Family Histone Acetyltransferase Research Articles

KAT8-mediated H4K16ac is essential for sustaining trophoblast self-renewal and proliferation via regulating CDX2

Abnormal trophoblast self-renewal and differentiation during early gestation is the major cause of miscarriage, yet the underlying regulatory mechanisms remain elusive. Here, we show that trophoblast specific deletion of Kat8, a MYST family histone acetyltransferase, leads to extraembryonic ectoderm abnormalities and embryonic lethality. Employing RNA-seq and CUT&Tag analyses on trophoblast stem cells (TSCs), we further discover that KAT8 regulates the transcriptional activation of the trophoblast stemness marker, CDX2, via acetylating H4K16. Remarkably, CDX2 overexpression partially rescues the defects arising from Kat8 knockout. Moreover, increasing H4K16ac via using deacetylase SIRT1 inhibitor, EX527, restores CDX2 levels and promoted placental development. Clinical analysis shows reduced KAT8, CDX2 and H4K16ac expression are associated with recurrent pregnancy loss (RPL). Trophoblast organoids derived from these patients exhibit impaired TSC self-renewal and growth, which are significantly ameliorated with EX527 treatment. These findings suggest the therapeutic potential of targeting the KAT8-H4K16ac-CDX2 axis for mitigating RPL, shedding light on early gestational abnormalities.

Chromodomain mutation in S. pombe Kat5/Mst1 affects centromere dynamics and DNA repair.

KAT5 (S. pombe Mst1, human TIP60) is a MYST family histone acetyltransferase conserved from yeast to humans that is involved in multiple cellular activities. This family is characterized in part by containing a chromodomain, a motif associated with binding methylated histones. We show that a chromodomain mutation in the S. pombe Kat5, mst1-W66R, has defects in pericentromere silencing. mst1-W66R is sensitive to camptothecin (CPT) but only at an increased temperature of 36°C, although it is proficient for growth at this temperature. We also describe a de-silencing effect at the pericentromere by CPT that is independent of RNAi and methylation machinery. We also show that mst1-W66R disrupts recruitment of proteins to repair foci in response to camptothecin-induced DNA damage. Our data suggest a function of Mst1 chromodomain in centromere heterochromatin formation and a separate role in genome-wide damage repair in CPT.

MYST family histone acetyltransferases regulate reproductive diapause initiation

Histone acetylation, a crucial epigenetic mechanism, has been suggested to play a role in diapause regulation, but this has not been confirmed through gene loss-of-function studies. In this work, we investigated the involvement of MYST family genes, which are key writers of histone acetylation, in initiating reproductive diapause using the cabbage beetle Colaphellus bowringi as a model. We identified C. bowringi orthologs of MYST, including Tip60, KAT6A, KAT7, and KAT8, from previous transcriptomes. Analyses of phylogenetic trees and protein domains indicated that these MYST proteins are structurally conserved across animal species. Expression of these MYST genes was found to be enriched in heads and ovaries of C. bowringi. Under reproductive photoperiod conditions, RNAi targeting MYST genes, especially KAT8, suppressed ovarian growth and yolk deposition, resembling the characteristics of diapausing ovaries. Additionally, KAT8 knockdown led to the upregulation of diapause-related genes, such as heat shock proteins and diapause protein 1, and the emergence of diapause-like guts. Moreover, KAT8 knockdown reduced the expression of a crucial enzyme involved in juvenile hormone (JH) biosynthesis, likely due to decreased H4K16ac levels. Consequently, our findings suggest that MYST family genes, specifically KAT8, influence the JH signal, thereby regulating the initiation of reproductive diapause.

The MYST Family Histone Acetyltransferase SasC Governs Diverse Biological Processes in Aspergillus fumigatus.

