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Scientific Reports Apr 2023The majority of proteins in mammalian cells are modified by covalent attachment of an acetyl-group to the N-terminus (Nt-acetylation). Paradoxically, Nt-acetylation has...
The majority of proteins in mammalian cells are modified by covalent attachment of an acetyl-group to the N-terminus (Nt-acetylation). Paradoxically, Nt-acetylation has been suggested to inhibit as well as to promote substrate degradation. Contrasting these findings, proteome-wide stability measurements failed to detect any correlation between Nt-acetylation status and protein stability. Accordingly, by analysis of protein stability datasets, we found that predicted Nt-acetylation positively correlates with protein stability in case of GFP, but this correlation does not hold for the entire proteome. To further resolve this conundrum, we systematically changed the Nt-acetylation and ubiquitination status of model substrates and assessed their stability. For wild-type Bcl-B, which is heavily modified by proteasome-targeting lysine ubiquitination, Nt-acetylation did not correlate with protein stability. For a lysine-less Bcl-B mutant, however, Nt-acetylation correlated with increased protein stability, likely due to prohibition of ubiquitin conjugation to the acetylated N-terminus. In case of GFP, Nt-acetylation correlated with increased protein stability, as predicted, but our data suggest that Nt-acetylation does not affect GFP ubiquitination. Similarly, in case of the naturally lysine-less protein p16, Nt-acetylation correlated with protein stability, regardless of ubiquitination on its N-terminus or on an introduced lysine residue. A direct effect of Nt-acetylation on p16 stability was supported by studies in NatB-deficient cells. Together, our studies argue that Nt-acetylation can stabilize proteins in human cells in a substrate-specific manner, by competition with N-terminal ubiquitination, but also by other mechanisms that are independent of protein ubiquitination status.
Topics: Animals; Humans; Lysine; Proteome; Acetylation; Protein Processing, Post-Translational; Ubiquitination; Mammals
PubMed: 37005459
DOI: 10.1038/s41598-023-32380-3 -
Journal of Proteome Research Jan 2021Acetylation was initially discovered as a post-translational modification (PTM) on the unstructured, highly basic N-terminal tails of eukaryotic histones in the 1960s.... (Review)
Review
Acetylation was initially discovered as a post-translational modification (PTM) on the unstructured, highly basic N-terminal tails of eukaryotic histones in the 1960s. Histone acetylation constitutes part of the "histone code", which regulates chromosome compaction and various DNA processes such as gene expression, recombination, and DNA replication. In bacteria, nucleoid-associated proteins (NAPs) are responsible these functions in that they organize and compact the chromosome and regulate some DNA processes. The highly conserved DNABII family of proteins are considered functional homologues of eukaryotic histones despite having no sequence or structural conservation. Within the past decade, a growing interest in N-lysine acetylation led to the discovery that hundreds of bacterial proteins are acetylated with diverse cellular functions, in direct contrast to the original thought that this was a rare phenomenon. Similarly, other previously undiscovered bacterial PTMs, like serine, threonine, and tyrosine phosphorylation, have also been characterized. In this review, the various PTMs that were discovered among DNABII family proteins, specifically histone-like protein (HU) orthologues, from large-scale proteomic studies are discussed. The functional significance of these modifications and the enzymes involved are also addressed. The discovery of novel PTMs on these proteins begs this question: is there a histone-like code in bacteria?
Topics: Acetylation; Bacteria; Histone Code; Histones; Protein Processing, Post-Translational; Proteomics
PubMed: 32962352
DOI: 10.1021/acs.jproteome.0c00442 -
PloS One 2014The emerging view of Nε-lysine acetylation in eukaryotes is of a relatively abundant post-translational modification (PTM) that has a major impact on the function,...
