Project description:In order to identify what RNA messengers are being translated at the ER membrane, membrane-associated polysomes were separated from free polysomes by subcellular fractionation of WT and delta-egd1/delta-egd2 yeast strains and associated RNA was isolated and then identified by microarray analysis.
Project description:In order to identify what RNA messengers are being translated at the ER membrane, membrane-associated polysomes were separated from free polysomes by subcellular fractionation of WT and delta-egd1/delta-egd2 yeast strains and associated RNA was isolated and then identified by microarray analysis. Set of arrays that are part of repeated experiments
Project description:Nonsense-mediated decay (NMD) is a translation-dependent RNA quality control mechanism that occurs in the cytoplasm. However, it is unknown how NMD regulates the stability of RNAs translated at the Endoplasmic Reticulum (ER). Here, we identify a localized NMD pathway dedicated to ER-translated mRNAs. We previously identified NBAS, a component of the Syntaxin 18 complex involved in Golgi-to-ER trafficking, as a novel NMD factor. Here, we show that NBAS fulfils an independent function in NMD. This ER-NMD pathway requires the interaction of NBAS with the core NMD factor UPF1, which is partially localized at the ER in the proximity of the translocon. NBAS and UPF1 co-regulate the stability of ER-associated transcripts, in particular those associated with the cellular stress response. We propose a model where NBAS recruits UPF1 to the membrane of the ER and activates an ER-dedicated NMD pathway, thus providing an ER protective function by ensuring quality control of ER-translated mRNAs.
Project description:Nonsense-mediated decay (NMD) is a translation-dependent RNA quality control mechanism that occurs in the cytoplasm. However, it is unknown how NMD regulates the stability of RNAs translated at the Endoplasmic Reticulum (ER). Here, we identify a localized NMD pathway dedicated to ER-translated mRNAs. We previously identified NBAS, a component of the Syntaxin 18 complex involved in Golgi-to-ER trafficking, as a novel NMD factor. Here, we show that NBAS fulfils an independent function in NMD. This ER-NMD pathway requires the interaction of NBAS with the core NMD factor UPF1, which is partially localized at the ER in the proximity of the translocon. NBAS and UPF1 co-regulate the stability of ER-associated transcripts, in particular those associated with the cellular stress response. We propose a model where NBAS recruits UPF1 to the membrane of the ER and activates an ER-dedicated NMD pathway, thus providing an ER protective function by ensuring quality control of ER-translated mRNAs.
Project description:In this study we focus on two Saccharomyces cerevisiae strains with varying production of heterologous α-amylase and we compare the metabolic fluxes and transcriptional regulation at aerobic and anaerobic conditions, in particular with the objective to identify the final electron acceptor for protein folding. We found that anaerobic conditions showed high amount of amylase productions when comparing to aerobic conditions and the genome-scale transcriptional analysis suggested that genes related to the endoplasmic reticulum (ER), lipid synthesis and stress responses were generally up-regulated at anaerobic conditions. Moreover, we proposed a model for the electron transfer from ER to the final electron acceptor, fumarate under anaerobic conditions.
Project description:Aneuploidy and aging are correlated; however, a causal link between these two phenomena has remained elusive. Here we show that yeast disomic for a single native yeast chromosome generally have a decreased replicative lifespan. In addition, the extent of this lifespan deficit correlates with the size of the extra chromosome. We identified a mutation in BUL1 that rescues both the lifespan deficit and a protein trafficking defect in yeast disomic for chromosome 5. Bul1 is an E4 ubiquitin ligase adaptor involved in a protein quality-control pathway that targets membrane proteins for endocytosis and destruction in the lysosomal vacuole thereby maintaining protein homeostasis. Concurrent suppression of the aging and trafficking phenotypes suggests that disrupted membrane protein homeostasis in aneuploid yeast may contribute to their accelerated aging. The data reported here demonstrate that aneuploidy can impair protein homeostasis, shorten lifespan, and may contribute to age-associated phenotypes.
Project description:In this study we focus on two Saccharomyces cerevisiae strains with varying production of heterologous M-NM-1-amylase and we compare the metabolic fluxes and transcriptional regulation at aerobic and anaerobic conditions, in particular with the objective to identify the final electron acceptor for protein folding. We found that anaerobic conditions showed high amount of amylase productions when comparing to aerobic conditions and the genome-scale transcriptional analysis suggested that genes related to the endoplasmic reticulum (ER), lipid synthesis and stress responses were generally up-regulated at anaerobic conditions. Moreover, we proposed a model for the electron transfer from ER to the final electron acceptor, fumarate under anaerobic conditions. Three Saccharomyces cerevisiae strains with varied amylase productions were selected at early glucose phase in batch fermentations for RNA extraction and hybridization on Affymetrix microarrays. Biological triplicates were applied, and strains with empty plasmid (no amylase productions) were used as control strain.
Project description:We developed a 'Virtual Northern' method, using DNA microarrays for genome-wide systematic analysis of mRNA lengths. We used this method to measure mRNAs corresponding to 84% of the annotated open reading frames (ORFs) in the S. cerevisiae genome, with high precision and accuracy (measurement errors 1 6-7%). We found a close linear relationship between mRNA lengths and the lengths of known or predicted translated sequences; mRNAs were typically around 300 nucleotides longer than the translated sequences. Analysis of genes deviating from that relationship identified ORFs with annotation errors, ORFs that appear not to be bona fide genes, and potentially novel genes. Interestingly, we found that systematic differences in the total length of the untranslated sequences in mRNAs were related to the functions of the encoded proteins. Groups of assays that are related as part of a time series. Keywords: time_series_design