Search Results (80)

The most serious complications of systemic sclerosis (SSc) and mixed connective tissue disease (MCTD) include lung fibrosis (LF) and pulmonary hypertension (PH). The aim of this study was to find any association between the serological profile and the incidence of these complications. The tested group included 121 persons (87 SSc, 34 MCTD); mean age 55.6 ± 13.4 years. Patients were qualified for the LF presence group based on HRCT. Likelihood of PH was determined using echocardiography. The presence of antinuclear antibodies (ANA) was assessed using indirect immunofluorescence, ANA-profile, sclerosis-profile (using EUROIMMUN kits), and antiphospholipid antibodies (aPL) (using the ELISA method). Distribution of individual antibody types was at a level similar to the previously described groups in the Polish population and differed from the American and African population. A positive correlation was found between LF and the presence of anti-Scl-70 (p = 0.024) antibodies, negative correlation was found between LF and the presence of anti-histone (p = 0.03), anti-centromere A (p = 0.009), anti-centromere B (p = 0.014), and anti-nucleosomes (p = 0.03) antibodies. No correlation between the presence of aPL and the above complications was found. The prevalence of individual antibody types in SSc and MCTD may have ethnic and geographical grounds. Scl-70 antibodies correlate positively with LF. Anti-centromere, anti-histone, and anti-nucleosome antibodies reduce its risk. No correlation between aPL and the occurrence of LF and elevated PH risk was found. Full article

The DNA base-excision repair (BER) pathway shares the second part of its enzymatic chain with the single-strand break (SSB) repair pathway. BER is initiated by a glycosylase, such as UDG, while SSBR is initiated by the multifunctional enzyme PARP1. The very early steps in the identification of the DNA damage are crucial to the correct initiation of the repair chains, and become even more complex when considering the realistic environment of damage to the DNA in the nucleosome. We performed molecular dynamics computer simulations of the interaction between the glycosylase UDG and a mutated uracil (as could result from oxidative deamination of cytosine), and between the Zn1-Zn2 fragment of PARP1 and a simulated SSB. The model system is a whole nucleosome in which DNA damage is inserted at various typical positions along the 145-bp sequence. It is shown that damage recognition by the enzymes requires very strict conditions, unlikely to be matched by pure random search along the DNA. We propose that mechanical deformation of the DNA around the defective sites may help signaling the presence of the defect, accelerating the search process. Full article

The maintenance of genome stability and the prevention of genotoxic damage to DNA require immediate DNA repair. In the cell, the repair process is usually preceded by a release of DNA from complexes with chromatin proteins accompanied by nucleosome sliding, relaxing or disassembly. Base excision DNA repair (BER) corrects the most common DNA lesions, which does not disturb the DNA helix dramatically. Notably, small DNA lesions can be repaired in chromatin without global chromatin decompaction. One of the regulatory mechanisms is poly(ADP-ribosyl)ation, leading to the relaxation of the nucleosome. In our work, we demonstrated that recently a discovered protein, HPF1, can modulate the efficiency of one of the key BER stages—DNA synthesis—via the regulation of total poly(ADP-ribosyl)ation. Accordingly, we investigated both short-patch and long-patch DNA synthesis catalyzed by DNA polymerase β (pol β; main polymerase in BER) and showed that HPF1’s influence on the poly(ADP-ribosyl)ation catalyzed by PARP1 and especially by PARP2 results in more efficient DNA synthesis in the case of the short-patch BER pathway in nucleosomes. Additionally, HPF1-dependent poly(ADP-ribosyl)ation was found to positively regulate long-patch BER. Full article

