DNA Modification Research Articles

Phage SEP1 hijacks Staphylococcus epidermidis stationary cells' metabolism to replicate.

In nature, bacteria often survive in a stationary state with low metabolic activity. Phages use the metabolic machinery of the host cell to replicate, and, therefore, their efficacy against non-dividing cells is usually limited. Nevertheless, it was previously shown that the Staphylococcus epidermidis phage SEP1 has the remarkable capacity to actively replicate in stationary-phase cells, reducing their numbers. Here, we studied for the first time the transcriptomic profiles of both exponential and stationary cells infected with SEP1 phage using RNA-seq to gain a better understanding of this rare phenomenon. We showed that SEP1 successfully takes over the transcriptional apparatus of both exponential and stationary cells. Infection was, however, delayed in stationary cells, with genes within the gp142-gp154 module putatively implicated in host takeover. S. epidermidis responded to SEP1 infection by upregulating three genes involved in a DNA modification system, with this being observed already 5 min after infection in exponential cells and later in stationary cells. In stationary cells, a significant number of genes involved in translation and RNA metabolic and biosynthetic processes were upregulated after 15 and 30 min of SEP1 infection in comparison with the uninfected control, showing that SEP1 activates metabolic and biosynthetic pathways necessary to its successful replication.IMPORTANCEMost phage-host interaction studies are performed with exponentially growing cells. However, this cell state is not representative of what happens in natural environments. Additionally, most phages fail to replicate in stationary cells. The Staphylococcus epidermidis phage SEP1 is one of the few phages reported to date to be able to infect stationary cells. Here, we unveiled the interaction of SEP1 with its host in both exponential and stationary states of growth at the transcriptomic level. The findings of this study provide valuable insights for a better implementation of phage therapy since phages able to infect stationary cells could be more efficient in the treatment of recalcitrant infections.

Natural Epigenetic DNA Modifications Cause Remote DNA Photodamage.

5-Formyl-2'-deoxycytidine, an intermediate during the erasure of epigenetic marker 5-methyl-2'-deoxycytidine, and 5-formyl-2'-deoxyuridine, an oxidative lesion of thymidine, are naturally occurring DNA modifications. The carbonyl groups of these DNA modifications are the smallest possible photosensitizers and have the potential to generate cyclobutane pyrimidine dimers upon irradiation with UV light. To evidence this damaging potential, ternary DNA architectures were used, in which the photosensitizer and the damage site were located at well-defined positions in the sequences. The quantitative and time-dependent analysis revealed not only the high photodamaging potential of both natural DNA modifications but also the mechanisms for this new pathway to photodamage. 5-Formyl-2'-deoxycytidine is more efficiently generating cyclobutane pyrimidine dimers than 5-formyl-2'-deoxyuridine because the latter is also photochemically converted to 5-carboxy-2'-deoxyuridine. This demonstrates for the first time that epigenetic DNA modifications regulating gene expression interact with sunlight and can induce DNA photodamages.

Associations between placental hydroxymethylation and birthweight.

5-hydroxymethylcytosine (5hmC), formed through the ten-eleven translocation (TET) methylcytosine dioxygenase mediated oxidation of 5-methylcytosine (5mC) at cytosine-phosphate-guanine (CpG) dinucleotides, is believed to mainly serve as an intermediate in the DNA demethylation pathway, though recent evidence suggests that 5hmC may also play a functionally relevant role. We have conducted an epigenome-wide association study (EWAS) to assess the association between placenta 5hmC, obtained through parallel bisulfite and oxidative bisulfite modification of DNA and array-based assessment, and newborn birthweight in the Rhode Island Child Health Study (RICHS). We also assessed whether the removal of 5hmC signal impacts the observed results from traditional epigenome-wide studies that rely on BS modification-based (combined 5mC and 5hmC) assessment alone. We identified 5hmC at one CpG in the CUBN gene to be significantly associated with birthweight (FDR < 0.05) and demonstrate that expression of that gene was also associated with birthweight. Comparison of 5hmC+5mC and 5mC EWAS effect estimates reveal a strong correlation (r = 0.77, p < 0.0001). Our study suggests that traditional assessment of 5mC through bisulfite modification alone provides an accurate assessment of CpG-specific DNA methylation for EWAS studies but was unable to provide evidence of widespread associations between placental 5hmC and birthweight.

