Uncovering the Role of the Yeast Lysine Acetyltransferase NuA4 in the Regulation of Nuclear Shape and Lipid Metabolism.

Here, we report a novel role for the yeast lysine acetyltransferase NuA4 in regulating phospholipid availability for organelle morphology. Disruption of the NuA4 complex results in 70% of cells displaying nuclear deformations and nearly 50% of cells exhibiting vacuolar fragmentation. Cells deficient in NuA4 also show severe defects in the formation of nuclear-vacuole junctions (NJV), as well as a decrease in piecemeal microautophagy of the nucleus (PMN). To determine the cause of these defects we focused on Pah1, an enzyme that converts phosphatidic acid into diacylglycerol, favoring accumulation of lipid droplets over phospholipids that are used for membrane expansion. NuA4 subunit Eaf1 was required for Pah1 localization to the inner nuclear membrane and artificially tethering of Pah1 to the nuclear membrane rescued nuclear deformation and vacuole fragmentation defects, but not defects related to the formation of NVJs. Mutation of a NuA4-dependent acetylation site on Pah1 also resulted in aberrant Pah1 localization and defects in nuclear morphology and NVJ. Our work suggests a critical role for NuA4 in organelle morphology that is partially mediated through the regulation of Pah1 subcellular localization.

Single-cell multiomics reveals ENL mutation perturbs kidney developmental trajectory by rewiring gene regulatory landscape.

How disruptions to normal cell differentiation link to tumorigenesis remains incompletely understood. Wilms tumor, an embryonal tumor associated with disrupted organogenesis, often harbors mutations in epigenetic regulators, but their role in kidney development remains unexplored. Here, we show at single-cell resolution that a Wilms tumor-associated mutation in the histone acetylation reader ENL disrupts kidney differentiation in mice by rewiring the gene regulatory landscape. Mutant ENL promotes nephron progenitor commitment while restricting their differentiation by dysregulating transcription factors such as Hox clusters. It also induces abnormal progenitors that lose kidney-associated chromatin identity. Furthermore, mutant ENL alters the transcriptome and chromatin accessibility of stromal progenitors, resulting in hyperactivation of Wnt signaling. The impacts of mutant ENL on both nephron and stroma lineages lead to profound kidney developmental defects and postnatal mortality in mice. Notably, a small molecule inhibiting mutant ENL's histone acetylation binding activity largely reverses these defects. This study provides insights into how mutations in epigenetic regulators disrupt kidney development and suggests a potential therapeutic approach.

Editing of endogenous tubulins reveals varying effects of tubulin posttranslational modifications on axonal growth and regeneration.

Tubulin posttranslational modifications (PTMs) modulate the dynamic properties of microtubules and their interactions with other proteins. However, the effects of tubulin PTMs were often revealed indirectly through the deletion of modifying enzymes or the overexpression of tubulin mutants. In this study, we directly edited the endogenous tubulin loci to install PTM-mimicking or -disabling mutations and studied their effects on microtubule stability, neurite outgrowth, axonal regeneration, cargo transport, and sensory functions in the touch receptor neurons of Caenorhabditis elegans. We found that the status of ß-tubulin S172 phosphorylation and K252 acetylation strongly affected microtubule dynamics, neurite growth, and regeneration, whereas α-tubulin K40 acetylation had little influence. Polyglutamylation and detyrosination in the tubulin C-terminal tail had more subtle effects on microtubule stability likely by modulating the interaction with kinesin-13. Overall, our study systematically assessed and compared several tubulin PTMs for their impacts on neuronal differentiation and regeneration and established an in vivo platform to test the function of tubulin PTMs in neurons.

Spermidine supplementation in honey bees: Autophagy and epigenetic modifications.

Polyamines (PAs), including putrescine (Put), spermidine (Spd), and spermine (Spm), are essential polycations with wide-ranging roles in cellular functions. PA levels decline with age, making exogenous PA supplementation, particularly Spd, an intriguing prospect. Previous research in honey bees demonstrated that millimolar Spd added to their diet increased lifespan and reinforced oxidative resilience. The present study is aimed to assess the anti-aging effects of spermidine supplementation at concentrations of 0.1 and 1 mM in honey bees, focusing on autophagy and associated epigenetic changes. Results showed a more pronounced effect at the lower Spd concentration, primarily in the abdomen. Spd induced site-specific histone 3 hypoacetylation at sites K18 and 27, hyperacetylation at K9, with no change at K14 in the entire body. Additionally, autophagy-related genes (ATG3, 5, 9, 13) and genes associated with epigenetic changes (HDAC1, HDAC3, SIRT1, KAT2A, KAT6B, P300, DNMT1A, DNMT1B) were upregulated in the abdomens of honey bees. In conclusion, our findings highlight profound epigenetic changes and autophagy promotion due to spermidine supplementation, contributing to increased honey bee longevity. Further research is needed to fully understand the precise mechanisms and the interplay between epigenetic alterations and autophagy in honey bees, underscoring the significance of autophagy as a geroprotective mechanism.

