Podcasts about nucleosomes

rWotD Episode 2770: HIST1H2AE Welcome to Random Wiki of the Day, your journey through Wikipedia’s vast and varied content, one random article at a time.The random article for Tuesday, 3 December 2024 is HIST1H2AE.Histone H2A type 1-B/E is a protein that in humans is encoded by the HIST1H2AE gene.Histones are basic nuclear proteins that are responsible for the nucleosome structure of the chromosomal fiber in eukaryotes. Nucleosomes consist of approximately 146 bp of DNA wrapped around a histone octamer composed of pairs of each of the four core histones (H2A, H2B, H3, and H4). The chromatin fiber is further compacted through the interaction of a linker histone, H1, with the DNA between the nucleosomes to form higher order chromatin structures. This gene is intronless and encodes a member of the histone H2A family. Transcripts from this gene lack polyA tails; instead, they contain a palindromic termination element. This gene is found in the large histone gene cluster on chromosome 6p22-p21.3.This recording reflects the Wikipedia text as of 00:57 UTC on Tuesday, 3 December 2024.For the full current version of the article, see HIST1H2AE on Wikipedia.This podcast uses content from Wikipedia under the Creative Commons Attribution-ShareAlike License.Visit our archives at wikioftheday.com and subscribe to stay updated on new episodes.Follow us on Mastodon at @wikioftheday@masto.ai.Also check out Curmudgeon's Corner, a current events podcast.Until next time, I'm neural Amy.

In this episode of the Epigenetics Podcast, we caught up with Lucas Farnung from Harvard Medical School to talk about his work on the structural analysis of nucleosomes during transcription. Lucas Farnung started his scientific career in Patrick Cramer's lab, trying to solve the cryo-EM structure of RNA polymerase II transcribing through a nucleosome. This project spanned some time before being published in 2018, during which time Dr. Farnung accomplished several other goals. The team solved the cryo-electron microscopy structure of Chd1 from the yeast Saccharomyces cerevisiae bound to a nucleosome at a resolution of 4.8 Å, solved the structure of the nucleosome-CHD4 chromatin remodeler, and investigated the structural basis of nucleosome transcription mediated by Chd1 and FACT. In 2021, he started his own lab and is now working on structural analysis of nucleosomes during transcription and how chromatin remodelers work on the chromatin template. References Farnung, L., Vos, S. M., Wigge, C., & Cramer, P. (2017). Nucleosome-Chd1 structure and implications for chromatin remodelling. Nature, 550(7677), 539–542. https://doi.org/10.1038/nature24046 Farnung, L., Vos, S. M., & Cramer, P. (2018). Structure of transcribing RNA polymerase II-nucleosome complex. Nature communications, 9(1), 5432. https://doi.org/10.1038/s41467-018-07870-y Filipovski, M., Soffers, J. H. M., Vos, S. M., & Farnung, L. (2022). Structural basis of nucleosome retention during transcription elongation. Science (New York, N.Y.), 376(6599), 1313–1316. https://doi.org/10.1126/science.abo3851   Related Episodes Molecular Mechanisms of Chromatin Modifying Enzymes (Karim-Jean Armache) Regulation of Chromatin Organization by Histone Chaperones (Geneviève Almouzni) Transcription Elongation Control by the Paf1 Complex (Karen Arndt) From Nucleosome Structure to Function (Karolin Luger)   Contact Epigenetics Podcast on Twitter Epigenetics Podcast on Instagram Epigenetics Podcast on Mastodon Active Motif on Twitter Active Motif on LinkedIn Email: podcast@activemotif.com

