Low Dose Proteasome Inhibition Affects Alternative Splicing

Protein degradation by the ubiquitin proteasome system ensures controlled degradation of structural proteins, signaling mediators, and transcription factors. Inhibition of proteasome function by specific proteasome inhibitors results in dose-dependent cellular effects ranging from induction of apoptosis to protective stress responses. The present study seeks to identify nuclear regulators mediating the protective stress response to low dose proteasome inhibition. Primary human endothelial cells were treated with low doses of the proteasome inhibitor MG132 for 2 h, and proteomic analysis of nuclear extracts was performed. Using a 2-D differential in gel electrophoresis (DIGE) approach, we identified more than 24 splice factors to be differentially regulated by low dose proteasome inhibition. In particular, several isoforms of hnRNPA1 were shown to be increased, pointing toward altered posttranslational modification of hnRNPA1 upon proteasome inhibition. Elevated levels of splice factors were associated with a different alternative splicing pattern in response to proteasome inhibition as determined by Affymetrix exon array profiling. Of note, we observed alternative RNA processing for stress associated genes such as caspases and heat shock proteins. Our study provides first evidence that low dose proteasome inhibition affects posttranscriptional regulation of splice factors and early alternative splicing events.