In eukaryotic nuclei, DNA is wrapped around an octamer of core histones to form nucleosomes. H1 binds to the linker DNA of nucleosome to form the chromatosome, the next structural unit of chromatin. Structural features on individual chromatosomes contribute to chromatin structure, but not fully characterized. In addition to canonical nucleosomes composed of two copies each of histones H2A, H2B, H3, and H4 (H3 nucleosomes), centromeres chromatin contain nucleosomes in which H3 is replaced with its analog CENP-A, changing structural properties of CENP-A nucleosomes. Nothing is known about the interaction of H1 with CENP-A nucleosomes. Here we filled this gap and characterized the interaction of H1 histone with both types of nucleosomes. H1 does bind both types of the nucleosomes forming more compact chromosome particles with elevated affinity to H3 nucleosomes. H1 binding significantly increases the stability of chromatosomes preventing their spontaneous dissociation. In addition to binding to the entry-exit position of the DNA arms identified earlier, H1 is capable of bridging of distant DNA segments. H1 binding leads to the assembly of mononucleosomes in aggregates, stabilized by internucleosome interactions as well as bridging of the DNA arms of chromatosomes. Contribution of these finding to the chromatin structure and functions are discussed. Full article

(1) Background: Nucleosomes represent the essential structural units of chromatin and serve as key regulators of cell function and gene expression. Oocytes in the germinal vesicle (GV) stage will later undergo meiosis and become haploid cells ready for fertilization, while somatic cells undergo mitosis after DNA replication. (2) Purpose: To furnish theoretical insights and data that support the process of cell reprogramming after nuclear transplantation. (3) Methods: We compared the nucleosome occupancy, distribution, and transcription of genes between two types of cells: fully grown GV oocytes from big follicles (BF) and somatic cells (porcine embryonic fibroblast, PEF). (4) Results: The nucleosome occupancy in the promoter of BF was 4.85%, which was significantly higher than that of 3.3% in PEF (p < 0.05), and the nucleosome distribution showed a noticeable increase surrounding transcriptional start sites (TSSs) in BF. Next, we reanalyzed the currently published transcriptome of fully grown GV oocytes and PEF, and a total of 51 genes in BF and 80 genes in PEF were identified as being uniquely expressed. The nucleosome distribution around gene TSSs correlated with expression levels in somatic cells, yet the results in BF differed from those in PEF. (5) Conclusion: This study uncovers the dynamic nature and significance of nucleosome positioning and chromatin organization across various cell types, providing a basis for nuclear transplantation. Full article

Background/Objectives: Transgene applications, ranging from gene therapy to the development of stable cell lines and organisms, rely on maintaining the expression of transgenes. To date, the use of plasmid-based transgenes has been limited by the loss of their expression shortly after their delivery into the target cells. The short-lived expression of plasmid-based transgenes has been largely attributed to host-cell-mediated degradation and/or silencing of transgenes. The development of chromatin-based strategies for gene delivery has the potential to facilitate defining the requirements for establishing epigenetic states and to enhance transgene expression for numerous applications. Methods: To assess the impact of “priming” plasmid-based transgenes to adopt accessible chromatin states to promote gene expression, nucleosome positioning elements were introduced at promoters of transgenes, and vectors were pre-assembled into nucleosomes containing unmodified histones or mutants mimicking constitutively acetylated states at residues 9 and 14 of histone H3 or residue 16 of histone H4 prior to their introduction into cells, then the transgene expression was monitored over time. Results: DNA sequences capable of positioning nucleosomes could positively impact the expression of adjacent transgenes in a distance-dependent manner in the absence of their pre-assembly into chromatin. Intriguingly, the pre-assembly of plasmids into chromatin facilitated the prolonged expression of transgenes relative to plasmids that were not pre-packaged into chromatin. Interactions between pre-assembled chromatin states and nucleosome positioning-derived effects on expression were also assessed and, generally, nucleosome positioning played the predominant role in influencing gene expression relative to priming with hyperacetylated chromatin states. Conclusions: Strategies incorporating nucleosome positioning elements and the pre-assembly of plasmids into chromatin prior to nuclear delivery can modulate the expression of plasmid-based transgenes. Full article