In this episode of the Epigenetics Podcast, we caught up with Dr. Srinivas Ramachandran, Assistant Professor at the University of Colorado, Anschutz Medical Campus, to talk about his work on ​in vivo nucleosome structure and dynamics. Dr. Srinivas Ramachandran studies the structure and dynamics of nucleosomes during cellular processes like transcription and DNA replication. During transcription, as the RNA polymerase transcribes along the DNA, it needs to pass nucleosomes. Dr. Ramachandran investigated the effect of nucleosomes on transcription and also studied how different histone variants affect this process. He found that the first nucleosome within a gene body is a barrier for the progression of RNA polymerase, and that presence of the histone variant H2A.Z in this first nucleosome lowers this barrier. Furthermore, Dr. Ramachandran developed a method called mapping in vivo nascent chromatin using EdU and sequencing (MINCE-Seq), enabling the study of chromatin landscapes right after DNA replication. In MINCE-Seq, newly replicated DNA is labeled right after the replication fork has passed by with the nucleotide analog ethynyl deoxyuridine (EdU), which can then be coupled with biotin using click chemistry. After the purification of newly replicated DNA and MNase digestion, the chromatin landscape can be analyzed. In this interview, we discuss the story behind how Dr. Ramachandran found his way into chromatin research, what it was like to start a wet lab postdoc with a bioinformatics background, and what he is working on now to unravel nucleosomal structure and dynamics in his own lab.   References Christopher M. Weber, Srinivas Ramachandran, Steven Henikoff (2014) Nucleosomes are context-specific, H2A.Z-modulated barriers to RNA polymerase (Molecular Cell) DOI: 10.1016/j.molcel.2014.02.014 Srinivas Ramachandran, Steven Henikoff (2016) Transcriptional Regulators Compete with Nucleosomes Post-replication (Cell) DOI: 10.1016/j.cell.2016.02.062 Srinivas Ramachandran, Kami Ahmad, Steven Henikoff (2017) Transcription and Remodeling Produce Asymmetrically Unwrapped Nucleosomal Intermediates (Molecular Cell) DOI: 10.1016/j.molcel.2017.11.015 Satyanarayan Rao, Kami Ahmad, Srinivas Ramachandran (2020) Cooperative Binding of Transcription Factors is a Hallmark of Active Enhancers (bioRxiv) DOI: 10.1101/2020.08.17.253146 Contact Active Motif on Twitter Epigenetics Podcast on Twitter Active Motif on LinkedIn Active Motif on Facebook Email: podcast@activemotif.com

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.11.03.366690v1?rss=1 Authors: Oberbeckmann, E., Krietenstein, N., Niebauer, V., Wang, Y., Schall, K., Moldt, M., Straub, T., Rohs, R., Hopfner, K.-P., Korber, P., Eustermann, S. Abstract: The fundamental molecular determinants by which ATP-dependent chromatin remodelers organize nucleosomes across eukaryotic genomes remain largely elusive. Here, chromatin reconstitutions on physiological, whole-genome templates reveal how remodelers read and translate genomic information into nucleosome positions. Using the yeast genome and the multi-subunit INO80 remodeler as a paradigm, we identify DNA shape/mechanics encoded signature motifs as sufficient for nucleosome positioning and distinct from known DNA sequence preferences of histones. INO80 processes such information through an allosteric interplay between its core- and Arp8-modules that probes mechanical properties of nucleosomal and linker DNA. At promoters, INO80 integrates this readout of DNA shape/mechanics with a readout of co-evolved sequence motifs via interaction with general regulatory factors bound to these motifs. Our findings establish a molecular mechanism for robust and yet adjustable +1 nucleosome positioning and, more generally, remodelers as information processing hubs that enable active organization and allosteric regulation of the first level of chromatin. Copy rights belong to original authors. Visit the link for more info

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.10.30.360990v1?rss=1 Authors: Peng, Y., Li, S., Onufriev, A., Landsman, D., Panchenko, A. R. Abstract: Despite histone tails' critical roles in epigenetic regulation, little is known about mechanisms of how histone tails modulate the nucleosomal DNA solvent accessibility and recognition of nucleosomes by other macromolecules. Here we generate extensive atomic level conformational ensembles of histone tails in the context of the full human nucleosome, totaling 26 microseconds of molecular dynamics simulations. We explore the histone tail binding with the nucleosomal and linker DNA and observe rapid conformational transitions between bound and unbound states allowing us to estimate kinetic and thermodynamic properties of the histone tail-DNA interactions. Different histone types exhibit distinct, although conformationally heterogeneous, binding modes and each histone type occludes specific DNA regions from the solvent. Using a comprehensive set of experimental data on nucleosome structural complexes, we find that majority of the studied nucleosome-binding proteins and histone tails target mutually exclusive regions on nucleosomal or linker DNA around the super-helical locations {+/-}1, {+/-}2, and {+/-}7. This finding is explained within the generalized competitive binding and tail displacement models of partners recruitment to nucleosomes. Finally, we demonstrate the crosstalk between different histone post-translational modifications, where charge-altering modifications and mutations typically suppress tail-DNA interactions and enhance histone tail dynamics. Copy rights belong to original authors. Visit the link for more info