Parkinson's disease-associated shifts between DNA methylation and DNA hydroxymethylation in human brain in PD-related genes, including PARK19 (DNAJC6) and PTPRN2 (IA-2β).

The majority of Parkinson's disease (PD) cases are due to a complex interaction between aging, genetics, and environmental factors; epigenetic mechanisms are thought to act as important mediators of these risk factors. While multiple studies to date have explored the role of DNA modifications in PD, few focus on 5-hydroxymethylcytosine (5hmC). Because 5hmC occurs at its highest levels in the brain and is thought to be particularly important in the central nervous system, particularly in the response to neurotoxicants, it is important to explore the potential role of 5hmC in PD. This study expands on our previously published epigenome-wide association study (EWAS) performed on DNA isolated from neuron-enriched nuclei from human postmortem parietal cortex from the Banner Sun Health Research Institute Brain Bank. The study aimed to identify paired changes in 5hmC and 5mC in PD in enriched neuronal nuclei isolated from PD post-mortem parietal cortex and age- and sex-matched controls. We performed oxidative bisulfite (oxBS) conversion and paired it with our previously published bisulfite (BS)-based EWAS on the same samples to identify cytosines with significant shifts between these two related epigenetic marks. Interaction differentially modified cytosines (iDMCs) were identified using our recently published mixed-effects model for co-analyzing βmC and βhmC data. We identified 1,030 iDMCs with paired changes in 5mC and 5hmC (FDR < 0.05) that map to 695 genes, including PARK19 (DNAJC6), a familial PD gene, and PTPRN2 (IA-2), which has been previously implicated in PD in both epigenetic and mechanistic studies. The majority of iDMC-containing genes have not previously been implicated in PD and were not identified in our previous BS-based EWAS. These data potentially link epigenetic regulation of the PARK19 and PTPRN2 loci in the pathogenesis of idiopathic PD. In addition, iDMC-containing genes have known functions in synaptic formation and function, cell cycle and senescence, neuroinflammation, and epigenetic regulation. These data suggest that there are significant shifts between 5mC and 5hmC associated with PD in genes relevant to PD pathogenesis that are not captured by analyzing BS-based data alone or by analyzing each mark as a distinct dataset.

Developmental origins of Parkinson's disease risk: perinatal exposure to the organochlorine pesticide dieldrin leads to sex-specific DNA modifications in critical neurodevelopmental pathways in the mouse midbrain.

Epidemiological studies show that exposure to the organochlorine pesticide dieldrin is associated with an increased risk of Parkinson's disease (PD). Animal studies support a link between developmental dieldrin exposure and increased neuronal susceptibility in the α-synuclein preformed fibril and MPTP models in adult male C57BL/6 mice. In a previous study, we showed that developmental dieldrin exposure was associated with sex-specific changes in DNA modifications within genes related to dopaminergic neuron development and maintenance at 12 wk of age. Here, we used capture hybridization-sequencing with custom baits to interrogate DNA modifications across the entire genetic loci of the previously identified genes at multiple time points-birth, 6, 12, and 36 wk old. We identified largely sex-specific dieldrin-induced changes in DNA modifications at each time point that annotated to pathways important for neurodevelopment, potentially related to critical steps in early neurodevelopment, dopaminergic neuron differentiation, synaptogenesis, synaptic plasticity, and glial-neuron interactions. Despite large numbers of age-specific DNA modifications, longitudinal analysis identified a small number of differential modification of cytosines with dieldrin-induced deflection of epigenetic aging. The sex-specificity of these results adds to evidence that sex-specific responses to PD-related exposures may underly sex-specific differences in disease. Overall, these data support the idea that developmental dieldrin exposure leads to changes in epigenetic patterns that persist after the exposure period and disrupt critical neurodevelopmental pathways, thereby impacting risk of late-life diseases, including PD.

Decoding the Epigenetic Landscape: Insights into 5mC and 5hmC Patterns in Mouse Cortical Cell Types.