The conserved MYST proteins form the largest family of histone acetyltransferases (HATs) that acetylate lysines within the N-terminal tails of histone, enabling active gene transcription. Here, we have investigated the biological and regulatory functions of the MYST family HAT SasC in the opportunistic human pathogenic fungus Aspergillus fumigatus using a series of genetic, biochemical, pathogenic, and transcriptomic analyses. The deletion (Δ) of sasC results in a drastically reduced colony growth, asexual development, spore germination, response to stresses, and the fungal virulence. Genome-wide expression analyses have revealed that the ΔsasC mutant showed 2402 significant differentially expressed genes: 1147 upregulated and 1255 downregulated. The representative upregulated gene resulting from ΔsasC is hacA, predicted to encode a bZIP transcription factor, whereas the UV-endonuclease UVE-1 was significantly downregulated by ΔsasC. Furthermore, our Western blot analyses suggest that SasC likely catalyzes the acetylation of H3K9, K3K14, and H3K29 in A. fumigatus. In conclusion, SasC is associated with diverse biological processes and can be a potential target for controlling pathogenic fungi.

HATS off to KAT6A/B inhibitors: A new way to target estrogen-receptor-positive breast cancer

Inhibitors for the KAT6 family of histone acetyltransferases (HATs) have been actively pursued due to the oncogenic roles of KAT6A in human cancer. CTx-648 is a novel KAT6A/B inhibitor with excellent pharmacokinetic properties and invivo efficacy that represents a promising new treatment strategy for estrogen-receptor-positive breast cancer.

Functional Characterization of the GNAT Family Histone Acetyltransferase Elp3 and GcnE in Aspergillus fumigatus.

Post-translational modifications of chromatin structure by histone acetyltransferase (HATs) play a pivotal role in the regulation of gene expression and diverse biological processes. However, the function of GNAT family HATs, especially Elp3, in the opportunistic human pathogenic fungus Aspergillus fumigatus is largely unknown. To investigate the roles of the GNAT family HATs Elp3 and GcnE in the A. fumigatus, we have generated and characterized individual null Δelp3 and ΔgcnE mutants. The radial growth of fungal colonies was significantly decreased by the loss of elp3 or gcnE, and the number of asexual spores (conidia) in the ΔgcnE mutant was significantly reduced. Moreover, the mRNA levels of the key asexual development regulators were also significantly low in the ΔgcnE mutant compared to wild type (WT). Whereas both the Δelp3 and ΔgcnE mutants were markedly impaired in the formation of adherent biofilms, the ΔgcnE mutant showed a complete loss of surface structure and of intercellular matrix. The ΔgcnE mutant responded differently to oxidative stressors and showed significant susceptibility to triazole antifungal agents. Furthermore, Elp3 and GcnE function oppositely in the production of secondary metabolites, and the ΔgcnE mutant showed attenuated virulence. In conclusion, Elp3 and GcnE are associated with diverse biological processes and can be potential targets for controlling the pathogenic fungus.

Molecular Insights into the Recognition of Acetylated Histone Modifications by the BRPF2 Bromodomain.

HBO1 [HAT bound to the origin recognition complex (ORC)], a member of the MYST family of histone acetyltransferases (HATs), was initially identified as a binding partner of ORC that acetylates free histone H3, H4, and nucleosomal H3. It functions as a quaternary complex with the BRPF (BRPF1/2/3) scaffolding protein and two accessory proteins, ING4/5 and Eaf6. Interaction of BRPF2 with HBO1 has been shown to be important for regulating H3K14 acetylation during embryonic development. However, how BRPF2 directs the HBO1 HAT complex to chromatin to regulate its HAT activity toward nucleosomal substrates remains unclear. Our findings reveal novel interacting partners of the BRPF2 bromodomain that recognizes different acetyllysine residues on the N-terminus of histone H4, H3, and H2A and preferentially binds to H4K5ac, H4K8ac, and H4K5acK12ac modifications. In addition, mutational analysis of the BRPF2 bromodomain coupled with isothermal titration calorimetry binding and pull-down assays on the histone substrates identified critical residues responsible for acetyllysine binding. Moreover, the BRPF2 bromodomain could enrich H4K5ac mark-bearing mononucleosomes compared to other acetylated H4 marks. Consistent with this, ChIP-seq analysis revealed that BRPF2 strongly co-localizes with HBO1 at histone H4K5ac and H4K8ac marks near the transcription start sites in the genome. Our study provides novel insights into how the histone binding function of the BRPF2 bromodomain directs the recruitment of the HBO1 HAT complex to chromatin to regulate gene expression.

