Protein Acetylation Microarray Reveals that NuA4 Controls Key Metabolic Target Regulating Gluconeogenesis

Yang, Lin; Lu, Jin; Zhang, Junmei; Walter, Wendy; Dang, Weiwei; Wan, Jun; Tao, Sheng Ce; Qian, Jiang; Zhao, Yingming; Boeke, Jef D.; Berger, Shelley L.; Zhu, Heng

doi:10.1016/j.cell.2009.01.033

Cited by 290 publications

(351 citation statements)

References 73 publications

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“…Accordingly, the analyses were performed with K290Q RV and K290R RV. There are several examples in which mutation of a lysine to glutamine may function as an acetyl lysine mimetic while mutation to arginine may represent a constitutively unacetylated lysine (23,46,47). Contrary to these assumptions, our kinetic analyses of histone acetylation showed both K290Q RV and K290R RV exhibited an ϳ100-fold decrease in k cat (ϳ0.001 s Ϫ1 ) compared with RV (0.1 s Ϫ1 ) (Table 1).…”

mentioning

confidence: 46%

Autoacetylation of the Histone Acetyltransferase Rtt109

Albaugh

Arnold

Lee

et al. 2011

Journal of Biological Chemistry

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Rtt109 is a yeast histone acetyltransferase (HAT) that associates with histone chaperones Asf1 and Vps75 to acetylate H3K56, H3K9, and H3K27 and is important in DNA replication and maintaining genomic integrity. Recently, mass spectrometry and structural studies of Rtt109 have shown that active site residue Lys-290 is acetylated. However, the functional role of this modification and how the acetyl group is added to Lys-290 was unclear. Here, we examined the mechanism of Lys-290 acetylation and found that Rtt109 catalyzes intramolecular autoacetylation of Lys-290 ϳ200-times slower than H3 acetylation. Deacetylated Rtt109 was prepared by reacting with a sirtuin protein deacetylase, producing an enzyme with negligible HAT activity. Autoacetylation of Rtt109 restored full HAT activity, indicating that autoacetylation is necessary for HAT activity and is a fully reversible process. To dissect the mechanism of activation, biochemical, and kinetic analyses were performed with Lys-290 variants of the Rtt109-Vps75 complex. We found that autoacetylation of Lys-290 increases the binding affinity for acetyl-CoA and enhances the rate of acetyl-transfer onto histone substrates. This study represents the first detailed investigation of a HAT enzyme regulated by single-site intramolecular autoacetylation.Compaction of the eukaryotic genome is achieved by the packaging of DNA into chromatin. The fundamental unit of chromatin, the nucleosome, is composed of 147 base pairs of DNA wrapped around an octamer of histones containing two each of H2A, H2B, H3, and H4 (1). The covalent modification of histones, such as phosphorylation, acetylation, and methylation regulates DNA replication, repair, and transcription (2-6). Histone acetyltransferases (HATs), 2 a particular class of modification enzymes, catalyze the transfer of the acetyl group from acetyl-CoA onto the ⑀-amine group of lysines on histone substrates (7-9). There are three distinct HAT families that are classified according to their primary sequence and structural homology. These include the GNAT (Gcn5-related N-acetyltransferase), MYST (MOZ, Ybf2/Sas3, Sas2, and Tip60) and p300/CBP/Rtt109 families (7,9,10). Despite the divergence in sequence homology among the families, these HAT enzymes contain structurally similar core catalytic domains and utilize direct lysine substrate attack mechanisms (sequential) to carry out acetyl transfer (7,9 -11).Rtt109 (regulator of Ty1 transposition gene product 109) is a unique fungal-specific HAT enzyme (also named KAT11) that shares little sequence homology with other HAT enzymes. First identified to regulate the mobility of the Ty1 transposon element, Rtt109 HAT activities are important in a number of nuclear processes including DNA replication and maintaining genome integrity (12-17). Unique from other HAT enzymes, Rtt109 requires association with histone chaperones to direct substrate specificity and enhance catalysis (15, 18 -20). Histone chaperones bind histones to prevent unfavorable histone:DNA interactions and function in the assem...

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confidence: 46%

Autoacetylation of the Histone Acetyltransferase Rtt109

Albaugh

Arnold

Lee

et al. 2011

Journal of Biological Chemistry

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“…1C), suggesting that our approach can identify sirtuin target proteins. No peptides from sirtuin target Pck1 (18) were observed in our data sets. Although they did not meet the stringent twofold cut-off employed here, peptides containing Hst2-regulated Snf2 acetylation sites (19) were up-regulated an average of 1.7-fold in FIG.…”

Section: Resultsmentioning

confidence: 91%

“…Using a candidate approach, we previously identified the ribosomal protein gene transcription factor Ifh1 as a highly acetylated protein regulated both by Sir2 and Hst1 (1). Sir2 also mediates the deacetylation of the Pck1 enzyme to regulate gluconeogenesis (18). Finally, the acetylation of the Snf2 subunit of the SWI/SNF ATPase is increased slightly in the absence of HST2 (19).…”