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.09.08.287656v1?rss=1 Authors: Schachner, L. F., Jooss, K., Morgan, M. A., Piunti, A., Meiners, M. J., Lee, A., Kafader, J. O., Iwanaszko, M., Cheek, M. A., Burg, J. M., Howard, S. A., Keogh, M.-C., Shilatifard, A., Kelleher, N. L. Abstract: Nuc-MS characterizes histone modifications and variants directly from intact endogenous nucleosomes. Preserving whole nucleosome particles enables precise interrogation of their protein content, as for H3.3-containing nucleosomes which had 6-fold co-enrichment of variant H2A.Z over bulk chromatin. Nuc-MS, validated by ChIP-seq, showed co-occurrence of oncogenic H3.3K27M with euchromatic marks (e.g., H4K16ac and >15-fold enrichment of H3K79me2). By capturing the entire epigenetic landscape, Nuc-MS provides a new, quantitative readout of nucleosome-level biology. Copy rights belong to original authors. Visit the link for more info

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.09.04.282921v1?rss=1 Authors: Huertas, J., Schöler, H. R., Cojocaru, V. Abstract: Genomic DNA is packaged in chromatin, a dynamic polymer that forms fibers variable in size and compaction. Chromatin's monomeric unit, the nucleosome, is formed by wrapping ~ 146 basepairs of DNA around histone proteins. Dynamic transitions in the chromatin structure, which depend on intra- and inter-nucleosome dynamics and on chemical modifications of histones, are key to initiating gene regulation programs. The histone modifications occur in unstructured terminal regions known as tails and are key to epigenetic regulation of gene expression. Here we elucidate how the interplay between the tails of two histones, H3 and H2A modulate ample breathing motions of genomic nucleosomes. From 18 s of atomistic molecular dynamics simulations of two genomic and one engineered nucleosomes, we observed two rare, large opening events, one in each genomic nucleosome, and only small amplitude breathing in the engineered nucleosome. Clustering of the sampled structures revealed a strong correlation between DNA conformations with different degree of opening and histone tail conformations and positions. The ample nucleosome motions were caused by changes in the interaction profiles between the DNA and clusters of lysine and arginine residues in these tails, many of which are targets for epigenetic regulatory modifications. Given that histone tail modulated nucleosome breathing can change the structure and compaction of chromatin fibers, the mechanism we describe provides the basis for understanding chromatin dynamics at atomic resolution. Copy rights belong to original authors. Visit the link for more info

Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.07.30.228908v1?rss=1 Authors: Rousova, D., Funk, S. K., Reichle, H., Weir, J. R. Abstract: One of the defining features of sexual reproduction is the recombination events that take place during meiosis I. Recombination is both evolutionarily advantageous, but also mechanistically necessary to form the crossovers that link homologous chromosomes. Meiotic recombination is initiated through the placement of programmed double-strand DNA breaks (DSBs) mediated by the protein Spo11. The timing, number, and physical placement of DSBs are carefully controlled through a variety of protein machinery. Previous work has implicated Mer2(IHO1 in mammals) to be involved in both the placement of breaks, and their timing. In this study we use a combination of protein biochemistry and biophysics to extensively characterise various roles of the Mer2. We gain further insights into the details of Mer2 interaction with the PHD protein Spp1, reveal that Mer2 is a novel nucleosome binder, and suggest how Mer2's interaction with the HORMA domain protein Hop1 (HORMAD1/2 in mammals) is controlled. Copy rights belong to original authors. Visit the link for more info