Cyclic Mechanical Strain Regulates Osteoblastic Differentiation of Mesenchymal Stem Cells on TiO2 Nanotubes Through GCN5 and Wnt/β-Catenin.

Bone marrow mesenchymal stem cells (BMSCs) play a critical role in bone formation and are extremely sensitive to external mechanical stimuli. Mechanical signals can regulate the biological behavior of cells on the surface of titanium-related prostheses and inducing osteogenic differentiation of BMSCs, which provides the integration of host bone and prosthesis benefits. But the mechanism is still unclear. In this study, BMSCs planted on the surface of TiO2 nanotubes were subjected to cyclic mechanical stress, and the related mechanisms were explored. The results of alkaline phosphatase staining, real-time PCR, and Western blot showed that cyclic mechanical stress can regulate the expression level of osteogenic differentiation markers in BMSCs on the surface of TiO2 nanotubes through Wnt/β-catenin. As an important member of the histone acetyltransferase family, GCN5 exerted regulatory effects on receiving mechanical signals. The results of the ChIP assay indicated that GCN5 could activate the Wnt promoter region. Hence, we concluded that the osteogenic differentiation ability of BMSCs on the surface of TiO2 nanotubes was enhanced under the stimulation of cyclic mechanical stress, and GCN5 mediated this process through Wnt/β-catenin.

O-GlcNAc modified-TIP60/KAT5 is required for PCK1 deficiency-induced HCC metastasis

Aberrant glucose metabolism and elevated O-linked β-N-acetylglucosamine modification (O-GlcNAcylation) are hallmarks of hepatocellular carcinoma (HCC). Loss of phosphoenolpyruvate carboxykinase 1 (PCK1), the major rate-limiting enzyme of hepatic gluconeogenesis, increases hexosamine biosynthetic pathway (HBP)-mediated protein O-GlcNAcylation in hepatoma cell and promotes cell growth and proliferation. However, whether PCK1 deficiency and hyper O-GlcNAcylation can induce HCC metastasis is largely unknown. Here, gain- and loss-of-function studies demonstrate that PCK1 suppresses HCC metastasis in vitro and in vivo. Specifically, lysine acetyltransferase 5 (KAT5), belonging to the MYST family of histone acetyltransferases (HAT), is highly modified by O-GlcNAcylation in PCK1 knockout hepatoma cells. Mechanistically, PCK1 depletion suppressed KAT5 ubiquitination by increasing its O-GlcNAcylation, thereby stabilizing KAT5. KAT5 O-GlcNAcylation epigenetically activates TWIST1 expression via histone H4 acetylation, and enhances MMP9 and MMP14 expression via c-Myc acetylation, thus promoting epithelial-mesenchymal transition (EMT) in HCC. In addition, targeting HBP-mediated O-GlcNAcylation of KAT5 inhibits lung metastasis of HCC in hepatospecific Pck1-deletion mice. Collectively, our findings demonstrate that PCK1 depletion increases O-GlcNAcylation of KAT5, epigenetically induces TWIST1 expression and promotes HCC metastasis, and link metabolic enzyme, post-translational modification (PTM) with epigenetic regulation.

HBO1 overexpression is important for hepatocellular carcinoma cell growth

Hepatocellular carcinoma (HCC) is a common primary liver malignancy lacking effective molecularly-targeted therapies. HBO1 (lysine acetyltransferase 7/KAT7) is a member of MYST histone acetyltransferase family. Its expression and potential function in HCC are studied. We show that HBO1 mRNA and protein expression is elevated in human HCC tissues and HCC cells. HBO1 expression is however low in cancer-surrounding normal liver tissues and hepatocytes. In HepG2 and primary human HCC cells, shRNA-induced HBO1 silencing or CRISPR/Cas9-induced HBO1 knockout potently inhibited cell viability, proliferation, migration, and invasion, while provoking mitochondrial depolarization and apoptosis induction. Conversely, ectopic overexpression of HBO1 by a lentiviral construct augmented HCC cell proliferation, migration and invasion. In vivo, xenografts-bearing HBO1-KO HCC cells grew significantly slower than xenografts with control HCC cells in severe combined immunodeficient mice. These results suggest HBO1 overexpression is important for HCC cell progression.

Schizosaccharomyces pombe KAT5 contributes to resection and repair of a DNA double-strand break.