Polyamine Catabolism Revisited: Acetylpolyamine Oxidase Plays a Minor Role due to Low Expression.

Biogenic polyamines are ubiquitous compounds. Dysregulation of their metabolism is associated with the development of various pathologies, including cancer, hyperproliferative diseases, and infections. The canonical pathway of polyamine catabolism includes acetylation of spermine and spermidine and subsequent acetylpolyamine oxidase (PAOX)-mediated oxidation of acetylpolyamines (back-conversion) or their direct efflux from the cell. PAOX is considered to catalyze a non-rate-limiting catabolic step. Here, we show that PAOX transcription levels are extremely low in various tumor- and non-tumor cell lines and, in most cases, do not change in response to altered polyamine metabolism. Its enzymatic activity is undetectable in the majority of cell lines except for neuroblastoma and low passage glioblastoma cell lines. Treatment of A549 cells with N1,N11-diethylnorspermine leads to PAOX induction, but its contribution to polyamine catabolism remains moderate. We also describe two alternative enzyme isoforms and show that isoform 4 has diminished oxidase activity and isoform 2 is inactive. PAOX overexpression correlates with the resistance of cancer cells to genotoxic antitumor drugs, indicating that PAOX may be a useful therapeutic target. Finally, PAOX is dispensable for the replication of various viruses. These data suggest that a decrease in polyamine levels is achieved predominantly by the secretion of acetylated spermine and spermidine rather than by back-conversion.

N-terminal acetylation of Set1-COMPASS fine-tunes H3K4 methylation patterns.

H3K4 methylation by Set1-COMPASS (complex of proteins associated with Set1) is a conserved histone modification. Although it is critical for gene regulation, the posttranslational modifications of this complex that affect its function are largely unexplored. This study showed that N-terminal acetylation of Set1-COMPASS proteins by N-terminal acetyltransferases (NATs) can modulate H3K4 methylation patterns. Specifically, deleting NatA substantially decreased global H3K4me3 levels and caused the H3K4me2 peak in the 5' transcribed regions to shift to the promoters. NatA was required for N-terminal acetylation of three subunits of Set1-COMPASS: Shg1, Spp1, and Swd2. Moreover, deleting Shg1 or blocking its N-terminal acetylation via proline mutation of the target residue drastically reduced H3K4 methylation. Thus, NatA-mediated N-terminal acetylation of Shg1 shapes H3K4 methylation patterns. NatB also regulates H3K4 methylation, likely via N-terminal acetylation of the Set1-COMPASS protein Swd1. Thus, N-terminal acetylation of Set1-COMPASS proteins can directly fine-tune the functions of this complex, thereby substantially shaping H3K4 methylation patterns.

Comprehensive Proteome and Acetylome Analysis of Needle Senescence in Larix gmelinii.

Leaf senescence is essential for the growth and development of deciduous trees in the next season. Larix gmelinii, a deciduous coniferous tree, exhibits its most distinctive feature by turning yellow in the autumn and eventually shedding its leaves, resulting in significant changes in its appearance during the fall. Lysine acetylation plays an important role in diverse cellular processes; however, limited knowledge is available regarding acetylations in the needle senescence of L. gmelinii. In this study, the proteomics and acetylated modification omics of two phenotypic leaves, yellow and green (senescent and non-senescent) needles, were analyzed before autumn defoliation. In total, 5022 proteins and 4469 unique acetylation sites in 2414 lysine acylated proteins were identified, and this resulted in the discovery of 1335 differentially expressed proteins (DEPs) and 605 differentially expressed acetylated proteins (DAPs) in yellow versus green needles. There are significant differences between the proteome and acetylome; only 269 proteins were found to be DEP and DAP, of which 136 proteins were consistently expressed in both the DEP and DAP, 91 proteins were upregulated, and 45 proteins were down-regulated. The DEPs participate in the metabolism of starch and sucrose, while the DAPs are involved in glycolysis and the tricarboxylic acid cycle. Among them, DEPs underwent significant changes in glycolysis and citric acid cycling. Most of the enzymes involved in glycolysis and the citrate cycle were acetylated. DAPs were down-regulated in glycolysis and up-regulated in the citrate cycle. In all, the results of this study reveal the important role of lysine acetylation in the senescence of L. gmelinii needles and provide a new perspective for understanding the molecular mechanism of leaf senescence and tree seasonal growth.