The Nucleosome is the basic building unit of chromatin. It consists out of 147 base pairs of double stranded DNA wrapped around the Histone core octamer that consists out of 2 copies of each dimer of H2A/H2B, and H3/H4. Nucleosomes are organized like "beads on a string" to form a modifiable regulatory basis for higher order structures of chromatin. The first images of the nucleosome as a particle was published by our guests Ada and Don Olins from the University of New England in 1974 (Olins, A. L. & Olins, D. E. Spheroid Chromatin Units (ν Bodies). Science 183, 330–332 (1974).). This observation lead the way to numerous discoveries around chromatin which ultimately culminated in the discovery of the 2.8 Angstrom high-resolution crystal structure 20 years ago in the year 1997 (Luger, K., Mäder, A. W., Richmond, R. K., Sargent, D. F. & Richmond, T. J. Crystal structure of the nucleosome core particle at 2.8 Å resolution. Nature 389, 251–260 (1997).) References for this episode Ada L. Olins, Donald E. Olins. Spheroid Chromatin Units (ν Bodies). Science. 25 Jan 1974: Vol. 183, Issue 4122, pp. 330-332. DOI: 10.1126/science.183.4122.330 Ada L. Olins, Donald E. Olins, et al. An epichromatin epitope. Nucleus. 2011 Jan-Feb; 2(1): 47–60. DOI: 10.4161/nucl.2.1.13271 Active Motif Contact Details Follow us on Twitter Join us on LinkedIn Like us on Facebook Email us @Active Motif Europe or Active Motif North America.

Eukaryotic genomes are organized inside the cell nucleus in a structured macromolecular DNA-protein polymer named chromatin, formed by single discrete unites called Nucleosomes. The packing of the genetic information into chromatin allows the efficient regulation of several nuclear processes, such as gene expression and transcription, DNA replication, cell cycle progression, chromosome segregation and DNA damage repair. Chromatin comes in two flavors: a transcriptionally active, more loosened state, called euchromatin and a transcriptionally silent or low expressed, more compact state, called heterochromatin. The assembly of silent chromatin or heterochromatin is fundamental for the regulation of every nuclear process and it is driven in most Eukaryotes by the deposition and the read-out of the histone H3 lysine 9 methylation (H3K9me) post-translational modification (PTM). H3K9me on the nucleosome is specifically bound by chromatin readers called chromodomains (CD) and this recognition is fundamental for the downstream processes that lead to the formation of heterochromatin and shut down the expression of single genes or entire gene clusters. Despite several studies have been done on different chromodomains binding to H3K9me histone tail peptides, to date there was no structural information on how chromodomains interact with their natural binding partners, the H3K9me3 Nucleosomes. In a preliminary structural study carried out in our laboratory we solved the cryo-electron microscopy (Cryo-EM) structure of the chromodomain of the fission yeast Chp1 protein (Chp1CD) in complex with an H3K9me nucleosome. The structure showed that the Chp1CD interacts not only with the histone H3 tail but also with the histone globular domains in the Nucleosome core, primarily with histone H3. Mutations in the residues of Chp1CD that form the binding interface with the Nucleosome core (two loops in the β-sheet of the domain) caused a drop of the affinity in vitro for the H3K9me Nucleosome, which was independent from the histone H3K9me tail interaction. Cells harboring the same Chp1CD loop mutations were defective in silencing centromeric transcripts and maintain the deposition of the H3K9me mark for heterochromatin formation. This indicated that Chp1CD-nucleosome core interaction is fundamental for heterochromatin formation in fission yeast and opened up to the possibility that chromodomains could read multiple histone PTMs, on both the recruiting histone tail and on the nucleosome core. This study substantially contributes to understand how chromodomains interact with chromatin, how much the nucleosome core interaction is conserved among different CDs and how different chromodomain proteins are regulated at the same loci. Understanding how chromodomain readers recognize nucleosomes is fundamental to uncover the basics of gene silencing and heterochromatin formation.