Chromatin remodeling is essential for effective repair of a DNA double-strand break (DSB). KAT5 (Schizosaccharomyces pombe Mst1, human TIP60) is a MYST family histone acetyltransferase conserved from yeast to humans that coordinates various DNA damage response activities at a DNA DSB, including histone remodeling and activation of the DNA damage checkpoint. In S. pombe, mutations in mst1+ causes sensitivity to DNA damaging drugs. Here we show that Mst1 is recruited to DSBs. Mutation of mst1+ disrupts recruitment of repair proteins and delays resection. These defects are partially rescued by deletion of pku70, which has been previously shown to antagonize repair by homologous recombination (HR). These phenotypes of mst1 are similar to pht1-4KR, a nonacetylatable form of histone variant H2A.Z, which has been proposed to affect resection. Our data suggest that Mst1 functions to direct repair of DSBs toward HR pathways by modulating resection at the DSB.

Dynamic histone acetylation in floral volatile synthesis and emission in petunia flowers.

Biosynthesis of secondary metabolites relies on primary metabolic pathways to provide precursors, energy, and cofactors, thus requiring coordinated regulation of primary and secondary metabolic networks. However, to date, it remains largely unknown how this coordination is achieved. Using Petunia hybrida flowers, which emit high levels of phenylpropanoid/benzenoid volatile organic compounds (VOCs), we uncovered genome-wide dynamic deposition of histone H3 lysine 9 acetylation (H3K9ac) during anthesis as an underlying mechanism to coordinate primary and secondary metabolic networks. The observed epigenome reprogramming is accompanied by transcriptional activation at gene loci involved in primary metabolic pathways that provide precursor phenylalanine, as well as secondary metabolic pathways to produce volatile compounds. We also observed transcriptional repression among genes involved in alternative phenylpropanoid branches that compete for metabolic precursors. We show that GNAT family histone acetyltransferase(s) (HATs) are required for the expression of genes involved in VOC biosynthesis and emission, by using chemical inhibitors of HATs, and by knocking down a specific HAT gene, ELP3, through transient RNAi. Together, our study supports that regulatory mechanisms at chromatin level may play an essential role in activating primary and secondary metabolic pathways to regulate VOC synthesis in petunia flowers.

CBP/p300 homologs CBP2 and CBP3 play distinct roles in planarian stem cell function

Chromatin modifications function as critical regulators of gene expression and cellular identity, especially in the regulation and maintenance of the pluripotent state. However, many studies of chromatin modification in stem cells-and pluripotent stem cells in particular-are performed in mammalian stem cell culture, an in vitro condition mimicking a very transient state during mammalian development. Thus, new models for studying pluripotent stem cells in vivo could be helpful for understanding the roles of chromatin modification, for confirming prior in vitro studies, and for exploring evolution of the pluripotent state. The freshwater flatworm, Schmidtea mediterranea, is an excellent model for studying adult pluripotent stem cells, particularly in the context of robust, whole-body regeneration. To identify chromatin modifying and remodeling enzymes critical for planarian regeneration and stem cell maintenance, we took a candidate approach and screened planarian homologs of 25 genes known to regulate chromatin biology in other organisms. Through our study, we identified six genes with novel functions in planarian homeostasis, regeneration, and behavior. Of the list of genes characterized, we identified five planarian homologs of the mammalian CREB-Binding Protein (CBP) and p300 family of histone acetyltransferases, representing an expansion of this family in planarians. We find that two planarian CBP family members are required for planarian survival, with knockdown of Smed-CBP2 and Smed-CBP3 causing distinct defects in stem cell maintenance or function. Loss of CBP2 causes a quick, dramatic loss of stem cells, while knockdown of CBP3 affects stem cells more narrowly, influencing differentiation of several cell types that include neuronal subtypes and cells of the eye. Further, we find that Smed-CBP1 is required for planarian fissioning behavior. We propose that the division of labor among a diversified CBP family in planarians presents an opportunity to dissect specific functions of a broadly important histone acetyltransferase family.

