Phenomic screen identifies a role for the yeast lysine acetyltransferase NuA4 in the control of Bcy1 subcellular localization, glycogen biosynthesis, and mitochondrial morphology

Abstract

Cellular metabolism is tightly regulated by many signaling pathways and processes, including lysine acetylation of proteins. While lysine acetylation of metabolic enzymes can directly influence enzyme activity, there is growing evidence that lysine acetylation can also impact protein localization. As the Saccharomyces cerevisiae lysine acetyltransferase complex NuA4 has been implicated in a variety of metabolic processes, we have explored whether NuA4 controls the localization and/or protein levels of metabolic proteins. We performed a high-throughput microscopy screen of over 360 GFP-tagged metabolic proteins and identified 23 proteins whose localization and/or abundance changed upon deletion of the NuA4 scaffolding subunit, EAF1. Within this, three proteins were required for glycogen synthesis and 14 proteins were associated with the mitochondria. We determined that in eaf1Δ cells the transcription of glycogen biosynthesis genes is upregulated resulting in increased proteins and glycogen production. Further, in the absence of EAF1, mitochondria are highly fused, increasing in volume approximately 3-fold, and are chaotically distributed but remain functional. Both the increased glycogen synthesis and mitochondrial elongation in eaf1Δ cells are dependent on Bcy1, the yeast regulatory subunit of PKA. Surprisingly, in the absence of EAF1, Bcy1 localization changes from being nuclear to cytoplasmic and PKA activity is altered. We found that NuA4-dependent localization of Bcy1 is dependent on a lysine residue at position 313 of Bcy1. However, the glycogen accumulation and mitochondrial elongation phenotypes of eaf1Δ, while dependent on Bcy1, were not fully dependent on Bcy1-K313 acetylation state and subcellular localization of Bcy1. As NuA4 is highly conserved with the human Tip60 complex, our work may inform human disease biology, revealing new avenues to investigate the role of Tip60 in metabolic diseases.

Cdk8 Kinase Module: A Mediator of Life and Death Decisions in Times of Stress

“…Similarly, yeast grown with a nonfermentable carbon source require increased OXPHOS activity that is accompanied by elongated mitochondrial networks [ 224 ]. This is also partially mediated through PKA activity [ 225 ] and supports the model that high OXPHOS activity correlates with extensive mitochondrial networks [ 226 ].…”

JAZF1: A metabolic actor subunit of the NuA4/TIP60 chromatin modifying complex

“…As a newly identified subunit of NuA4/TIP60 (Sudarshan et al, 2022), it is unclear if or how JAZF1 regulates enzymatic activity of the complex on or off chromatin. It is especially interesting that the role of NuA4/ TIP60 activity in transcription of metabolic genes and direct regulation of metabolic enzymes is well supported (van Beekum et al, 2008;Lin et al, 2009;Grönniger et al, 2010;Couture et al, 2012;Rossetto et al, 2014;Filteau et al, 2015;Jacquet et al, 2016;Dacquay et al, 2017;Rollins et al, 2017;Li et al, 2018;Cheng et al, 2019;Walden et al, 2020;Liu et al, 2021;Pham et al, 2022). Therefore, it is tempting to speculate that JAZF1 specifically modulates metabolism-related activity of the NuA4/ TIP60 complex.…”

“…Additionally, enzymatic activity of PCK1, the rate-limiting enzyme in gluconeogenesis, is positively controlled by Esa1/Tip60-mediated acetylation in yeast and human, supporting a role in glucose homeostasis (Lin et al, 2009). In parallel, disruption of yeast NuA4 leads to increased glycogen biosynthesis through direct regulation of protein kinase A (Filteau et al, 2015;Walden et al, 2020). Further, implication of the NuA4/ TIP60 complex in lipid homeostasis is clear, albeit somewhat controversial.…”

Global analysis of Saccharomyces cerevisiae growth in mucin

“…Once they reached log phase, cultures were washed twice in YP, and diluted to an OD 600 of 0.2 in YP. Dot assays were performed as described previously ( Walden et al 2020 ) with some modifications. Briefly, 3 μL of 10-fold serial dilutions (OD 600 = 0.2, 0.02, 0.002, 0.0002) were spotted on indicated media and incubated for 2–6 days at indicated temperatures.…”

“…Oxygen consumption rate (OCR) assays were performed with a Seahorse XFe96 instrument (Agilent) as described previously ( Walden et al 2020 ) with some modifications. Briefly, overnight cultures grown in YPD were washed in YP, reinoculated into 120 mL of YP or YPM to an OD 600 of 0.1, and incubated at 30° for 24 h. To make 2% ethanol supplemented YP (YPE) cultures, overnight strains were reinoculated into 50 mL of YPE to an OD 600 of 0.1 and incubated for 6 h on the day of the experiment.…”