The emerging view of Nε-lysine acetylation in eukaryotes is of a relatively abundant post-translational modification (PTM) that has a major impact on the function, structure, stability and/or location of thousands of proteins involved in diverse cellular processes. This PTM is typically considered to arise by the donation of the acetyl group from acetyl-coenzyme A (acCoA) to the ε-amino group of a lysine residue that is reversibly catalyzed by lysine acetyltransferases and deacetylases. Here, we provide genetic, mass spectrometric, biochemical and structural evidence that Nε-lysine acetylation is an equally abundant and important PTM in bacteria. Applying a recently developed, label-free and global mass spectrometric approach to an isogenic set of mutants, we detected acetylation of thousands of lysine residues on hundreds of Escherichia coli proteins that participate in diverse and often essential cellular processes, including translation, transcription and central metabolism. Many of these acetylations were regulated in an acetyl phosphate (acP)-dependent manner, providing compelling evidence for a recently reported mechanism of bacterial Nε-lysine acetylation. These mass spectrometric data, coupled with observations made by crystallography, biochemistry, and additional mass spectrometry showed that this acP-dependent acetylation is both non-enzymatic and specific, with specificity determined by the accessibility, reactivity and three-dimensional microenvironment of the target lysine. Crystallographic evidence shows acP can bind to proteins in active sites and cofactor binding sites, but also potentially anywhere molecules with a phosphate moiety could bind. Finally, we provide evidence that acP-dependent acetylation can impact the function of critical enzymes, including glyceraldehyde-3-phosphate dehydrogenase, triosephosphate isomerase, and RNA polymerase.
Topics: Acetylation; Amino Acid Sequence; Binding Sites; Blotting, Western; Crystallography, X-Ray; Escherichia coli; Escherichia coli Proteins; Glucose; Kinetics; Lysine; Mass Spectrometry; Molecular Sequence Data; Mutant Proteins; Organophosphates; Proteomics; Staining and Labeling
PubMed: 24756028
DOI: 10.1371/journal.pone.0094816 -
The Indian Journal of Medical Research Jul 2008Acetylation is one of the most important post-translational modification of proteins determining the structure, function and intracellular localization that plays an... (Review)
Review
Acetylation is one of the most important post-translational modification of proteins determining the structure, function and intracellular localization that plays an important role in the signal transduction pathways related to diverse cell functions, both during unstimulated and stress conditions. Protein acetylation in cells is regulated by a co-ordinated action of histone acetyl transferases (HAT) and histone deacetylases(HDAC) that ensures the maintenance of homeostasis and execution of activities related to damage response viz. DNA repair, cell cycle delay, apoptosis and senescence. Since inhibition of histone deacetylation, stalls the progress of many nuclear events including proliferation and damage response events on the one hand and the levels of deacetylases are elevated in many tumours on the other. Histone deacetylase has been among the targets for the development of anticancer drugs and adjuvant. The recent observation showing acetylation of proteins by calreticulin (an endoplasmic reticulum resident protein) with a high efficiency when polyphenolic acetates are the acetyl group donating molecules and acetyl CoA as weak substrate extends the realm of protein acetylation beyond HAT/HDAC combination. Elucidation of the relative roles of HAT/HDAC mediated acetylation viz. a calreticulin mediated acetylation in cell function under a variety of stress conditions would hold key to the design of drugs targeting protein acetylation system.
Topics: Acetylation; Antineoplastic Agents; Histones; Humans; Neoplasms; Signal Transduction
PubMed: 18820353
DOI: No ID Found -
Acetylation of Lactate Dehydrogenase Negatively Regulates the Acidogenicity of Streptococcus mutans.MBio Oct 2022Lysine acetylation, a ubiquitous and dynamic regulatory posttranslational modification (PTM), affects hundreds of proteins across all domains of life. In bacteria,...