Genome-wide identification of citrus histone acetyltransferase and deacetylase families and their expression in response to arbuscular mycorrhizal fungi and drought

ABSTRACT Histone acetyltransferase (HAT) and histone deacetylase (HDAC) family members control histone acetylation levels to regulate gene expression and play significant roles in microbe interactions and abioticstress responses. However, the HAT and HDAC family members in citrus have not yet been identified, and their responses to drought (abiotic stress) and arbuscular mycorrhizal fungi (AMF) colonization, remain unknown. In the present study, HAT and HDAC family genes were characterized in the citrus genome. A total of 14 CsHATs and 11 CsHDACs were identified in the citrus genome, of which 8 CsHATs and 7 CsHDACs showed syntenic relationships with Arabidopsis HATs and HDACs, respectively. Furthermore, the 14 CsHATs were classified into GNAT-MYST, TAFII250, MYST, CBP, and one citrus-unique family. The 11 CsHDACs were classified into SIR2, HD2, and RPD3/HDA1. Gene expression analysis showed AMF colonizationor drought induced the expression of CsHAT1, 2, 6, 7, 9, 11, 12, and 14, and repressed CsHDAC1, 2, 5, 6, 8, and 10. The regulation of HAT and HDAC expression was not always consistent with the cis-elements identified in their promoters. The present study offers useful insights into the citrus HAT and HDAC gene families and their expression in response to AMF colonization and drought stress.

KAT8/MOF-Mediated Anti-Cancer Mechanism of Gemcitabine in Human Bladder Cancer Cells.

Histone acetylation is a well-characterized epigenetic modification controlled by histone acetyltransferases (HATs) and histone deacetylases (HDACs). Imbalanced histone acetylation has been observed in many primary cancers. Therefore, efforts have been made to find drugs or small molecules such as HDAC inhibitors that can revert acetylation levels to normal in cancer cells. We observed dose-dependent reduction in the endogenous and exogenous protein expression levels of KAT8 (also known as human MOF), a member of the MYST family of HATs, and its corresponding histone acetylation at H4K5, H4K8, and H4K16 in chemotherapy drug gemcitabine (GEM)-exposed T24 bladder cancer (BLCA) cells. Interestingly, the reduction in MOF and histone H4 acetylation was inversely proportional to GEM-induced γH2AX, an indicator of chemotherapy drug effectiveness. Furthermore, pGL4-MOF-Luc reporter activities were significantly inhibited by GEM, thereby suggesting that GEM utilizes an MOF-mediated anti-BLCA mechanism of action. In the CCK-8, wound healing assays and Transwell® experiments, the additive effects on cell proliferation and migration were observed in the presence of exogenous MOF and GEM. In addition, the promoted cell sensitivity to GEM by exogenous MOF in BLCA cells was confirmed using an Annexin V-FITC/PI assay. Taken together, our results provide the theoretical basis for elucidating the anti-BLCA mechanism of GEM.

SWI/SNF complexes act through CBP-1 histone acetyltransferase to regulate acute functional tolerance to alcohol

BackgroundSWI/SNF chromatin remodeling genes are required for normal acute responses to alcohol in C. elegans and are associated with alcohol use disorder in two human populations. In an effort to discover the downstream genes that are mediating this effect, we identified SWI/SNF-regulated genes in C. elegans.ResultsTo identify SWI/SNF-regulated genes in adults, we compared mRNA expression in wild type and swsn-1(os22ts) worms under conditions that produce inactive swsn-1 in mature cells. To identify SWI/SNF-regulated genes in neurons, we compared gene expression in swsn-9(ok1354) null mutant worms that harbor a neuronal rescue or a control construct. RNA sequencing was performed to an average depth of 25 million reads per sample using 50-base, paired-end reads. We found that 6813 transcripts were significantly differentially expressed between swsn-1(os22ts) mutants and wild-type worms and 2412 transcripts were significantly differentially expressed between swsn-9(ok1354) mutants and swsn-9(ok1354) mutants with neuronal rescue. We examined the intersection between these two datasets and identified 603 genes that were differentially expressed in the same direction in both comparisons; we defined these as SWI/SNF-regulated genes in neurons and in adults. Among the differentially expressed genes was cbp-1, a C. elegans homolog of the mammalian CBP/p300 family of histone acetyltransferases. CBP has been implicated in the epigenetic regulation in response to alcohol in animal models and a polymorphism in the human CBP gene, CREBBP, has been associated with alcohol-related phenotypes. We found that cbp-1 is required for the development of acute functional tolerance to alcohol in C. elegans.ConclusionsWe identified 603 transcripts that were regulated by two different SWI/SNF complex subunits in adults and in neurons. The SWI/SNF-regulated genes were highly enriched for genes involved in membrane rafts, suggesting an important role for this membrane microdomain in the acute alcohol response. Among the differentially expressed genes was cbp-1; CBP-1 homologs have been implicated in alcohol responses across phyla and we found that C. elegans cbp-1 was required for the acute alcohol response in worms.

