Abstract
Although polyubiquitin chains linked through all lysines of ubiquitin exist, specific functions are well-established only for lysine-48 and lysine-63 linkages in Saccharomyces cerevisiae. To uncover pathways regulated by distinct linkages, genetic interactions between a gene deletion library and a panel of lysine-to-arginine ubiquitin mutants were systematically identified. The K11R mutant had strong genetic interactions with threonine biosynthetic genes. Consistently, we found that K11R mutants import threonine poorly. The K11R mutant also exhibited a strong genetic interaction with a subunit of the anaphase-promoting complex (APC), suggesting a role in cell cycle regulation. K11-linkages are important for vertebrate APC function, but this was not previously described in yeast. We show that the yeast APC also modifies substrates with K11-linkages in vitro, and that those chains contribute to normal APC-substrate turnover in vivo. This study reveals comprehensive genetic interactomes of polyubiquitin chains and characterizes the role of K11-chains in two biological pathways.
Highlights
The ubiquitination of proteins is a highly conserved posttranslational modification that can alter the stability, localization, and function of proteins (Finley et al, 2012)
A synthetic genetic array analysis (SGA) was undertaken to identify pathways that are regulated by specific polyubiquitin chain types (Tong et al, 2004)
Some large-scale proteomics studies have advanced our understanding of the physiological roles of specific polyubiquitin chain types, much of the progress has occurred through the study of single pathways
Summary
The ubiquitination of proteins is a highly conserved posttranslational modification that can alter the stability, localization, and function of proteins (Finley et al, 2012). Three enzyme classes mediate the covalent attachment of ubiquitin, a 76 amino acid protein, to target proteins: An E1 ubiquitin-activating enzyme activates ubiquitin for transfer to an E2 ubiquitin-conjugating enzyme, which in turn interacts with an E3 ubiquitin ligase to transfer ubiquitin to lysine residues on the target protein (Hershko and Ciechanover, 1998). This enzymatic cascade can transfer ubiquitin onto lysine residues of ubiquitin itself, thereby generating chains of polyubiquitin on target proteins (Hershko and Ciechanover, 1998). K48 and K11 are the most abundant ubiquitin linkage types—each accounting for about a third of ubiquitin linkages in yeast—while
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