Background: Lupus nephritis (LN) is an inflammation of the kidneys that is related to systemic lupus erythematosus (SLE). This study aimed to evaluate the differences in clinical and laboratory characteristics between LN and non-LN SLE patients. Methods: We conducted a retrospective analysis of medical records collected from SLE patients treated at the University Hospital in Kraków, Poland, from 2012 to 2022. All patients met the 2019 European League Against Rheumatism and the American College of Rheumatology (EULAR/ACR) criteria for SLE. Results: Among 921 SLE patients, LN was documented in 331 (35.94%). LN patients were younger at SLE diagnosis (29 vs. 37 years; p < 0.001) and had a male proportion that was 2.09 times higher than the non-LN group (16.62% vs. 7.97%; p < 0.001). They were more often diagnosed with serositis and hematological or neurological involvement (p < 0.001 for all). Hypertension and hypercholesterolemia occurred more frequently in these patients (p < 0.001 for both). LN patients exhibited a higher frequency of anti-dsDNA, anti-histone, and anti-nucleosome antibodies (p < 0.001 for all). Conversely, the non-LN group had a 1.24-fold (95% CI: 1.03–1.50; p = 0.021) increase in the odds ratio of having positive anti-cardiolipin IgM antibody results. LN patients were more frequently treated with immunosuppressants. The risk factors for experiencing at least three LN flares included female sex, younger age at the onset of LN or SLE, LN occurring later than SLE onset, the presence of anti-nucleosome or anti-dsDNA antibodies, and certain SLE manifestations such as myalgia, arthritis, proteinuria > 3.5 g/day, and pathological urinary casts in the urine sediment. Conclusions: LN patients differ from non-LN patients in the age of SLE diagnosis, treatment modalities, and autoantibody profile and have more frequent, severe manifestations of SLE. However, we still need more prospective studies to understand the diversity of LN and its progression in SLE patients. Full article

Technological advancements in cell-free DNA (cfDNA) liquid biopsy have triggered exponential growth in numerous clinical applications. While cfDNA-based liquid biopsy has made significant strides in personalizing cancer treatment, the exploration and translation of epigenetics in liquid biopsy to clinical practice is still nascent. This comprehensive review seeks to provide a broad yet in-depth narrative of the present status of epigenetics in cfDNA liquid biopsy and its associated challenges. It highlights the potential of epigenetics in cfDNA liquid biopsy technologies with the hopes of enhancing its clinical translation. The momentum of cfDNA liquid biopsy technologies in recent years has propelled epigenetics to the forefront of molecular biology. We have only begun to reveal the true potential of epigenetics in both our understanding of disease and leveraging epigenetics in the diagnostic and therapeutic domains. Recent clinical applications of epigenetics-based cfDNA liquid biopsy revolve around DNA methylation in screening and early cancer detection, leading to the development of multi-cancer early detection tests and the capability to pinpoint tissues of origin. The clinical application of epigenetics in cfDNA liquid biopsy in minimal residual disease, monitoring, and surveillance are at their initial stages. A notable advancement in fragmentation patterns analysis has created a new avenue for epigenetic biomarkers. However, the widespread application of cfDNA liquid biopsy has many challenges, including biomarker sensitivity, specificity, logistics including infrastructure and personnel, data processing, handling, results interpretation, accessibility, and cost effectiveness. Exploring and translating epigenetics in cfDNA liquid biopsy technology can transform our understanding and perception of cancer prevention and management. cfDNA liquid biopsy has great potential in precision oncology to revolutionize conventional ways of early cancer detection, monitoring residual disease, treatment response, surveillance, and drug development. Adapting the implementation of liquid biopsy workflow to the local policy worldwide and developing point-of-care testing holds great potential to overcome global cancer disparity and improve cancer outcomes. Full article