Summary: Structure of the FACT chaperone domain in complex with histones H2A-H2B, and a model for FACT-mediated nucleosome reorganization Nucleosomes are the smalles unit of chromatin: two coils of DNA are wrapped around a histone octamer core, which neutralizes its charge and `packs' the lengthy molecule. Nucleosomes confer a barrier to processes that require access to the eukaryotic genome such as transcription, DNA replication and repair. A variety of nucleosome remodeling machines and histone chaperones facilitate nucleosome dynamics by depositing or evicting histones and unwrapping the DNA. The eukaryotic FACT complex (composed of the subunits Spt16 and Pob3) is an essential and highly conserved chaperone. It assists the progression of DNA and RNA polymerases, for example by facilitating transcriptional initiation and elongation. Further, it promotes the genome-wide integrity of chromatin structure, including the suppression of cryptic transcription. Genetic and biochemical assays have shown that FACT's chaperone activity is crucially mediated by a direct interaction with histones H2A-H2B. However, the structural basis for how H2A-H2B are recognized and how this integrates with FACT’s other functions, including the recognition of histones H3-H4 and of other nuclear factors, is unknown. In my PhD research project, I was able to reveal the structure of the yeast chaperone domain in complex with the H2A-H2B heterodimer and show that the Spt16M module in FACT’s Spt16 subunit establishes the evolutionarily conserved H2A-H2B binding and chaperoning function. The structure shows how an alpha-helical `U-turn' motif in Spt16M interacts with the alpha-1-helix of H2B. The U-turn motif scaffolds onto a tandem pleckstrin-homology-like (PHL) module, which is structurally and functionally related to the H3-H4 chaperone Rtt106 and the Pob3M domain of FACT. Biochemical and in vivo assays validate the crystal structure and dissect the contribution of histone tails and H3-H4 toward FACT binding. My results show that Spt16M makes multiple interactions with histones, which I suggest allow the module to gradually invade the nucleosome and ultimately block the strongest interaction surface of H2B with nucleosomal DNA by binding the H2B alpha-1-helix. Together, these multiple contact points establish an extended surface that could reorganize the first 30 base-pairs of nucleosomal histone–DNA contacts. Further, I report a brief biochemical analysis of FACT’s heterodimerization domain. Its PHL fold indicates shared evolutionary origin with the H3-H4-binding Spt16M, Pob3M and Rtt106 tandem PHL modules. However, the Spt16D–Pob3N heterodimer does not bind histones, rather it connects FACT to replicative DNA polymerases. The snapshots of FACT’s engagement with H2A-H2B and structure-function analysis of all its domains lay the foundation for the systematic analysis of FACT’s vital chaperoning functions and how the complex promotes the activity of enzymes that require nucleosome reorganization.

Background Selective Internal Radiation Therapy (SIRT) is a new and effective locoregional anticancer therapy for colorectal cancer patients with liver metastases. Markers for prediction of therapy response and prognosis are needed for the individual management of those patients undergoing SIRT. Methods Blood samples were prospectively and consecutively taken from 49 colorectal cancer patients with extensive hepatic metastases before, three, six, 24 and 48 h after SIRT to analyze the concentrations of nucleosomes and further laboratory parameters, and to compare them with the response to therapy regularly determined 3 months after therapy and with overall survival. Results Circulating nucleosomes, cytokeratin-19 fragments (CYFRA 21-1), carcinoembryonic antigen (CEA), C-reactive protein (CRP) and various liver markers increased already 24 h after SIRT. Pretherapeutical levels of CYFRA 21-1, CEA, cancer antigen 19-9 (CA 19-9), asparate-aminotransferase (AST) and lactate dehydrogenase (LDH) as well as 24 h values of nucleosomes were significantly higher in patients suffering from disease progression (N = 35) than in non-progressive patients (N = 14). Concerning overall survival, CEA, CA 19-9, CYFRA 21-1, CRP, LDH, AST, choline esterase (CHE), gamma-glutamyl-transferase, alkaline phosphatase, and amylase (all 0 h, 24 h) and nucleosomes (24 h) were found to be prognostic relevant markers in univariate analyses. In multivariate Cox-Regression analysis, the best prognostic model was obtained for the combination of CRP and AST. When 24 h values were additionally included, nucleosomes (24 h) further improved the existing model. Conclusion Panels of biochemical markers are helpful to stratify pretherapeutically colorectal cancer patients for SIR-therapy and to early estimate the response to SIR-therapy.

Background: As cell-free circulating DNA exists predominantly as mono-and oligonucleosomes, the focus of the current study was to examine the interplay of circulating nucleosomes, DNA, proteases and caspases in blood of patients with benign and malignant breast diseases. Methods: The concentrations of cell-free DNA and nucleosomes as well as the protease and caspase activities were measured in serum of patients with benign breast disease (n = 20), primary breast cancer (M0, n = 31), metastatic breast cancer (M1, n = 32), and healthy individuals (n = 28) by PicoGreen, Cell Death Detection ELISA, Protease Fluorescent Detection Kit and Caspase-Glo (R) 3/7 Assay, respectively. Results: Patients with benign and malignant tumors had significantly higher levels of circulating nucleic acids in their blood than healthy individuals (p = 0.001, p = 0.0001), whereas these levels could not discriminate between benign and malignant lesions. Our analyses of all serum samples revealed significant correlations of circulating nucleosome with DNA concentrations (p = 0.001), nucleosome concentrations with caspase activities (p = 0.008), and caspase with protease activities (p = 0.0001). High serum levels of protease and caspase activities associated with advanced tumor stages (p = 0.009). Patients with lymph node-positive breast cancer had significantly higher nucleosome levels in their blood than node-negative patients (p = 0.004). The presence of distant metastases associated with a significant increase in serum nucleosome (p = 0.01) and DNA levels (p = 0.04), and protease activities (p = 0.008). Conclusion: Our findings demonstrate that high circulating nucleic acid concentrations in blood are no indicators of a malignant breast tumor. However, the observed changes in apoptosis-related deregulation of proteolytic activities along with the elevated serum levels of nucleosomes and DNA in blood are linked to breast cancer progression.