The DNA modifications, 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC), represent powerful epigenetic regulators of temporal and spatial gene expression. Yet, how the cooperation of these genome-wide, epigenetic marks determine unique transcriptional signatures across different brain cell populations is unclear. Here we applied Nanopore sequencing of native DNA to obtain a complete, genome-wide, single-base resolution atlas of 5mC and 5hmC modifications in neurons, astrocytes and microglia in the mouse cortex (99% genome coverage, 40 million CpG sites). In tandem with RNA sequencing, analysis of 5mC and 5hmC patterns across cell types reveals astrocytes drive uniquely high brain 5hmC levels and support two decades of research regarding methylation patterns, gene expression and alternative splicing, benchmarking this resource. As such, we provide the most comprehensive DNA methylation data in mouse brain as an interactive, online tool (NAM-Me, https://olsenlab.shinyapps.io/NAMME/) to serve as a resource dataset for those interested in the methylome landscape.

A Preliminary Study of the Occurrence of Genetic Changes in mtDNA in the Muscles in Children Treated for Strabismus.

Background: The dysregulation of extraocular muscles (EOMs) in the strabismus may be partly due to modification in the mitochondrial DNA (mtDNA). Currently, little is known about changes occurring in mtDNA of EOMs in patients with strabismus, therefore the aim of our study was to analyze if there are any changes occurring in the mitochondrial DNA of extraocular muscles in children that underwent strabismus surgery in our clinic. Methods: MtDNA was isolated from the tissue material using the Qiagen kit. Assessment of mtDNA mutations was performed by next-generation sequencing (NGS) using the Illumina MiSeq protocol. Results: The examination revealed the presence of atrophic changes in muscle fibers. NGS evaluation revealed a dominant genetic mutation in the ANT1 gene in 12 of the 15 patients examined. Conclusions: The presented results constitute the beginning of research on changes in mtDNA occurring in the muscles of children with strabismus surgery. Further studies are necessary in the context of resolving the transcriptomic differences between strabismic and non-strabismic EOMs. Better understanding of the molecular genetics of strabismus will lead to improved knowledge of the disease mechanisms and ultimately to a more effective treatment.

Overview of chromatin regulatory processes during surface ectodermal development and homeostasis

The ectoderm is the outermost of the three germ layers of the early embryo that arise during gastrulation. Once the germ layers are established, the complex interplay of cellular proliferation, differentiation, and migration results in organogenesis. The ectoderm is the progenitor of both the surface ectoderm and the neural ectoderm. Notably, the surface ectoderm develops into the epidermis and its associated appendages, nails, external exocrine glands, olfactory epithelium, and the anterior pituitary. Specification, development, and homeostasis of these organs demand a tightly orchestrated gene expression program that is often dictated by epigenetic regulation. In this review, we discuss the recent discoveries that have highlighted the importance of chromatin regulatory mechanisms mediated by transcription factors, histone and DNA modifications that aid in the development of surface ectodermal organs and maintain their homeostasis post-development.

Rockfish: A transformer-based model for accurate 5-methylcytosine prediction from nanopore sequencing

DNA methylation plays an important role in various biological processes, including cell differentiation, ageing, and cancer development. The most important methylation in mammals is 5-methylcytosine mostly occurring in the context of CpG dinucleotides. Sequencing methods such as whole-genome bisulfite sequencing successfully detect 5-methylcytosine DNA modifications. However, they suffer from the serious drawbacks of short read lengths and might introduce an amplification bias. Here we present Rockfish, a deep learning algorithm that significantly improves read-level 5-methylcytosine detection by using Nanopore sequencing. Rockfish is compared with other methods based on Nanopore sequencing on R9.4.1 and R10.4.1 datasets. There is an increase in the single-base accuracy and the F1 measure of up to 5 percentage points on R.9.4.1 datasets, and up to 0.82 percentage points on R10.4.1 datasets. Moreover, Rockfish shows a high correlation with whole-genome bisulfite sequencing, requires lower read depth, and achieves higher confidence in biologically important regions such as CpG-rich promoters while being computationally efficient. Its superior performance in human and mouse samples highlights its versatility for studying 5-methylcytosine methylation across varied organisms and diseases. Finally, its adaptable architecture ensures compatibility with new versions of pores and chemistry as well as modification types.