Histone Deacetylases in Retinoblastoma.

Retinoblastoma, a pediatric ocular malignancy, presents significant challenges in comprehending its molecular underpinnings and targeted therapeutic approaches. The dysregulated activity of histone deacetylases (HDACs) has been associated with retinoblastoma pathogenesis, influencing critical cellular processes like cell cycle regulation or retinal ganglion cell apoptosis. Through their deacetylase activity, HDACs exert control over key tumor suppressors and oncogenes, influencing the delicate equilibrium between proliferation and cell death. Furthermore, the interplay between HDACs and the retinoblastoma protein pathway, a pivotal aspect of retinoblastoma etiology, reveals a complex network of interactions influencing the tumor microenvironment. The examination of HDAC inhibitors, encompassing both established and novel compounds, offers insights into potential approaches to restore acetylation balance and impede retinoblastoma progression. Moreover, the identification of specific HDAC isoforms exhibiting varying expression in retinoblastoma provides avenues for personalized therapeutic strategies, allowing for interventions tailored to individual patient profiles. This review focuses on the intricate interrelationship between HDACs and retinoblastoma, shedding light on epigenetic mechanisms that control tumor development and progression. The exploration of HDAC-targeted therapies underscores the potential for innovative treatment modalities in the pursuit of more efficacious and personalized management strategies for this disease.

Lactate modulates zygotic genome activation through H3K18 lactylation rather than H3K27 acetylation.

In spite of its essential role in culture media, the precise influence of lactate on early mouse embryonic development remains elusive. Previous studies have implicated lactate accumulation in medium affecting histone acetylation. Recent research has underscored lactate-derived histone lactylation as a novel epigenetic modification in diverse cellular processes and diseases. Our investigation demonstrated that the absence of sodium lactate in the medium resulted in a pronounced 2-cell arrest at the late G2 phase in embryos. RNA-seq analysis revealed that the absence of sodium lactate significantly impaired the maternal-to-zygotic transition (MZT), particularly in zygotic gene activation (ZGA). Investigations were conducted employing Cut&Tag assays targeting the well-studied histone acetylation and lactylation sites, H3K18la and H3K27ac, respectively. The findings revealed a noticeable reduction in H3K18la modification under lactate deficiency, and this alteration showed a significant correlation with changes in gene expression. In contrast, H3K27ac exhibited minimal correlation. These results suggest that lactate may preferentially influence early embryonic development through H3K18la rather than H3K27ac modifications.

Sirt1 Deficiency Promotes Age-Related AF Through Enhancing Atrial Necroptosis by Activation of RIPK1 Acetylation.

BACKGROUND: Aging is one of the most potent risk determinants for the onset of atrial fibrillation (AF). Sirts (sirtuins) have been implicated in the pathogenesis of cardiovascular disease, and their expression declines with aging. However, whether Sirts involved in age-related AF and its underlying mechanisms remain unknown. The present study aims to explore the role of Sirts in age-related AF and delineate the underlying molecular mechanisms. METHODS: Sirt1 levels in the atria of both elderly individuals and aging rats were evaluated using quantitative real-time polymerase chain reaction and Western blot analysis. Mice were engineered to specifically knockout Sirt1 in the atria and right ventricle (Sirt1mef2c/mef2c). Various techniques, such as echocardiography, atrial electrophysiology, and protein acetylation modification omics were employed. Additionally, coimmunoprecipitation was utilized to substantiate the interaction between Sirt1 and RIPK1 (receptor-interacting protein kinase 1). RESULTS: We discerned that among the diverse subtypes of sirtuin proteins, only Sirt1 expression was significantly diminished in the atria of elderly people and aged rats. The Sirt1mef2c/mef2c mice exhibited an enlarged atrial diameter and heightened vulnerability to AF. Acetylated proteomics and cell experiments identified that Sirt1 deficiency activated atrial necroptosis through increasing RIPK1 acetylation and subsequent pseudokinase MLKL (mixed lineage kinase domain-like protein) phosphorylation. Consistently, necroptotic inhibitor necrosulfonamide mitigated atrial necroptosis and diminished both the atrial diameter and AF susceptibility of Sirt1mef2c/mef2c mice. Resveratrol prevented age-related AF in rats by activating atrial Sirt1 and inhibiting necroptosis. CONCLUSIONS: Our findings first demonstrated that Sirt1 exerts significant efficacy in countering age-related AF by impeding atrial necroptosis through regulation of RIPK1 acetylation, highlighting that the activation of Sirt1 or the inhibition of necroptosis could potentially serve as a therapeutic strategy for age-related AF.