Background: Transarterial chemoembolization (TACE) therapy is an effective locoregional treatment in hepatocellular cancer (HCC) patients. For early modification of therapy, markers predicting therapy response are urgently required. Methods: Here, sera of 50 prospectively and consecutively included HCC patients undergoing 71 TACE therapies were taken before and 3 h, 6 h and 24 h after TACE application to analyze concentrations of circulating nucleosomes, cytokeratin-19 fragments (CYFRA 21-1), alpha fetoprotein (AFP), C-reactive protein (CRP) and several liver biomarkers, and to compare these with radiological response to therapy. Results: While nucleosomes, CYFRA 21-1, CRP and some liver biomarkers increased already 24 h after TACE, percental changes of nucleosome concentrations before and 24 h after TACE and pre- and posttherapeutic values of AFP, gamma-glutamyl-transferase (GGT) and alkaline phosphatase (AP) significantly indicated the later therapy response (39 progression versus 32 no progression). In multivariate analysis, nucleosomes (24 h), AP (24 h) and TACE number were independent predictive markers. The risk score of this combination model achieved an AUC of 81.8% in receiver operating characteristic (ROC) curves and a sensitivity for prediction of non-response to therapy of 41% at 97% specificity, and of 72% at 78% specificity. Conclusion: Circulating nucleosomes and liver markers are valuable tools for early estimation of the efficacy of TACE therapy in HCC patients.

The positions of nucleosomes in eukaryotic genomes determine which parts of the DNA sequence are readily accessible for regulatory proteins and which are not. Genome-wide maps of nucleosome positions have revealed a salient pattern around transcription start sites, involving a nucleosome-free region (NFR) flanked by a pronounced periodic pattern in the average nucleosome density. While the periodic pattern clearly reflects well-positioned nucleosomes, the positioning mechanism is less clear. A recent experimental study by Mavrich et al. argued that the pattern observed in Saccharomyces cerevisiae is qualitatively consistent with a "barrier nucleosome model," in which the oscillatory pattern is created by the statistical positioning mechanism of Kornberg and Stryer. On the other hand, there is clear evidence for intrinsic sequence preferences of nucleosomes, and it is unclear to what extent these sequence preferences affect the observed pattern. To test the barrier nucleosome model, we quantitatively analyze yeast nucleosome positioning data both up- and downstream from NFRs. Our analysis is based on the Tonks model of statistical physics which quantifies the interplay between the excluded-volume interaction of nucleosomes and their positional entropy. We find that although the typical patterns on the two sides of the NFR are different, they are both quantitatively described by the same physical model with the same parameters, but different boundary conditions. The inferred boundary conditions suggest that the first nucleosome downstream from the NFR (the +1 nucleosome) is typically directly positioned while the first nucleosome upstream is statistically positioned via a nucleosome-repelling DNA region. These boundary conditions, which can be locally encoded into the genome sequence, significantly shape the statistical distribution of nucleosomes over a range of up to approximately 1,000 bp to each side.