Insights Into the Function of the NuA4 Complex in Plants.

Chromatin remodeling plays a key role in the establishment and maintenance of gene expression patterns essential for plant development and responses to environmental factors. Post-translational modification of histones, including acetylation, is one of the most relevant chromatin remodeling mechanisms that operate in eukaryotic cells. Histone acetylation is an evolutionarily conserved chromatin signature commonly associated with transcriptional activation. Histone acetylation levels are tightly regulated through the antagonistic activity of histone acetyltransferases (HATs) and histone deacetylases (HDACs). In plants, different families of HATs are present, including the MYST family, which comprises homologs of the catalytic subunit of the Nucleosome Acetyltransferase of H4 (NuA4) complex in yeast. This complex mediates acetylation of histones H4, H2A, and H2A.Z, and is involved in transcriptional regulation, heterochromatin silencing, cell cycle progression, and DNA repair in yeast. In Arabidopsis and, other plant species, homologs for most of the yeast NuA4 subunits are present and although the existence of this complex has not been demonstrated yet, compelling evidence supports the notion that this type of HAT complex functions from mosses to angiosperms. Recent proteomic studies show that several Arabidopsis homologs of NuA4 components, including the assembly platform proteins and the catalytic subunit, are associated in vivo with additional members of this complex suggesting that a NuA4-like HAT complex is present in plants. Furthermore, the functional characterization of some Arabidopsis NuA4 subunits has uncovered the involvement of these proteins in the regulation of different plant biological processes. Interestingly, for most of the mutant plants deficient in subunits of this complex characterized so far, conspicuous defects in flowering time are observed, suggesting a role for NuA4 in the control of this plant developmental program. Moreover, the participation of Arabidopsis NuA4 homologs in other developmental processes, such as gametophyte development, as well as in cell proliferation and stress and hormone responses, has also been reported. In this review, we summarize the current state of knowledge on plant putative NuA4 subunits and discuss the latest progress concerning the function of this chromatin modifying complex.

Prognostic role of upregulated P300 expression in human cancers: A clinical study of synovial sarcoma and a meta-analysis.

E1A binding protein p300 (P300) is a member of the histone acetyltransferase family of transcriptional co-activators, which are associated with various types of cancer. Numerous studies have evaluated the diagnostic value of P300, but their results are not consistent. Therefore, a clinical study and a meta-analysis were performed in the present study to investigate the prognostic value of P300 expression in human malignant neoplasms. Immunohistochemical (IHC) analysis was used to assess P300 expression in 43 paraffin-embedded primary synovial sarcoma (SS) samples. For the meta-analysis, eligible studies published until January 21, 2018 were identified by searching the PubMed, EMBASE and Web of Science databases. The IHC analysis indicated a high P300 expression rate in 33.3% (10/30) of biphasic SS (BSSs) and in 60% (6/10) of monophasic fibrous SS tissues. In BSS, the expression rate was significantly higher in the epithelial component (80.0%, 24/30) than that in the spindle-cell component (30.0%, 9/30; P<0.05). The meta-analysis indicated that high expression of P300 was associated with poor overall survival (OS) in digestive system malignant neoplasms (HR=1.54, 95% CI: 1.20-2.23), as well as with poor progression-free survival, recurrence-free survival and disease-free survival combined (HR=1.84, 95% CI: 1.36-2.47). Analysis of subgroups by ethnicity demonstrated that high expression of P300 was associated with poor OS in Asians (HR=1.72, 95% CI: 1.20-2.47) but favourable OS in Caucasians (HR=0.59, 95% CI: 0.47-0.73). Furthermore, high expression of P300 was associated with clinical stage [Relative Risk (RR)=1.30, 95% CI: 1.07-1.58], lymph node metastasis (RR=1.30, 95% CI: 1.03-1.64) and depth of invasion (RR=1.31, 95% CI: 1.07-1.60). P300 expression may therefore be a useful biomarker for predicting patient prognosis in various types of human cancer.