A disposable electrochemical magnetic immunosensor for the rapid and sensitive detection of 5-formylcytosine and 5-carboxylcytosine in DNA

5-formylcytosine (5 fC) and 5-carboxylcytosine (5caC) serve as key intermediates in DNA demethylation process with significant implications for gene regulation and disease progression. In this study, we introduce a novel electrochemical sensing platform specifically designed for the sensitive and selective detection of 5 fC and 5caC in DNA. Protein A-modified magnetic beads (ProtA-MBs) coupled with specific antibodies facilitate the immunorecognition and enrichment of these modified bases. Signal amplification is achieved through several chemical reactions involving the interaction between N3-kethonaxl and guanine, copper-free click chemistry for the attachment of dibenzocyclooctyne (DBCO)-Biotin, and the subsequent recognition by streptavidin-conjugated horseradish peroxidase (SA-HRP). The assay's readout is performed on a disposable laser-induced graphene (LIG) electrode, modified with the bead-antibody-DNA complex in a magnetic field, and analyzed using differential pulse voltammetry in a system employing hydroquinone (HQ) as the redox mediator and H2O2 as the substrate. This immunosensor displayed excellent sensitivity, with detection limits of 14.8 fM for 5 fC across a 0.1–1000 pM linear range and 87.4 fM for 5caC across a 0.5–5000 pM linear range, and maintained high selectivity even in the presence of interferences from other DNA modifications. Successful application in quantifying 5 fC and 5caC in genomic DNA from cell extracts, with recovery rates between 97.7% to 102.9%, underscores its potential for clinical diagnostics. N3-kethoxal was used for the first time in an electrochemical sensor. This work not only broadens the toolkit for detecting DNA modifications but also provides a fresh impetus for the development of point-of-care testing (POCT) technologies.

Unveiling nuclear chromatin distribution using IsoConcentraChromJ: A flourescence imaging plugin for IsoRegional and IsoVolumetric based ratios analysis.

Chromatin exhibits non-random distribution within the nucleus being arranged into discrete domains that are spatially organized throughout the nuclear space. Both the spatial distribution and structural rearrangement of chromatin domains in the nucleus depend on epigenetic modifications of DNA and/or histones and structural elements such as the nuclear envelope. These components collectively contribute to the organization and rearrangement of chromatin domains, thereby influencing genome architecture and functional regulation. This study develops an innovative, user-friendly, ImageJ-based plugin, called IsoConcentraChromJ, aimed quantitatively delineating the spatial distribution of chromatin regions in concentric patterns. The IsoConcentraChromJ can be applied to quantitative chromatin analysis in both two- and three-dimensional spaces. After DNA and histone staining with fluorescent probes, high-resolution images of nuclei have been obtained using advanced fluorescence microscopy approaches, including confocal and stimulated emission depletion (STED) microscopy. IsoConcentraChromJ workflow comprises the following sequential steps: nucleus segmentation, thresholding, masking, normalization, and trisection with specified ratios for either 2D or 3D acquisitions. The effectiveness of the IsoConcentraChromJ has been validated and demonstrated using experimental datasets consisting in nuclei images of pre-adipocytes and mature adipocytes, encompassing both 2D and 3D imaging. The outcomes allow to characterize the nuclear architecture by calculating the ratios between specific concentric nuclear areas/volumes of acetylated chromatin with respect to total acetylated chromatin and/or total DNA. The novel IsoConcentrapChromJ plugin could represent a valuable resource for researchers investigating the rearrangement of chromatin architecture driven by epigenetic mechanisms using nuclear images obtained by different fluorescence microscopy methods.

Engineered Methionine Adenosyltransferase Cascades for Metabolic Labeling of Individual DNA Methylomes in Live Cells.