Periodontitis is a chronic inflammatory disease. We have previously shown that salivary DNA is higher in patients with periodontitis. Neutrophil extracellular traps (NETs) are involved in the pathogenesis of chronic inflammatory diseases. The objective of this case–control study was to compare patients with periodontitis and healthy controls regarding the salivary concentrations of extracellular DNA and NET components. Unstimulated saliva samples were collected from 49 patients with periodontitis and 71 controls before an oral examination. Salivary extracellular DNA was isolated and quantified fluorometrically and using PCR. NET-associated markers were assessed using ELISA. We have found significantly higher concentrations of salivary extracellular DNA in samples from periodontitis patients (five-times higher for supernatant and three times for pellet). Our results show that patients also have three-times-higher salivary nucleosomes and NET-associated enzymes—myeloperoxidase and neutrophil elastase (both two-times higher). Neutrophil elastase and salivary DNA in the pellet correlated positively with the pocket depth/clinical attachment level in periodontitis patients (r = 0.31—weak correlation; p = 0.03 and r = 0.41—moderate correlation, p = 0.004). Correlations between salivary extracellular DNA and NET enzymes were positive and significant. Based on our results, the higher salivary extracellular DNA in periodontitis seems to be related to components of NETs, albeit with weak to moderate correlations indicating that NETs are produced in periodontitis and can play a role in its pathogenesis similarly to other inflammatory diseases. Further studies should prove this assumption with potential diagnostic and therapeutic consequences. Full article

The coordination of activities between nuclei and organelles in plant cells involves information exchange, in which phytohormones may play essential roles. Therefore, the dissection of the mechanisms of hormone-related integration between phytohormones and mitochondria is an important and challenging task. Here, we found that inputs from multiple hormones may cause changes in the transcript accumulation of mitochondrial-encoded genes and nuclear genes encoding mitochondrial (mt) proteins. In particular, treatments with exogenous hormones induced changes in the GUS expression in the reporter line possessing a 5′-deletion fragment of the RPOTmp promoter. These changes corresponded in part to the up- or downregulation of RPOTmp in wild-type plants, which affects the transcription of mt-encoded genes, implying that the promoter fragment of the RPOTmp gene is functionally involved in the responses to IAA (indole-3-acetic acid), ACC (1-aminocyclopropane-1-carboxylic acid), and ABA (abscisic acid). Hormone-dependent modulations in the expression of mt-encoded genes can also be mediated through mitochondrial transcription termination factors 15, 17, and 18 of the mTERF family and genes for tetratricopeptide repeat proteins that are coexpressed with mTERF genes, in addition to SWIB5 encoding a mitochondrial SWI/SNF (nucleosome remodeling) complex B protein. These genes specifically respond to hormone treatment, displaying both negative and positive regulation in a context-dependent manner. According to bioinformatic resources, their promoter region possesses putative cis-acting elements involved in responses to phytohormones. Alternatively, the hormone-related transcriptional activity of these genes may be modulated indirectly, which is especially relevant for brassinosteroids (BS). In general, the results of this study indicate that hormones are essential mediators that are able to cause alterations in the transcript accumulation of mt-related nuclear genes, which, in turn, trigger the expression of mt genes. Full article

Chromatin is now regarded as a heterogeneous and dynamic structure occupying a non-random position within the cell nucleus, where it plays a key role in regulating various functions of the genome. This current view of chromatin has emerged thanks to high spatiotemporal resolution imaging, among other new technologies developed in the last decade. In addition to challenging early assumptions of chromatin being regular and static, high spatiotemporal resolution imaging made it possible to visualize and characterize different chromatin structures such as clutches, domains and compartments. More specifically, super-resolution microscopy facilitates the study of different cellular processes at a nucleosome scale, providing a multi-scale view of chromatin behavior within the nucleus in different environments. In this review, we describe recent imaging techniques to study the dynamic organization of chromatin at high spatiotemporal resolution. We also discuss recent findings, elucidated by these techniques, on the chromatin landscape during different cellular processes, with an emphasis on the DNA damage response. Full article

Nucleosomes not only serve as the basic building blocks for eukaryotic chromatin but also regulate many biological processes, such as DNA replication, repair, and recombination. To modulate gene expression in vivo, the histone variant H2A.Z can be dynamically incorporated into the nucleosome. However, the assembly dynamics of H2A.Z-containing nucleosomes remain elusive. Here, we demonstrate that our previous chemical kinetic model for nucleosome assembly can be extended to H2A.Z-containing nucleosome assembly processes. The efficiency of H2A.Z-containing nucleosome assembly, like that of canonical nucleosome assembly, was also positively correlated with the total histone octamer concentration, reaction rate constant, and reaction time. We expanded the kinetic model to represent the competitive dynamics of H2A and H2A.Z in nucleosome assembly, thus providing a novel method through which to assess the competitive ability of histones to assemble nucleosomes. Based on this model, we confirmed that histone H2A has a higher competitive ability to assemble nucleosomes in vitro than histone H2A.Z. Our competitive kinetic model and experimental results also confirmed that in vitro H2A.Z-containing nucleosome assembly is governed by chemical kinetic principles. Full article