A MYST family histone acetyltransferase, MoSAS3, is required for development and pathogenicity in the rice blast fungus.

SummaryHistone acetylation has been established as a principal epigenetic regulatory mechanism in eukaryotes. Sas3, a histone acetyltransferase belonging to the largest family of acetyltransferase, MYST, is the catalytic subunit of a conserved histone acetyltransferase complex. To date, the functions of Sas3 and its orthologues have been extensively studied in yeast, humans and flies in relation to global acetylation and transcriptional regulation. However, its precise impact on development and pathogenicity in fungal plant pathogens has yet to be elucidated. Considering the importance of Sas3 in H3K14 acetylation, here we investigate the roles of its orthologue in the rice blast fungus, Magnaporthe oryzae (Pyricularia oryzae). Unlike a previously reported Sas3 deletion in yeast, which led to no remarkable phenotypic changes, we found that MoSAS3 deletion alone had a profound effect on fungal growth and development, including asexual reproduction, germination and appressorium formation in M. oryzae. Such defects in pre‐penetration development resulted in complete loss of pathogenicity in the deletion mutant. Furthermore, genetic analysis of MoSAS3 and MoGCN5 encoding a Gcn5‐related N‐acetyltransferase family histone acetyltransferase suggested that two conserved components of histone acetylation are integrated differently into epigenetic regulatory mechanisms in the yeast and a filamentous fungus. RNA‐seq analysis of ΔMosas3 showed two general trends: many DNA repair and DNA damage response genes are up‐regulated, while carbon and nitrogen metabolism genes are down‐regulated in ΔMosas3. Our work demonstrates the importance of MYST family histone acetyltransferase as a developmental regulator and illuminates a degree of functional variation in conserved catalytic subunits among different fungal species.

Crosstalk between cellular metabolism and histone acetylation.

Dynamic interplay between cellular metabolism and histone acetylation is a key mechanism underlying metabolic control of epigenetics. In particular, the central metabolite acetyl-coenzyme A (acetyl-CoA) acts as the acetyl-donor for histone acetylation in both an enzymatic and non-enzymatic manner. Since members of the family of histone acetyl transferases (HATs) that catalyze the acetylation of histone tails possess a Michaelis constant (Km) within the range of physiological cellular acetyl-CoA concentrations, changing concentrations of acetyl-CoA can restrict or promote enzymatic histone acetylation. Likewise, non-enzymatic histone acetylation occurs at physiological concentrations. These concepts implicate acetyl-CoA as a rheostat for nutrient availability acting, in part, by controlling histone acetylation. Histone acetylation is an important epigenetic modification that controls gene expression and acetyl-CoA dependent changes in both histone acetylation and gene expression have been shown in yeast and mammalian systems. However, quantifying the metabolic conditions required to achieve specific changes in histone acetylation is a major challenge. The relationship between acetyl-CoA and histone acetylation may be influenced by a variety of factors including sub-cellular location of metabolites and enzymes, relative quantities of metabolites, and substrate availability/preference. A diversity of substrates can contribute the two-carbon acyl-chain to acetyl-CoA, a number of pathways can create or degrade acetyl-CoA, and only a handful of potential mechanisms for the crosstalk between metabolism and histone acetylation have been explored. The centrality of acetyl-CoA in intermediary metabolism means that acetyl-CoA levels may change, or be resistant to change, in unexpected ways. Thus, quantification of relevant metabolites is critical evidence in understanding how the nutrient rheostat is set in normal and pathological contexts. Coupling metabolite quantitation with isotope tracing to examine fate of specific metabolites is critical to the crosstalk between metabolism and histone acetylation, including but not limited to acetyl-CoA provides necessary context. This chapter provides guidance on experimental design of quantification with isotope dilution and/or tracing of acetyl-CoA within a targeted or highly multiplexed multi-analyte workflow.