The role of NAD-dependent deacetylase sirtuin-2 in liver metabolic stress through regulating pyruvate kinase M2 ubiquitination.

NAD-dependent deacetylase Sirt2 is involved in mammalian metabolic activities, matching energy demand with energy production and expenditure, and is relevant to a variety of metabolic diseases. Here, we constructed Sirt2 knockout and adeno-associated virus overexpression mice and found that deletion of hepatic Sirt2 accelerated primary obesity and insulin resistance in mice with concomitant hepatic metabolic dysfunction. However, the key targets of Sirt2 are unknown. We identified the M2 isoform of pyruvate kinase (PKM2) as a key Sirt2 target involved in glycolysis in metabolic stress. Through yeast two-hybrid and mass spectrometry combined with multi-omics analysis, we identified candidate acetylation modification targets of Sirt2 on PKM2 lysine 135 (K135). The Sirt2-mediated deacetylation-ubiquitination switch of PKM2 regulated the development of glycolysis. Here, we found that Sirt2 deficiency led to impaired glucose tolerance and insulin resistance and induced primary obesity. Sirt2 severely disrupted liver function in mice under metabolic stress, exacerbated the metabolic burden on the liver, and affected glucose metabolism. Sirt2 underwent acetylation modification of lysine 135 of PKM2 through a histidine 187 enzyme active site-dependent effect and reduced ubiquitination of the K48 ubiquitin chain of PKM2. Our findings reveal that the hepatic glucose metabolism links nutrient state to whole-body energetics through the rhythmic regulation of Sirt2.

Epigenetic-based differentiation therapy for Acute Myeloid Leukemia.

Despite the development of novel therapies for acute myeloid leukemia, outcomes remain poor for most patients, and therapeutic improvements are an urgent unmet need. Although treatment regimens promoting differentiation have succeeded in the treatment of acute promyelocytic leukemia, their role in other acute myeloid leukemia subtypes needs to be explored. Here we identify and characterize two lysine deacetylase inhibitors, CM-444 and CM-1758, exhibiting the capacity to promote myeloid differentiation in all acute myeloid leukemia subtypes at low non-cytotoxic doses, unlike other commercial histone deacetylase inhibitors. Analyzing the acetylome after CM-444 and CM-1758 treatment reveals modulation of non-histone proteins involved in the enhancer-promoter chromatin regulatory complex, including bromodomain proteins. This acetylation is essential for enhancing the expression of key transcription factors directly involved in the differentiation therapy induced by CM-444/CM-1758 in acute myeloid leukemia. In summary, these compounds may represent effective differentiation-based therapeutic agents across acute myeloid leukemia subtypes with a potential mechanism for the treatment of acute myeloid leukemia.

Myeloid-specific Hdac10 deletion protects against LPS-induced acute lung injury via P62 acetylation at lysine 165.

BACKGROUND: Aberrant activation of macrophages is associated with pathogenesis of acute lung injury (ALI). However, the potential pathogenesis has not been explored. OBJECTIVES: We aimed to identify whether histone deacetylase (HDAC) 10 is involved in lipopolysaccharide (LPS)-exposed ALI and reveal the underlying pathogenesis by which it promotes lung inflammation in LPS-exposed ALI via modifying P62 with deacetylation. METHODS: We constructed an ALI mice model stimulated with LPS to determine the positive effect of Hdac10 deficiency. Moreover, we cultured murine alveolar macrophage cell line (MH-S cells) and primary bone marrow-derived macrophages (BMDMs) to explore the pro-inflammatory activity and mechanism of HDAC10 after LPS challenge. RESULTS: HDAC10 expression was increased both in mice lung tissues and macrophage cell lines and promoted inflammatory cytokines production exposed to LPS. Hdac10 deficiency inhibited autophagy and inflammatory response after LPS stimulation. In vivo, Hdac10fl/fl-LysMCre mice considerably attenuated lung inflammation and inflammatory cytokines release exposed to LPS. Mechanistically, HDAC10 interacts with P62 and mediates P62 deacetylation at lysine 165 (K165), by which it promotes P62 expression and increases inflammatory cytokines production. Importantly, we identified that Salvianolic acid B (SAB), an HDAC10 inhibitor, reduces lung inflammatory response in LPS-stimulated ALI. CONCLUSION: These results uncover a previously unknown role for HDAC10 in regulating P62 deacetylation and aggravating lung inflammation in LPS-induced ALI, implicating that targeting HDAC10 is an effective therapy for LPS-exposed ALI.