Background: Nucleosomes are cell death products that are elevated in serum of patients with diseases that are associated with massive cell destruction. We investigated the kinetics of circulating nucleosomes after cerebral stroke and their correlation with the clinical status. Methods: In total, we analyzed nucleosomes by ELISA in sera of 63 patients with early stroke daily during the first week after onset. For correlation with the clinical pathology, patients were grouped into those with medium to slight functional impairment (Barthel Index BI >= 50) and those with severe functional impairment (BI < 50). Results: Patients with BI >= 50 showed a continuous increase in nucleosomes until day 5 (median: 523 arbitrary units, AU) followed by a slow decline. In contrast, patients with BI < 50 showed a steeper initial increase reaching a maximum already on day 3 (869 AU). Both, days after stroke (p < 0.001) and BI (p < 0.001), had a significant influence on nucleosome concentrations, respectively. Consistently, patients with BI < 50 had a significantly larger area under the curve (AUC/day) of nucleosome values during the first week after stroke (800 AU) than patients with BI >= 50 (497 AU; p = 0.031). Concerning the infarction volume, nucleosomes showed significant correlations for the concentrations on day 3 (r = 0.43; p = 0.001) and for the area under the curve (r = 0.34; p = 0.016). Conclusion: Even if nucleosomes are nonspecific cell death markers, their release into serum after cerebral stroke correlates with the gross functional status as well as with the infarction volume and can be considered as biochemical correlative to the severity of stroke. Copyright (c) 2006 S. Karger AG, Basel.

Apoptotic markers and tumor-associated antigens might be suitable to indicate the response to radiochemotherapy early. We analyzed the courses of nucleosomes, CEA, CA 19-9 and CYFRA 21-1 in 25 colorectal cancer patients during radiochemotherapy (4 postoperative, 13 preoperative, 8 local relapse therapy). Blood was taken before therapy, daily during the first week, once weekly during the following weeks, and at the end of the radiochemotherapy. After a temporary decline 6 h after the first irradiation, nucleosomes rose in most patients rapidly reaching a maximum during the first days which was followed by a subsequent decrease. In patients receiving postoperative therapy after complete resection of tumor, nucleosome levels generally were lower than in patients with preoperative or relapse therapy. Correspondingly, CEA, CA 19-9 and CYFRA 21-1 levels of postoperatively treated patients were the lowest whereas those with tumor relapse had the highest ones. During preoperative therapy, lower nucleosome concentrations were found in patients with response to therapy resulting in a smaller area under the curve of days 1-3 (AUC) than in those with progressive disease (p = 0.028). The other parameters did not indicate the response to therapy at the initial treatment phase. In conclusion, the course of nucleosomes (AUC) might be valuable for the early prediction of therapy response in preoperatively treated colorectal cancer patients. Copyright (c) 2006 S. Karger AG, Basel.

Nucleosomes appear spontaneously in elevated concentrations in the serum of patients with malignant diseases as well as during chemo- and radiotherapy. We analyzed whether their kinetics show typical characteristics during radiochemotherapy and enable an early estimation of therapy efficacy. We used the Cell Death Detection Elisaplus ( Roche Diagnostics) and investigated the course of nucleosomes in the serum of 32 patients with a local stage of pancreatic cancer who were treated with radiochemotherapy for several weeks. Ten of them received postsurgical therapy, 21 received primary therapy and 1 received therapy for local relapse. Blood was taken before the beginning of therapy, daily during the first week, once weekly during the following weeks and at the end of radiochemotherapy. The response to therapy was defined according to the kinetics of CA 19-9: a decrease of CA 19-9 650% after radiochemotherapy was considered as `remission'; an increase of >= 100% ( which was confirmed by two following values) was defined as `progression'. Patients with `stable disease' ranged intermediately. Most of the examined patients showed a decrease of the concentration of nucleosomes within 6 h after the first dose of radiation. Afterwards, nucleosome levels increased rapidly, reaching their maximum during the following days. Patients receiving postsurgery, primary or relapse therapies did not show significant differences in nucleosome values during the time of treatment. Single nucleosome values, measured at 6, 24 and 48 h after the application of therapy, could not discriminate significantly between patients with no progression and those with progression of disease. However, the area under the curve of the first 3 days, which integrated all variables of the initial therapeutic phase, showed a significant correlation with the progression-free interval ( p = 0.008). Our results indicate that the area under the curve of nucleosomes during the initial phase of radiochemotherapy could be valuable for the early prediction of the progression-free interval. Copyright (C) 2005 S. Karger AG, Basel.

