cytosine methylation
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2022 ◽  
Marco Regolini

Every adult male of the little roundworm Caenorhabditis elegans is always and invariably comprised of exactly 1031 somatic cells, not one more, not one less; and so it is for the adult hermaphrodite (959 somatic cells); its intestine founder cell (the ‘E’ blastomere), if isolated and cultured, undergoes the same number of divisions as in the whole embryo (Robertson et al., 2014); the zygote of Drosophila melanogaster executes 13 cycles of asynchronous cell divisions without cellularization: how are these numbers counted? Artificial Intelligence (First and Second Order Logic, Knowledge graph Engineering) infers that, to perform precise stereotypical numbers of asynchronous cell divisions, a nucleic (genomic) counter is indispensable. Made up of tandemly repeated similar monomers, satellite DNA (satDNA) corresponds to iterable objects used in programming. The purpose of this article is to show how satDNA sequences can be iterated over to count a deterministic number of cell divisions: computational models (attached for free download) are introduced that handle DNA repeated sequences as iterable counters and simulate their use in cells through an epigenetic marker (cytosine methylation) as an iterator. SatDNA, because of its propensity to remodel its structure, can also operate as a strong accelerator in the evolution of complex organs and provides a basis to control interspecific variability of shapes.

2022 ◽  
Vol 13 (1) ◽  
Steve Horvath ◽  
Amin Haghani ◽  
Sichong Peng ◽  
Erin N. Hales ◽  
Joseph A. Zoller ◽  

AbstractCytosine methylation patterns have not yet been thoroughly studied in horses. Here, we profile n = 333 samples from 42 horse tissue types at loci that are highly conserved between mammalian species using a custom array (HorvathMammalMethylChip40). Using the blood and liver tissues from horses, we develop five epigenetic aging clocks: a multi-tissue clock, a blood clock, a liver clock and two dual-species clocks that apply to both horses and humans. In addition, using blood methylation data from three additional equid species (plains zebra, Grevy’s zebras and Somali asses), we develop another clock that applies across all equid species. Castration does not significantly impact the epigenetic aging rate of blood or liver samples from horses. Methylation and RNA data from the same tissues define the relationship between methylation and RNA expression across horse tissues. We expect that the multi-tissue atlas will become a valuable resource.

2022 ◽  
Vol 12 ◽  
Hideki Takahashi ◽  
Midori Tabara ◽  
Shuhei Miyashita ◽  
Sugihiro Ando ◽  
Shuichi Kawano ◽  

A cucumber mosaic virus isolate, named Ho [CMV(Ho)], was isolated from a symptomless Arabidopsis halleri field sample containing low virus titers. An analysis of CMV(Ho) RNA molecules indicated that the virus isolate, besides the usual cucumovirus tripartite RNA genome, additionally contained defective RNA3 molecules and a satellite RNA. To study the underlying mechanism of the persistent CMV(Ho) infection in perennial A. halleri, infectious cDNA clones were generated for all its genetic elements. CMV, which consists of synthetic transcripts from the infectious tripartite RNA genomes, and designated CMV(Ho)tr, multiplied in A. halleri and annual Arabidopsis thaliana Col-0 to a similar level as the virulent strain CMV(Y), but did not induce any symptoms in them. The response of Col-0 to a series of reassortant CMVs between CMV(Ho)tr and CMV(Y) suggested that the establishment of an asymptomatic phenotype of CMV(Ho) infection was due to the 2b gene of CMV RNA2, but not due to the presence of the defective RNA3 and satellite RNA. The accumulation of CMV(Ho) 2b protein tagged with the FLAG epitope (2b.Ho-FLAG) in 2b.Ho-FLAG-transformed Col-0 did not induce any symptoms, suggesting a 2b-dependent persistency of CMV(Ho)tr infection in Arabidopsis. The 2b protein interacted with Argonaute 4, which is known to regulate the cytosine methylation levels of host genomic DNA. Whole genomic bisulfite sequencing analysis of CMV(Ho)tr- and mock-inoculated Col-0 revealed that cytosine hypomethylation in the promoter regions of 82 genes, including two genes encoding transcriptional regulators (DOF1.7 and CBP1), was induced in response to CMV(Ho)tr infection. Moreover, the increased levels of hypomethylation in the promoter region of both genes, during CMV(Ho)tr infection, were correlated with the up- or down-regulation of their expression. Taken altogether, the results indicate that during persistent CMV(Ho) infection in Arabidopsis, host gene expression may be epigenetically modulated resulting from a 2b-mediated cytosine hypomethylation of host genomic DNA.

2021 ◽  
Artur Scherf ◽  
Elie Hammam ◽  
Samia Miled ◽  
Frederic Bonhomme ◽  
Benoit Arcangioli ◽  

DNA cytosine methylation and its oxidized products are important epigenetic modifications in mammalian cells. Although 5-methylcytosine (5mC) was detected in the human malaria parasite Plasmodium falciparum, the presence of oxidized 5mC forms remain to be characterized.Here we establish a protocol to explore nuclease-based DNA digestion for the extremely AT-rich genome of P. falciparum (>80% A+T) for quantitative LC-MS/MS analysis of 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). We demonstrate the presence of 5hmC, 5fC and 5caC cytosine modifications in a DNMT2-only organism and observe striking ratio changes between 5mC and 5hmC during the 48-hour blood stage parasite development. Parasite-infected red blood cells cultured in different physiological oxygen concentrations revealed a shift in the cytosine modifications distribution towards the oxidized 5hmC and 5caC forms. In the absence of the canonical C5-DNA methyltransferase (DNMT1 and DNMT3A/B) in P. falciparum, we show that all cytosine modifications depend on the presence of DNMT2. We conclude that DNMT2 and oxygen levels are critical determinants that shape the dynamic cytosine epigenetic landscape in this human pathogen.