Unconventional posttranslational modification in innate immunity.

Pattern recognition receptors (PRRs) play a crucial role in innate immunity, and a complex network tightly controls their signaling cascades to maintain immune homeostasis. Within the modification network, posttranslational modifications (PTMs) are at the core of signaling cascades. Conventional PTMs, which include phosphorylation and ubiquitination, have been extensively studied. The regulatory role of unconventional PTMs, involving unanchored ubiquitination, ISGylation, SUMOylation, NEDDylation, methylation, acetylation, palmitoylation, glycosylation, and myristylation, in the modulation of innate immune signaling pathways has been increasingly investigated. This comprehensive review delves into the emerging field of unconventional PTMs and highlights their pivotal role in innate immunity.

Acetyl-CoA synthetase activity is enzymatically regulated by lysine acetylation using acetyl-CoA or acetyl-phosphate as donor molecule.

The AMP-forming acetyl-CoA synthetase is regulated by lysine acetylation both in bacteria and eukaryotes. However, the underlying mechanism is poorly understood. The Bacillus subtilis acetyltransferase AcuA and the AMP-forming acetyl-CoA synthetase AcsA form an AcuA•AcsA complex, dissociating upon lysine acetylation of AcsA by AcuA. Crystal structures of AcsA from Chloroflexota bacterium in the apo form and in complex with acetyl-adenosine-5'-monophosphate (acetyl-AMP) support the flexible C-terminal domain adopting different conformations. AlphaFold2 predictions suggest binding of AcuA stabilizes AcsA in an undescribed conformation. We show the AcuA•AcsA complex dissociates upon acetyl-coenzyme A (acetyl-CoA) dependent acetylation of AcsA by AcuA. We discover an intrinsic phosphotransacetylase activity enabling AcuA•AcsA generating acetyl-CoA from acetyl-phosphate (AcP) and coenzyme A (CoA) used by AcuA to acetylate and inactivate AcsA. Here, we provide mechanistic insights into the regulation of AMP-forming acetyl-CoA synthetases by lysine acetylation and discover an intrinsic phosphotransacetylase allowing modulation of its activity based on AcP and CoA levels.

DCAF15 control of cohesin dynamics sustains acute myeloid leukemia.

The CRL4-DCAF15 E3 ubiquitin ligase complex is targeted by the aryl-sulfonamide molecular glues, leading to neo-substrate recruitment, ubiquitination, and proteasomal degradation. However, the physiological function of DCAF15 remains unknown. Using a domain-focused genetic screening approach, we reveal DCAF15 as an acute myeloid leukemia (AML)-biased dependency. Loss of DCAF15 results in suppression of AML through compromised replication fork integrity and consequent accumulation of DNA damage. Accordingly, DCAF15 loss sensitizes AML to replication stress-inducing therapeutics. Mechanistically, we discover that DCAF15 directly interacts with the SMC1A protein of the cohesin complex and destabilizes the cohesin regulatory factors PDS5A and CDCA5. Loss of PDS5A and CDCA5 removal precludes cohesin acetylation on chromatin, resulting in uncontrolled chromatin loop extrusion, defective DNA replication, and apoptosis. Collectively, our findings uncover an endogenous, cell autonomous function of DCAF15 in sustaining AML proliferation through post-translational control of cohesin dynamics.

Acetylation model predicts prognosis of patients and affects immune microenvironment infiltration in epithelial ovarian carcinoma.

