.g. to progressive loss of cognitive capabilities, spastic paralysis, ataxia and epilepsy. 90% of Rett-syndrome cases are ascribed to mutations of MECP2, but FOXG1 haploinsufficiency results in similar phenotypes called atypical Rett-syndrome. Recent data suggest that expression of non-coding RNAs (ncRNA) is under control of FOXG1 as well as MECP2, and that long as well as micro RNAs are implicated in the disease. Specifically, our preliminary data showed that the miR200 family and the long ncRNA (lncRNA) Pantr1 have decreased transcriptional levels in the hippocampus of adult mice with reduced expression of FOXG1. Consequently, we aim to analyse molecular functions of Pantr1 within this project. Bridging the fields of bioinformatics, molecular biology, and biochemistry our approaches to elucidate the so far unknown function of Pantr1 in neural cells include target prediction and validation, as well as prediction and validation of transcriptional and post-transcriptional regulatory mechanisms. Specifically, we will explore the functionality of the interaction of Pantr1 with PURβ that we identified in our preliminary work. Within the proposed project we will (1) determine alterations of the transcriptome after changing expression levels of Pantr1 and PURβ using RNA-seq. (2) We will use chromatin immunoprecipitation by RNA pull down (ChIRP) to analyse whether PURβ promotes or hampers Pantr1 from binding to DNA. (3) We will employ Pantr1 ChIRP followed by mass spectrometry in conditions with and without PURβ expression to elucidate whether PURβ-binding interferes with association of Pantr1 to other RNA binding proteins (RBPs). (4) We will use iCLIP and ChIP to examine whether binding to Pantr1 interferes with binding of PURβ to RNA or DNA. In further experiments we will validate whether the rescued expression of Pantr1 and miR200 family can revert phenotypical alterations in vivo in the adult mouse hippocampus. In addition, we will explore whether reduced expression of FOXG1 in patient-derived neurons that will be generated from iPS cells, leads to comparable alterations of miR200 and Pantr1 expression and function. Together, these data on the role of ncRNAs in atypical Rett-syndrome will increase our understanding of the molecular basis of this neurodevelopmental disorder and inspire prospective concepts for therapy.

Start of project: 2019
End of project: 2023

Albert-Ludwigs-University Freiburg
Prof. Dr. Rolf Backofen
Bioinformatik
Prof. Dr. Rolf Backofen
Georges-Köhler-Allee 106
79110 Freiburg
Germany

Phone: +49 (0) 761-203-7461
Fax: +49 (0) 761-203-7462
Email: backofen@informatik.uni-freiburg.de
http://www.bioinf.uni-freiburg.de
Actual Research Report

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