All species are well adapted to their environment. Stress causes a magnitude of biochemical and molecular responses in plants, leading to physiological or pathological changes. The response to various stresses is genetically predetermined, but is also controlled on the epigenetic level. Most plants are adapted to their environments through generations of exposure to all elements. Many plant species have the capacity to acclimate or adapt to certain stresses using the mechanism of priming. In most cases, priming is a somatic response allowing plants to deal with the same or similar stress more efficiently, with fewer resources diverted from growth and development. Priming likely relies on multiple mechanisms, but the differential expression of non-coding RNAs, changes in DNA methylation, histone modifications, and nucleosome repositioning play a crucial role. Specifically, we emphasize the role of BRM/CHR17, BRU1, FGT1, HFSA2, and H2A.Z proteins as positive regulators, and CAF-1, MOM1, DDM1, and SGS3 as potential negative regulators of somatic stress memory. In this review, we will discuss the role of epigenetic factors in response to stress, priming, and the somatic memory of stress exposures. Full article

The establishment and maintenance of nucleosome-free regions (NFRs) are prominent processes within chromatin dynamics. Transcription factors, ATP-dependent chromatin remodeling complexes (CRCs) and DNA sequences are the main factors involved. In Saccharomyces cerevisiae, CRCs such as RSC contribute to chromatin opening at NFRs, while other complexes, including ISW1a, contribute to NFR shrinking. Regarding DNA sequences, growing evidence points to poly(dA:dT) tracts as playing a direct role in active processes involved in nucleosome positioning dynamics. Intriguingly, poly(dA:dT)-tract-containing NFRs span asymmetrically relative to the location of the tract by a currently unknown mechanism. In order to obtain insight into the role of poly(dA:dT) tracts in nucleosome remodeling, we performed a systematic analysis of their influence on the activity of ISW1a and RSC complexes. Our results show that poly(dA:dT) tracts differentially affect the activity of these CRCs. Moreover, we found differences between the effects exerted by the two alternative tract orientations. Remarkably, tract-containing linker DNA is taken as exit DNA for nucleosome sliding catalyzed by RSC. Our findings show that defined DNA sequences, when present in linker DNA, can dictate in which direction a remodeling complex has to slide nucleosomes and shed light into the mechanisms underlying asymmetrical chromatin opening around poly(dA:dT) tracts. Full article

Much of today’s molecular science revolves around next-generation sequencing. Frequently, the first step in analyzing such data is aligning sequencing reads to a reference genome. This step is often taken for granted, but any analysis downstream of the alignment will be affected by the aligner’s ability to correctly map sequences. In most cases, for research into chromatin structure and nucleosome positioning, ATAC-seq, ChIP-seq, and MNase-seq experiments use short read lengths. How well aligners manage these reads is critical. Most aligner programs will output mapped reads and unmapped reads. However, from a biological point of view, reads will fall into one of three categories: correctly mapped, incorrectly mapped, and unmapped. While increased sequencing depth can often compensate for unmapped reads, incorrectly and correctly mapped reads appear algorithmically identical but can produce biologically significant alterations in the results. For this reason, we are benchmarking various alignment programs to determine their propensity to incorrectly map short reads. As short-read alignment is an important step in ATAC-seq, ChIP-seq, and MNase-seq experiments, caution should be taken in mapping reads to ensure that the most accurate conclusions can be made from the data generated. Our analysis is intended to help investigators new to the field pick the alignment program best suited for their experimental conditions. In general, the aligners we tested performed well. BWA, Bowtie2, and Chromap were all exceptionally accurate, and we recommend using them. Furthermore, we show that longer read lengths do in fact lead to more accurate mappings. Full article