BACKGROUND: Epithelial ovarian carcinoma (EOC) is a prevalent gynaecological malignancy. The prognosis of patients with EOC is related to acetylation modifications and immune responses in the tumour microenvironment (TME). However, the relationships between acetylation-related genes, patient prognosis, and the tumour immune microenvironment (TIME) are not yet understood. Our research aims to investigate the link between acetylation and the tumour microenvironment, with the goal of identifying new biomarkers for estimating survival of patients with EOC. METHODS: Using data downloaded from the tumour genome atlas (TCGA), genotypic tissue expression (GTEx), and gene expression master table (GEO), we comprehensively evaluated acetylation-related genes in 375 ovarian cancer specimens and identified molecular subtypes using unsupervised clustering. The prognosis, TIME, stem cell index and functional concentration analysis were compared among the three groups. A risk model based on differential expression of acetylation-related genes was established through minimum absolute contraction and selection operator (LASSO) regression analysis, and the predictive validity of this feature was validated using GEO data sets. A nomogram is used to predict a patient's likelihood of survival. In addition, different EOC risk groups were evaluated for timing, tumour immune dysfunction and exclusion (TIDE) score, stemness index, somatic mutation, and drug sensitivity. RESULTS: We used the mRNA levels of the differentially expressed genes related to acetylation to classify them into three distinct clusters. Patients with increased immune cell infiltration and lower stemness scores in cluster 2 (C2) exhibited poorer prognosis. Immunity and tumourigenesis-related pathways were highly abundant in cluster 3 (C3). We developed a prognostic model for ten differentially expressed acetylation-related genes. Kaplan-Meier analysis demonstrated significantly worse overall survival (OS) in high-risk patients. Furthermore, the TIME, tumour immune dysfunction and exclusion (TIDE) score, stemness index, tumour mutation burden (TMB), immunotherapy response, and drug sensitivity all showed significant correlations with the risk scores. CONCLUSIONS: Our study demonstrated a complex regulatory mechanism of acetylation in EOC. The assessment of acetylation patterns could provide new therapeutic strategies for EOC immunotherapy to improve the prognosis of patients.

SAHA/5-AZA Enhances Acetylation and Degradation of mutp53, Upregulates p21 and Downregulates c-Myc and BRCA-1 in Pancreatic Cancer Cells.

Epigenetic changes are common in cancer and include aberrant DNA methylation and histone modifications, including both acetylation or methylation. DNA methylation in the promoter regions and histone deacetylation are usually accompanied by gene silencing, and may lead to the suppression of tumor suppressors in cancer cells. An interaction between epigenetic pathways has been reported that could be exploited to more efficiently target aggressive cancer cells, particularly those against which current treatments usually fail, such as pancreatic cancer. In this study, we explored the possibility to combine the DNA demethylating agent 5-AZA with HDAC inhibitor SAHA to treat pancreatic cancer cell lines, focusing on the acetylation of mutp53 and the consequences on its stability, as well as on the interaction of this protein with c-myc and BRCA-1, key molecules in cancer survival. The results obtained suggest that SAHA/5-AZA combination was more effective than single treatments to promote the degradation of mutp53, to upregulate p21 and downregulate c-Myc and BRCA-1, thus increasing DNA damage and cytotoxicity in pancreatic cancer cells.

Bacterial protein acetylation: mechanisms, functions, and methods for study.

Lysine acetylation is an evolutionarily conserved protein modification that changes protein functions and plays an essential role in many cellular processes, such as central metabolism, transcriptional regulation, chemotaxis, and pathogen virulence. It can alter DNA binding, enzymatic activity, protein-protein interactions, protein stability, or protein localization. In prokaryotes, lysine acetylation occurs non-enzymatically and by the action of lysine acetyltransferases (KAT). In enzymatic acetylation, KAT transfers the acetyl group from acetyl-CoA (AcCoA) to the lysine side chain. In contrast, acetyl phosphate (AcP) is the acetyl donor of chemical acetylation. Regardless of the acetylation type, the removal of acetyl groups from acetyl lysines occurs only enzymatically by lysine deacetylases (KDAC). KATs are grouped into three main superfamilies based on their catalytic domain sequences and biochemical characteristics of catalysis. Specifically, members of the GNAT are found in eukaryotes and prokaryotes and have a core structural domain architecture. These enzymes can acetylate small molecules, metabolites, peptides, and proteins. This review presents current knowledge of acetylation mechanisms and functional implications in bacterial metabolism, pathogenicity, stress response, translation, and the emerging topic of protein acetylation in the gut microbiome. Additionally, the methods used to elucidate the biological significance of acetylation in bacteria, such as relative quantification and stoichiometry quantification, and the genetic code expansion tool (CGE), are reviewed.