Dataset Information

Purification of ubiquitylated proteins from HCT116 cells

ABSTRACT: By hyperosmotic stimuli, a slight increase in the level of ubiquitylated proteins was observed. So we tried to identify the di-Gly peptides.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Colon

SUBMITTER: Hikaru Tsuchiya

LAB HEAD: Yasushi Saeki

PROVIDER: PXD016369 | Pride | 2020-02-18

REPOSITORIES: Pride

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Publications

Stress- and ubiquitylation-dependent phase separation of the proteasome.

Yasuda Sayaka S Tsuchiya Hikaru H Kaiho Ai A Guo Qiang Q Ikeuchi Ken K Endo Akinori A Arai Naoko N Ohtake Fumiaki F Murata Shigeo S Inada Toshifumi T Baumeister Wolfgang W Fernández-Busnadiego Rubén R Tanaka Keiji K Saeki Yasushi Y

Nature 20200205 7794

The proteasome is a major proteolytic machine that regulates cellular proteostasis through selective degradation of ubiquitylated proteins<sup>1,2</sup>. A number of ubiquitin-related molecules have recently been found to be involved in the regulation of biomolecular condensates or membraneless organelles, which arise by liquid-liquid phase separation of specific biomolecules, including stress granules, nuclear speckles and autophagosomes<sup>3-8</sup>, but it remains unclear whether the proteas ...[more]

PMID: 32025036

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Project description:Myotonic dystrophy type 1 (DM1) is a neuromuscular disorder caused by a non-coding CTG repeat expansion in the DMPK gene. This mutation generates a toxic CUG RNA that interferes with the RNA processing of target genes in multiple tissues. Despite debilitating neurological impairment, the pathophysiological cascade of molecular and cellular events in the central nervous system has been less extensively characterized than the molecular pathogenesis of muscle/cardiac dysfunction. Particularly, the contribution of different cell types to DM1 brain disease is not clearly understood. We first used transcriptomics to compare the impact of expanded CUG RNA on the transcriptome of primary neurons, astrocytes and oligodendrocytes derived from DMSXL mice, a transgenic model of DM1. RNA sequencing revealed more frequent expression and splicing changes in glia than neuronal cells. In particular, primary DMSXL oligodendrocytes showed the highest number of transcripts differentially expressed, while DMSXL astrocytes displayed the most severe splicing dysregulation. Interestingly, the expression and splicing defects of DMSXL glia recreated molecular signatures suggestive of impaired cell differentiation: while DMSXL oligodendrocytes failed to upregulate a subset of genes that are naturally activated during the oligodendroglia differentiation, a significant proportion of missplicing events in DMSXL oligodendrocytes and astrocytes increased the expression of RNA isoforms typical of precursor cell stages. Together these data suggest that expanded CUG RNA in glial cells affects preferentially differentiation-regulated molecular events. This hypothesis was corroborated by gene ontology (GO) analyses, which revealed an enrichment for biological processes and cellular components with critical roles during cell differentiation. Finally, we combined exon ontology with phosphoproteomics and cell imaging to explore the functional impact of CUG-associated spliceopathy on downstream protein metabolism. Changes in phosphorylation, protein isoform expression and intracellular localization in DMSXL astrocytes demonstrate the far-reaching impact of the DM1 repeat expansion on cell metabolism. Our multi-omics approaches provide insight into the mechanisms of CUG RNA toxicity in the CNS with cell type resolution, and support the priority for future research on non-neuronal mechanisms and proteomic changes in DM1 brain disease.

Dataset Information

Purification of ubiquitylated proteins from HCT116 cells

Publications

Stress- and ubiquitylation-dependent phase separation of the proteasome.

Similar Datasets

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