Lysine acetylation, a ubiquitous and dynamic regulatory posttranslational modification (PTM), affects hundreds of proteins across all domains of life. In bacteria, lysine acetylation can be found in many essential pathways, and it is also crucial for bacterial virulence. However, the biological significance of lysine acetylation events to bacterial virulence factors remains poorly characterized. In Streptococcus mutans, the acetylome profiles help identify several lysine acetylation sites of lactate dehydrogenase (LDH), which catalyzes the conversion of pyruvate to lactic acid, causing the deterioration of teeth. We investigated the regulatory mechanism of LDH acetylation and characterized the effect of LDH acetylation on its function. We overexpressed the 15 Gcn5 -acetyltransferases (GNAT) family members in S. mutans and showed that the acetyltransferase ActA impaired its acidogenicity by acetylating LDH. Additionally, enzymatic acetyltransferase reactions demonstrated that purified ActA could acetylate LDH , and 10 potential lysine acetylation sites of LDH were identified by mass spectrometry, 70% of which were also detected . We further demonstrated that the lysine acetylation of LDH inhibited its enzymatic activity, and a subsequent rat caries model showed that ActA impaired the cariogenicity of S. mutans. Collectively, we demonstrated that ActA, the first identified and characterized acetyltransferase in S. mutans, acetylated the LDH enzymatically and inhibited its enzymatic activity, thereby providing a starting point for the further analysis of the biological significance of lysine acetylation in the virulence of S. mutans. Lysine acetylation, a dynamic regulatory posttranslational modification, remains poorly characterized in bacteria. Hundreds of proteins have been identified to be acetylated in bacteria, with advances made in acetylome analyses. However, the regulatory mechanisms and functional significance of the majority of these acetylated proteins remain unclear. We analyzed the acetylome profiles of Streptococcus mutans and found that lactate dehydrogenase (LDH) contains several lysine acetylation sites. We also demonstrated that the acetyltransferase ActA, a member of the Gcn5 -acetyltransferases (GNAT) family in S. mutans, acetylated LDH, inhibited its enzymatic ability to catalyze the conversion of pyruvate to lactic acid, and impaired its cariogenicity in a rat caries model. Therefore, LDH acetylation might be a potential target that can be exploited in the design of novel therapeutics to prevent dental caries.
Topics: Rats; Animals; Acetylation; Streptococcus mutans; Lysine; L-Lactate Dehydrogenase; Dental Caries; Protein Processing, Post-Translational; Virulence Factors; Acetyltransferases; Lactic Acid; Pyruvates
PubMed: 36043788
DOI: 10.1128/mbio.02013-22 -
Science Advances Apr 2020About 80% of human proteins are amino-terminally acetylated (Nt-acetylated) by one of seven Nt-acetyltransferases (NATs). Actin, the most abundant protein in the...
About 80% of human proteins are amino-terminally acetylated (Nt-acetylated) by one of seven Nt-acetyltransferases (NATs). Actin, the most abundant protein in the cytoplasm, has its own dedicated NAT, NAA80, which acts posttranslationally and affects cytoskeleton assembly and cell motility. Here, we show that NAA80 does not associate with filamentous actin in cells, and its natural substrate is the monomeric actin-profilin complex, consistent with Nt-acetylation preceding polymerization. NAA80 Nt-acetylates actin-profilin much more efficiently than actin alone, suggesting that profilin acts as a chaperone for actin Nt-acetylation. We determined crystal structures of the NAA80-actin-profilin ternary complex, representing different actin isoforms and different states of the catalytic reaction and revealing the first structure of NAT-substrate complex at atomic resolution. The structural, biochemical, and cellular analysis of mutants shows how NAA80 has evolved to specifically recognize actin among all cellular proteins while targeting all six actin isoforms, which differ the most at the amino terminus.
Topics: Acetylation; Acetyltransferases; Actins; Amino Acid Sequence; Binding Sites; Fluorescent Antibody Technique; Humans; Models, Molecular; Molecular Conformation; Profilins; Protein Binding; Protein Domains; Protein Isoforms; Protein Multimerization; Structure-Activity Relationship; Substrate Specificity
PubMed: 32284999
DOI: 10.1126/sciadv.aay8793 -
Genes Feb 2021Acetylation on lysine 56 of histone H3 of the yeast has been implicated in many cellular processes that affect genome stability. Despite being the object of much... (Review)
Review
Acetylation on lysine 56 of histone H3 of the yeast has been implicated in many cellular processes that affect genome stability. Despite being the object of much research, the complete scope of the roles played by K56 acetylation is not fully understood even today. The acetylation is put in place at the S-phase of the cell cycle, in order to flag newly synthesized histones that are incorporated during DNA replication. The signal is removed by two redundant deacetylases, Hst3 and Hst4, at the entry to G2/M phase. Its crucial location, at the entry and exit points of the DNA into and out of the nucleosome, makes this a central modification, and dictates that if acetylation and deacetylation are not well concerted and executed in a timely fashion, severe genomic instability arises. In this review, we explore the wealth of information available on the many roles played by H3K56 acetylation and the deacetylases Hst3 and Hst4 in DNA replication and repair.