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confidence: 99%

Acetylome Profiling Reveals Overlap in the Regulation of Diverse Processes by Sirtuins, Gcn5, and Esa1

Downey

Johnson

Davey

et al. 2015

Molecular & Cellular Proteomics

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Although histone acetylation and deacetylation machineries (HATs and HDACs) regulate important aspects of cell function by targeting histone tails, recent work highlights that non-histone protein acetylation is also pervasive in eukaryotes. Here, we use quantitative mass-spectrometry to define acetylations targeted by the sirtuin family, previously implicated in the regulation of non-histone protein acetylation. To identify HATs that promote acetylation of these sites, we also performed this analysis in gcn5 (SAGA) and esa1 (NuA4) mutants. We observed strong sequence specificity for the sirtuins and for each of these HATs. Although the Gcn5 and Esa1 consensus sequences are entirely distinct, the sirtuin consensus overlaps almost entirely with that of Gcn5, suggesting a strong coordination between these two regulatory enzymes. Furthermore, by examining global acetylation in an ada2 mutant, which dissociates Gcn5 from the SAGA complex, we found that a subset of Gcn5 targets did not depend on an intact SAGA complex for targeting. Our work provides a framework for understanding how HAT and HDAC enzymes collaborate to regulate critical cellular processes related to growth and division. Molecular & Cellular

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“…With conventional protein microarrays it is necessary to purify thousands of in vivo expressed proteins and to spot these purified proteins on a solid surface (5)(6)(7)(8). In contrast, in situ synthesis protein microarray technologies simplify protein microarray fabrication by circumventing the steps of in vivo protein expression and purification (9,10,14,15).…”

Section: Arabidopsis Thalianamentioning

confidence: 99%

Mapping transcription factor interactome networks using HaloTag protein arrays

Yazaki

Galli

Kim

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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Protein microarrays enable investigation of diverse biochemical properties for thousands of proteins in a single experiment, an unparalleled capacity. Using a high-density system called HaloTag nucleic acid programmable protein array (HaloTag-NAPPA), we created high-density protein arrays comprising 12,000 Arabidopsis ORFs. We used these arrays to query protein-protein interactions for a set of 38 transcription factors and transcriptional regulators (TFs) that function in diverse plant hormone regulatory pathways. The resulting transcription factor interactome network, TF-NAPPA, contains thousands of novel interactions. Validation in a benchmarked in vitro pull-down assay revealed that a random subset of TF-NAPPA validated at the same rate of 64% as a positive reference set of literaturecurated interactions. Moreover, using a bimolecular fluorescence complementation (BiFC) assay, we confirmed in planta several interactions of biological interest and determined the interaction localizations for seven pairs. The application of HaloTag-NAPPA technology to plant hormone signaling pathways allowed the identification of many novel transcription factor-protein interactions and led to the development of a proteome-wide plant hormone TF interactome network.protein arrays | interactome | hormone | systems biology | Arabidopsis thalianaA major objective in the postgenomic era is to assign detailed molecular function(s) to the many protein-coding genes that remain uncharacterized even in model organisms such as the reference plant Arabidopsis thaliana (1). It is estimated that Arabidopsis contains more than 2,000 transcription factors and transcriptional regulators (hereafter "TFs"), most of which are uncharacterized (2). TFs function as key players in plant hormone signal transduction pathways and are responsible for directing the widespread changes in gene expression that are essential for the regulation of growth and development in plants (3, 4). The TFs within these transcriptional regulatory networks do not work independently but rather undergo complex interactions with other proteins (2). Understanding how these TFs interact with other proteins will ultimately lead to a greater comprehension of biological systems.For determination of physical protein-protein interactions (PPIs), protein microarrays (5-10) are complementary to other PPI technologies such as the yeast two-hybrid system (Y2H) (11) and protein complex purification coupled with mass spectrometry (AP-MS) (12, 13). With conventional protein microarrays it is necessary to purify thousands of in vivo expressed proteins and to spot these purified proteins on a solid surface (5-8). In contrast, in situ synthesis protein microarray technologies simplify protein microarray fabrication by circumventing the steps of in vivo protein expression and purification (9,10,14,15). This streamlining facilitates an increase in the number of target genes that can be assayed, allowing for thousands of protein-encoding plasmids to be spotted at lower cost and in less time. Such ...

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Protein Acetylation Microarray Reveals that NuA4 Controls Key Metabolic Target Regulating Gluconeogenesis

Cited by 290 publications

References 73 publications

Autoacetylation of the Histone Acetyltransferase Rtt109

Autoacetylation of the Histone Acetyltransferase Rtt109

Acetylome Profiling Reveals Overlap in the Regulation of Diverse Processes by Sirtuins, Gcn5, and Esa1

Mapping transcription factor interactome networks using HaloTag protein arrays

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