An adaptive stress response that confers cellular resilience to decreased ubiquitination

Ubiquitination is a post-translational modification that regulates protein function and turnover. E2 ubiquitin-conjugating enzymes are key for ubiquitination but it remains uncharted what fraction of the proteome is E2-modulated. Here, we utilized deep-coverage TMT mass spectrometry to determine how protein abundance is modulated by E2s. Analysis of human cells with ~25% reduction in ubiquitination and with individual E2 knockdown indicates that 48% of the proteome is modulated by partial E2 loss, and that this rewires organelle and metabolic functions by reducing the turnover of E2-sensitive targets. In particular, several peroxins are upregulated by UBA1/E2 RNAi, and this promotes peroxisomal protein import and rewires cellular metabolism. Altogether, this study provides a framework for understanding how moderate reduction in E2 function rewires the proteome and cellular function. 116 RNA-seq samples from human HEK293T cells treated with siRNAs targetingUBA1 and E2 ubiquitin-conjugating enzymes compared to control treatment with non-targeting (NT) siRNAs