Topics: Acetylation; DNA Repair; DNA Replication; Genomic Instability; Histone Deacetylases; Histones; S Phase; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 33668997
DOI: 10.3390/genes12030342 -
The EMBO Journal Mar 2000The fact that histones are modified by acetylation has been known for almost 30 years. The recent identification of enzymes that regulate histone acetylation has... (Review)
Review
The fact that histones are modified by acetylation has been known for almost 30 years. The recent identification of enzymes that regulate histone acetylation has revealed a broader use of this modification than was suspected previously. Acetylases are now known to modify a variety of proteins, including transcription factors, nuclear import factors and alpha-tubulin. Acetylation regulates many diverse functions, including DNA recognition, protein-protein interaction and protein stability. There is even a conserved structure, the bromodomain, that recognizes acetylated residues and may serve as a signalling domain. If you think all this sounds familiar, it should be. These are features characteristic of kinases. So, is acetylation a modification analogous to phosphorylation? This review sets out what we know about the broader substrate specificity and regulation of acetyl- ases and goes on to compare acetylation with the process of phosphorylation.
Topics: Acetylation; Acetylesterase; Acetyltransferases; Amidohydrolases; Histones; Phosphorylation; Proteins; Substrate Specificity
PubMed: 10716917
DOI: 10.1093/emboj/19.6.1176 -
Molecules and Cells Mar 2016Although Nα-terminal acetylation (Nt-acetylation) is a pervasive protein modification in eukaryotes, its general functions in a majority of proteins are poorly... (Review)
Review
Although Nα-terminal acetylation (Nt-acetylation) is a pervasive protein modification in eukaryotes, its general functions in a majority of proteins are poorly understood. In 2010, it was discovered that Nt-acetylation creates a specific protein degradation signal that is targeted by a new class of the N-end rule proteolytic system, called the Ac/N-end rule pathway. Here, we review recent advances in our understanding of the mechanism and biological functions of the Ac/N-end rule pathway, and its crosstalk with the Arg/N-end rule pathway (the classical N-end rule pathway).
Topics: Acetylation; Humans; Protein Processing, Post-Translational; Proteins; Proteolysis; Signal Transduction
PubMed: 26883906
DOI: 10.14348/molcells.2016.2329 -
Current Biology : CB Sep 2013In eukaryotes, ribosome biosynthesis involves the coordination of ribosomal RNA and ribosomal protein (RP) production. In S. cerevisiae, the regulation of ribosome...
BACKGROUND
In eukaryotes, ribosome biosynthesis involves the coordination of ribosomal RNA and ribosomal protein (RP) production. In S. cerevisiae, the regulation of ribosome biosynthesis occurs largely at the level of transcription. The transcription factor Ifh1 binds at RP genes and promotes their transcription when growth conditions are favorable. Although Ifh1 recruitment to RP genes has been characterized, little is known about the regulation of promoter-bound Ifh1.
RESULTS
We used a novel whole-cell-extract screening approach to identify Spt7, a member of the SAGA transcription complex, and the RP transactivator Ifh1 as highly acetylated nonhistone species. We report that Ifh1 is modified by acetylation specifically in an N-terminal domain. These acetylations require the Gcn5 histone acetyltransferase and are reversed by the sirtuin deacetylases Hst1 and Sir2. Ifh1 acetylation is regulated by rapamycin treatment and stress and limits the ability of Ifh1 to act as a transactivator at RP genes.
CONCLUSIONS
Our data suggest a novel mechanism of regulation whereby Gcn5 functions to titrate the activity of Ifh1 following its recruitment to RP promoters to provide more than an all-or-nothing mode of transcriptional regulation. We provide insights into how the action of histone acetylation machineries converges with nutrient-sensing pathways to regulate important aspects of cell growth.
Topics: Acetylation; Histone Acetyltransferases; Ribosomal Proteins; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins; Sirtuins; Trans-Activators; Transcription Factors
PubMed: 23973296
DOI: 10.1016/j.cub.2013.06.050