Nucleosomes, which are typical cell death products, are elevated in the serum of cancer patients and are known to rapidly increase during radiotherapy. As both normal and malignant cells are damaged by irradiation, we investigated to which extent both cell types contribute to the release of nucleosomes. We cultured monolayers of normal bronchoepithelial lung cells (BEAS-2B, n = 18) and epithelial lung cancer cells (EPLC, n = 18), exposed them to various radiation doses (0, 10 and 30 Gy) and observed them for 5 days. Culture medium was changed every 24 h. Subsequently, nucleosomes were determined in the supernatant by the Cell Death Detection-ELISA(plus) ( Roche Diagnostics). Additionally, the cell number was estimated after harvesting the cells in a second preparation. After 5 days, the cell number of BEAS-2B cultures in the irradiated groups (10 Gy: median 0.03 x 10(6) cells/culture, range 0.02-0.08 x 10(6) cells/culture; 30 Gy: median 0.08 x 10(6) cells/culture, range 0.02-0.14 x 10(6) cells/culture) decreased significantly (10 Gy: p = 0.005; 30 Gy p = 0.005; Wilcoxon test) compared to the non-irradiated control group (median 4.81 x 10(6) cells/culture, range 1.50-9.54 x 10(6) cells/culture). Consistently, nucleosomes remained low in the supernatant of nonirradiated BEAS-2B. However, at 10 Gy, BEAS-2B showed a considerably increasing release of nucleosomes, with a maximum at 72 h ( before irradiation: 0.24 x 10(3) arbitrary units, AU, range 0.13-4.09 x 10(3) AU, and after 72 h: 1.94 x 10(3) AU, range 0.11-5.70 x 10(3) AU). At 30 Gy, the release was even stronger, reaching the maximum earlier (at 48 h, 11.09 x 10(3) AU, range 6.89-18.28 x 10(3) AU). In non-irradiated EPLC, nucleosomes constantly increased slightly. At 10 Gy, we observed a considerably higher release of nucleosomes in EPLC, with a maximum at 72 h (before irradiation: 2.79 x 10(3) AU, range 2.42-3.80 x 10(3) AU, and after 72 h: 7.16 x 10(3) AU, range 4.30-16.20 x 10(3) AU), which was more than 3.5 times higher than in BEAS-2B. At 30 Gy, the maximum (6.22 x 10(3) AU, range 5.13-9.71 x 10(3) AU) was observed already after 24 h. These results indicate that normal bronchoepithelial and malignant lung cancer cells contribute to the release of nucleosomes during irradiation in a dose-and time-dependent manner with cancer cells having a stronger impact at low doses. Copyright (C) 2004 S. Karger AG, Basel.

The concentration of nucleosomes is elevated in blood of patients with diseases which are associated with enhanced cell death. In order to detect these circulating nucleosomes, we used the Cell Death Detection-ELISA(Plus) (CDDE) from Roche Diagnostics (Mannheim, Germany) (details at http:textbackslash{}textbackslash{}biochem.roche.com). For its application in liquid materials we performed various modifications: we introduced a standard curve with nucleosome-rich material, which enabled direct quantification and improved comparability of the values within (CVinterassay:3.0-4.1%) and between several runs (CVinterassay:8.6-13.5%), and tested the analytical specificity of the ELISA. Because of the fast elimination of nucleosomes from circulation and their limited stability, we compared plasma and serum matrix and investigated in detail the pre-analytical handling of serum samples which can considerably influence the test results. Careless venipuncture producing hemolysis, delayed centrifugation and bacterial contamination of the blood samples led to false-positive results; delayed stabilization with EDTA and insufficient storage conditions resulted in false-negative values. At temperatures of -20 degreesC, serum samples which were treated with 10 mM EDTA were stable for at least 6 months. In order to avoid possible interfering factors, we recommend a schedule for the pre-analytical handling of the samples. As the first stage, the possible clinical application was investigated in the sera of 310 persons. Patients with solid tumors (n = 220; mean = 361 Arbitrary Units (AU)) had considerably higher values than healthy persons (n = 50; mean = 30 AU; P = 0.0001) and patients with inflammatory diseases (n = 40; mean = 296 AU; p = 0.096). Within the group of patients with tumors, those in advanced stages (UICC 4) showed significantly higher values than those in early stages (UICC 1-3) (P = 0.0004).

Sat, 1 Jan 1994 12:00:00 +0100 https://epub.ub.uni-muenchen.de/7388/1/transcriptional_repression_by_nucleosomes_7388.pdf Becker, Peter B.; Blank, T.; Sandaltzopoulos, R.