Methylation, a widely occurring natural modification serving diverse regulatory and structural functions, is carried out by a myriad of S-adenosyl-l-methionine (AdoMet)-dependent methyltransferases (MTases). The AdoMet cofactor is produced from l-methionine (Met) and ATP by a family of multimeric methionine adenosyltransferases (MAT). To advance mechanistic and functional studies, strategies for repurposing the MAT and MTase reactions to accept extended versions of the transferable group from the corresponding precursors have been exploited. Here, we used structure-guided engineering of mouse MAT2A to enable biocatalytic production of an extended AdoMet analogue, Ado-6-azide, from a synthetic methionine analogue, S-(6-azidohex-2-ynyl)-l-homocysteine (N3-Met). Three engineered MAT2A variants showed catalytic proficiency with the extended analogues and supported DNA derivatization in cascade reactions with M.TaqI and an engineered variant of mouse DNMT1 both in the absence and presence of competing Met. We then installed two of the engineered variants as MAT2A-DNMT1 cascades in mouse embryonic stem cells by using CRISPR-Cas genome editing. The resulting cell lines maintained normal viability and DNA methylation levels and showed Dnmt1-dependent DNA modification with extended azide tags upon exposure to N3-Met in the presence of physiological levels of Met. This for the first time demonstrates a genetically stable system for biosynthetic production of an extended AdoMet analogue, which enables mild metabolic labeling of a DNMT-specific methylome in live mammalian cells.

CCAAT Promoter element regulates transgenerational expression of the MHC class I gene

Transgenerational gene expression depends on both underlying DNA sequences and epigenetic modifications. The latter, which can result in transmission of variegated gene expression patterns across multiple generations without DNA alterations, has been termed epigenetic inheritance and has been documented in plants, worms, flies and mammals. Whereas transcription factors binding to cognate DNA sequence elements regulate gene expression, the molecular basis for epigenetic inheritance has been linked to histone and DNA modifications and non-coding RNA. Here we report that mutation of the CCAAT box promoter element abrogates NF-Y binding and disrupts the stable transgenerational expression of an MHC class I transgene. Transgenic mice with a mutated CCAAT box in the MHC class I transgene display variegated expression of the transgene among littermates and progeny in multiple independently derived transgenic lines. After 4 generations, CCAAT mutant transgenic lines derived from a single founder stably displayed distinct patterns of expression. Histone modifications and RNA polymerase II binding correlate with expression of CCAAT mutant transgenic lines, whereas DNA methylation and nucleosome occupancy do not. Mutation of the CCAAT box also results in changes to CTCF binding and DNA looping patterns across the transgene that correlate with expression status. These studies identify the CCAAT promoter element as a regulator of stable transgenerational gene expression such that mutation of the CCAAT box results in variegated transgenerational inheritance. Considering that the CCAAT box is present in 30% of eukaryotic promoters, this study provides insights into how fidelity of gene expression patterns is maintained through multiple generations.

Pangenome-wide association study reveals the selective absence of CRISPR genes (Rv2816c-19c) in drug-resistant Mycobacterium tuberculosis.

The presence of intermittently dispersed insertion sequences and transposases in the Mycobacterium tuberculosis (Mtb) genome makes intra-genome recombination events inevitable. Understanding their effect on the gene repertoires (GR), which may contribute to the development of drug-resistant Mtb, is critical. In this study, publicly available WGS data of clinical Mtb isolates (endemic region n = 2,601; non-endemic region n = 1,130) were de novo assembled, filtered, scaffolded into assemblies, and functionally annotated. Out of 2,601 Mtb WGS data sets from endemic regions, 2,184 (drug resistant/sensitive: 1,386/798) qualified as high quality. We identified 3,784 core genes, 123 softcore genes, 224 shell genes, and 762 cloud genes in the pangenome of Mtb clinical isolates from endemic regions. Sets of 33 and 39 genes showed positive and negative associations (P < 0.01) with drug resistance status, respectively. Gene ontology clustering showed compromised immunity to phages and impaired DNA repair in drug-resistant Mtb clinical isolates compared to the sensitive ones. Multidrug efflux pump repressor genes (Rv3830c and Rv3855c) and CRISPR genes (Rv2816c-19c) were absent in the drug-resistant Mtb. A separate WGS data analysis of drug-resistant Mtb clinical isolates from the Netherlands (n = 1130) also showed the absence of CRISPR genes (Rv2816c-17c). This study highlights the role of CRISPR genes in drug resistance development in Mtb clinical isolates and helps in understanding its evolutionary trajectory and as useful targets for diagnostics development.IMPORTANCEThe results from the present Pan-GWAS study comparing gene sets in drug-resistant and drug-sensitive Mtb clinical isolates revealed intricate presence-absence patterns of genes encoding DNA-binding proteins having gene regulatory as well as DNA modification and DNA repair roles. Apart from the genes with known functions, some uncharacterized and hypothetical genes that seem to have a potential role in drug resistance development in Mtb were identified. We have been able to extrapolate many findings of the present study with the existing literature on the molecular aspects of drug-resistant Mtb, further strengthening the relevance of the results presented in this study.

