Project description:We performed SlamSeq (thiol(SH)-linked alkylation for metabolic sequencing) to estimate mRNA half-lives in subcellular compartments (neurites, soma-cytoplasm and nucleus) of primary cortical neurons.
Project description:We performed a SLAMseq Metabolic RNA Labeling on neuronal subcellular compartments, e.g. neurites and soma, derived from Ascl1-induced neurons (iNeurons). This experimental approach provides a snapshot of mRNA kinetics which allows to estimate the half-lives of mRNAs. These data was used to investigate the influence of mRNA degradation machinery in both neuronal compartments.
Project description:To study the correlation between mRNA stability and subcellular mRNA localisation we globally interferred with mRNA degradation in primary cortical neurons by overexpressing a catalytic mutant of deadenylase CAF1 that functions as a dominant negative form. Neurons were separated into subcellular compartments (neurite, soma-cytoplasm and nucleus) and sequenced in parallel with respective compartments from GFP- expressing control neuons.
Project description:We perturbed mRNA degradation machinery in mouse primary cortical neurons (mPCN) and investigated the change in mRNA half-lives. We performed SLAMseq Metabolic RNA Labeling on Mutant mPCN line harbouring a ponasteroneA-inducible heterozygous dominant-negative Caf1 (dnCaf1) and on GFP-transfected mPCNs. The Slamseq technique provided snapshots of mRNA kinetics allowing to estimate mRNA half-lives.
Project description:To study the effect of Larp1 on the abundance and subcellular localization of 5'TOP containing mRNAs, Larp1 was depleted from mouse primary cortical neurons using shRNAs. RNA from subcellular compartments (neurite and soma cytoplasm) was isolated and sequenced in parallel with scrambled control shRNA expressing samples.
Project description:To study how the codon composition of the coding sequence affects subcellular localization of mRNA in an unbiased way, mouse primary cortical neurons were transfected with a collection of human open reading frames (ORFs) with identical untranslated regions, separated into soma and neurites, and subjected to RNA sequencing with specific amplification of the ORFs.
Project description:We have perturbed mRNA degradation machinery and investigated the change in subcellular localization of mRNA in mouse primary cortical neurons (mPCNs). Mutant mPCN line harbouring a ponasteroneA-inducible heterozygous dominant-negative Caf1 (dnCaf1) was generated, separated into neuronal compartments (soma and neurites) and sequenced in parallel with compartments from the GFP-transfected cells (GFP, negative control).
Project description:To study the effect of m6A modifications on subcellular mRNA localization we depleted m6A writer Mettl3 with shRNA and small molecule from mouse primary cortical neurons (mPCN) and sequenced neuritic and somatic compartments in parallel with scrambled shRNA control.
Project description:To study the effect of m6A modifications on subcellular mRNA localization we depleted m6A readers Ythdf1, -2 and -3 with shRNAs from mouse primary cortical neurons (mPCN) and sequenced neuritic and somatic compartments in parallel with scrambled shRNA control.