Bypass suppression defines critical genomic regulation by the NuA4 acetyltransferase complex

The organization and regulation of eukaryotic DNA is a critical genomic process that is controlled by many key enzymes and multimeric complexes. One mode of regulation involves the acetylation of histones, as coordinated by the opposing enzymatic activities of histone acetyltransferases and deacetylases. Some of the enzymatic and regulatory subunits involved in chromatin regulation are essential, and thus present additional challenges to comprehensively study their function. Subunits within the NuA4 complex, such as the catalytic subunit Esa1, and non-catalytic Epl1 subunit, are examples of essential chromatin-modifying components. In this thesis, the powerful genetic tool of bypass suppression, is used to study null ESA1 and EPL1 mutants by concurrent deletion of the Rpd3L deacetylase complex, promoting a more balanced cellular acetylation state. Upon bypass, critical functions of Epl1 were defined for the first time in vivo, and implicated Epl1 as a key co-factor required for Esa1 catalytic activity. Here, Epl1 is also found to be important for targeting Esa1 to chromatin, thereby supporting Esa1’s critical activity as an acetyltransferase. Furthermore, bypass suppression of NuA4 revealed a key interaction network between NuA4 and the Rpd3L and Hda1 histone deacetylases. Despite deleting an essential acetyltransferase and two important deacetylases, growth of the epl1∆ sds3∆ hda1∆ mutant is significantly more robust than the epl1∆ sds3∆ mutant and can withstand various stresses such as high temperature and exposure to genotoxic agents. The strong genetic interaction between NuA4, Rpd3L, and Hda1 underscores the importance of understanding how acetyltransferases and deacetylases act both in opposition and cooperatively. Finally, bypass suppression of ESA1 enabled the first transcriptomics study of esa1∆ cells, defining Esa1 as a critical regulator of ribosome biogenesis. Together, the studies presented and discussed in this thesis highlight the power of bypass suppression and define key functions and regulatory targets of critical chromatin-modifiers in Saccharomyces cerevisiae