Discovery and Accurate Detection of Rare Nucleic Acid Modifications

AbstractNucleic acid modifications play essential roles in diverse biological processes, ranging from gene expression regulation to stress response. While traditional research focused on common modifications like methylation, recent discoveries are unveiling a wide range of rare modifications with potentially crucial functions. However, accurately detecting and mapping these modifications pose significant challenges due to their low abundance and diverse chemical properties. This article summarizes the recent discoveries of rare DNA and RNA modifications across various organisms, highlighting their potential biological significance. Furthermore, it critically evaluates the limitations of current mapping techniques, including potential sources of false positives and negatives. Finally, the article discusses emerging strategies for overcoming these challenges and future opportunities in the field of rare nucleic acid modification detection.

Detection of DNA N6-Methyladenine Modification through SMRT-seq Features and Machine Learning Model

Introduction: N6-methyldeoxyadenine (6mA) is the most prevalent DNA modification in both prokaryotes and eukaryotes. While single-molecule real-time sequencing (SMRT-seq) can detect 6mA events at the individual nucleotide level, its practical application is hindered by a high rate of false positives. Methods: We propose a computational model for identifying DNA 6mA that incorporates comprehensive site features from SMRT-seq and employs machine learning classifiers. Results: The results demonstrate that 99.54% and 96.55% of the identified DNA 6mA instances in C.reinhardtii correspond with motifs and peak regions identified by methylated DNA immunoprecipitation sequencing (MeDIP-seq), respectively. Compared to SMRT-seq, the proportion of predicted DNA 6mA instances within MeDIP-seq peak regions increases by 2% to 70% across the six bacterial strains Conclusion: Our proposed method effectively reduces the false-positive rate in DNA 6mA prediction.

Selective non-Watson-Crick pairing-guided DNA synthesis for locus-specific analysis of 8-oxo-7,8-dihydroguanine in telomeres at ultrahigh resolution

8-Oxo-7,8-dihydroguanine (OG) is the most common oxidative modification in chromosomal DNA induced by reactive oxygen species (ROS). OG damage promotes telomere shortening and dysfunction, contributing to numerous degenerative/aging-related diseases and cancers. However, few methods are available for locus-specific quantification of OG in telomeres because of trace level of OG in DNA and low sensitivity of existing assays. Herein, we demonstrate selective non-Watson-Crick pairing-guided DNA synthesis for locus-specific analysis of OG in telomeres at ultrahigh resolution. When target telomere-OG is present, it hybridizes with the padlock probe to generate a one-base gap opposite OG. Deoxyadenosine triphosphate (dATP) is then incorporated into the gap by Bsu DNA polymerase (Bsu Pol) to mediate single-base extension. Through toehold-mediated strand displacement (TMSD), the primer probe displaces telomere-OG, and then it hybridizes with the extended padlock probe to induce proximity ligation-dependent cyclization and subsequent RCA with the aid of Taq ligase and phi29 polymerase, producing numerous ssDNAs that can bind with SYBR Green II to generate a significant fluorescence signal. This method can detect telomere-OG with a limit of detection (LOD) of 3.50 × 10−18 M in vitro and 1.38 × 10−3 ng in vivo under hydrogen peroxide (H2O2) stress. Moreover, it can discriminate as little as 0.001 % telomere-OG in excessive undamaged DNAs, and quantify the OG level in telomere DNAs from diverse human cell lines under oxidative stresses. Notably, this method can be rapidly implemented in one tube under isothermal conditions, without any labeled probes, complex steps, and high sample consumption, facilitating oxidative damage-related biomedical research and clinical diagnosis.

Epigenetic remodeling in insect immune memory.

The innate immune system of insects can respond more swiftly and efficiently to pathogens based on previous experience of encountering antigens. The understanding of molecular mechanisms governing immune priming, a form of immune memory in insects, including its transgenerational inheritance, remains elusive. It is still unclear if the enhanced expression of immune genes observed in primed insects can persist and be regulated through changes in chromatin structure via epigenetic modifications of DNA or histones, mirroring observations in mammals. Increasing experimental evidence suggests that epigenetic changes at the level of DNA/RNA methylation and histone acetylation can modulate the activation of insects' immune responses to pathogen exposure. Moreover, transgenerational inheritance of certain epigenetic modifications in model insect hosts can influence the transmission of pre-programmed immune responses to the offspring, leading to the development of evolved resistance. Epigenetic research in model insect hosts is on the brink of significant progress in the mechanistic understanding of chromatin remodeling within innate immunity, particularly the direct relationships between immunological priming and epigenetic alterations. In this review, we discuss the latest discoveries concerning the involvement of DNA methylation and histone acetylation in shaping the development, maintenance, and inheritance of immune memory in insects, culminating in the evolution of resistance against pathogens.

TB and HIV induced immunosenescence: where do vaccines play a role?

This paper tackles the complex interplay between Human Immunodeficiency virus (HIV-1) and Mycobacterium tuberculosis (M. tuberculosis) infections, particularly their contribution to immunosenescence, the age-related decline in immune function. Using the current literature, we discuss the immunological mechanisms behind TB and HIV-induced immunosenescence and critically evaluate the BCG (Bacillus Calmette-Guérin) vaccine's role. Both HIV-1 and M. tuberculosis demonstrably accelerate immunosenescence: M. tuberculosis through DNA modification and heightened inflammation, and HIV-1 through chronic immune activation and T cell production compromise. HIV-1 and M. tuberculosis co-infection further hastens immunosenescence by affecting T cell differentiation, underscoring the need for prevention and treatment. Furthermore, the use of the BCG tuberculosis vaccine is contraindicated in patients who are HIV positive and there is a lack of investigation regarding the use of this vaccine in patients who develop HIV co-infection with possible immunosenescence. As HIV does not currently have a vaccine, we focus our review more so on the BCG vaccine response as a result of immunosenescence. We found that there are overall limitations with the BCG vaccine, one of which is that it cannot necessarily prevent re-occurrence of infection due to effects of immunosenescence or protect the elderly due to this reason. Overall, there is conflicting evidence to show the vaccine's usage due to factors involving its production and administration. Further research into developing a vaccine for HIV and improving the BCG vaccine is warranted to expand scientific understanding for public health and beyond.

PT-seq: A method for metagenomic analysis of phosphorothioate epigenetics in complex microbial communities.

Among dozens of known epigenetic marks, naturally occurring phosphorothioate (PT) DNA modifications are unique in replacing a non-bridging phosphate oxygen with redox-active sulfur and function in prokaryotic restriction-modification and transcriptional regulation. Interest in PTs has grown due to the widespread distribution of the dnd, ssp, and brx genes among bacteria and archaea, as well as the discovery of PTs in 5-10% of gut microbes. Efforts to map PTs in complex microbiomes using existing next-generation and direct sequencing technologies have failed due to poor sensitivity. Here we developed PT-seq as a high-sensitivity method to quantitatively map PTs across genomes and metagenomically identify PT-containing microbes in complex genomic mixtures. Like other methods for mapping PTs in individual genomes, PT-seq exploits targeted DNA strand cleavage at PTs by iodine, followed by sequencing library construction using ligation or template switching approaches. However, PT-specific sequencing reads are dramatically increased by adding steps to heat denature the DNA, block pre-existing 3'-ends, fragment DNA after T-tailing, and enrich iodine-induced breaks using biotin-labeling and streptavidin beads capture. Iterative optimization of the sensitivity and specificity of PT-seq is demonstrated with individual bacteria and human fecal DNA.