2021 ◽  
Vol 22 (23) ◽  
pp. 13159
Hijiri Hasegawa ◽  
Ikkei Sasaki ◽  
Kaori Tsukakoshi ◽  
Yue Ma ◽  
Kazuo Nagasawa ◽  

Genomic DNA methylation is involved in many diseases and is expected to be a specific biomarker for even the pre-symptomatic diagnosis of many diseases. Thus, a rapid and inexpensive detection method is required for disease diagnosis. We have previously reported that cytosine methylation in G-quadruplex (G4)-forming oligonucleotides develops different G4 topologies. In this study, we developed a method for detecting CpG methylation in G4-forming oligonucleotides based on the structural differences between methylated and unmethylated G4 DNAs. The differences in G4 topologies due to CpG methylation can be discriminated by G4 ligands. We performed a binding assay between methylated or unmethylated G4 DNAs and G4 ligands. The binding abilities of fluorescent G4 ligands to BCL-2, HRAS1, HRAS2, VEGF G4-forming sequences were examined by fluorescence-based microtiter plate assay. The differences in fluorescence intensities between methylated and unmethylated G4 DNAs were statistically significant. In addition to fluorescence detection, the binding of G4 ligand to DNA was detected by chemiluminescence. A significant difference was also detected in chemiluminescence intensity between methylated and unmethylated DNA. This is the first study on the detection of CpG methylation in G4 structures, focusing on structural changes using G4 ligands.

Elizabeth B Lewis ◽  
Edwin Chen ◽  
Matthew J Culyba

Abstract The bacterial DNA damage response pathway (SOS response) is composed of a network of genes regulated by a single transcriptional repressor, LexA. The lexA promoter, itself, contains two LexA operators, enabling negative feedback. In Escherichia coli, the downstream operator contains a conserved DNA cytosine methyltransferase (Dcm) site that is predicted to be methylated to 5-methylcytosine (5mC) specifically during stationary phase growth, suggesting a regulatory role for DNA methylation in the SOS response. To test this, we quantified 5mC at the lexA locus, and then examined the effect of LexA on Dcm activity, as well as the impact of this 5mC mark on LexA binding, lexA transcription, and SOS response induction. We found that 5mC at the lexA promoter is specific to stationary phase growth, but that it does not affect lexA expression. Our data support a model where LexA binding at the promoter inhibits Dcm activity without an effect on the SOS regulon.

2021 ◽  
Vol 22 (22) ◽  
pp. 12594
Virginia Veronica Visconti ◽  
Federica Centofanti ◽  
Simona Fittipaldi ◽  
Elisa Macrì ◽  
Giuseppe Novelli ◽  

Myotonic dystrophy type 1 and 2 (DM1 and DM2) are two multisystemic autosomal dominant disorders with clinical and genetic similarities. The prevailing paradigm for DMs is that they are mediated by an in trans toxic RNA mechanism, triggered by untranslated CTG and CCTG repeat expansions in the DMPK and CNBP genes for DM1 and DM2, respectively. Nevertheless, increasing evidences suggest that epigenetics can also play a role in the pathogenesis of both diseases. In this review, we discuss the available information on epigenetic mechanisms that could contribute to the DMs outcome and progression. Changes in DNA cytosine methylation, chromatin remodeling and expression of regulatory noncoding RNAs are described, with the intent of depicting an epigenetic signature of DMs. Epigenetic biomarkers have a strong potential for clinical application since they could be used as targets for therapeutic interventions avoiding changes in DNA sequences. Moreover, understanding their clinical significance may serve as a diagnostic indicator in genetic counselling in order to improve genotype–phenotype correlations in DM patients.

2021 ◽  
Vol 12 ◽  
Corien M. Voorburg ◽  
Yuling Bai ◽  
Richard Kormelink

Ty-1 presents an atypical dominant resistance gene that codes for an RNA-dependent RNA polymerase (RDR) of the gamma class and confers resistance to tomato yellow leaf curl virus (TYLCV) and other geminiviruses. Tomato lines bearing Ty-1 not only produce relatively higher amounts of viral small interfering (vsi)RNAs, but viral DNA also exhibits a higher amount of cytosine methylation. Whether Ty-1 specifically enhances posttranscriptional gene silencing (PTGS), leading to a degradation of RNA target molecules and primarily relying on 21–22 nucleotides (nts) siRNAs, and/or transcriptional gene silencing (TGS), leading to the methylation of cytosines within DNA target sequences and relying on 24-nts siRNAs, was unknown. In this study, small RNAs were isolated from systemically TYLCV-infected leaves of Ty-1 encoding tomato plants and susceptible tomato Moneymaker (MM) and sequence analyzed. While in susceptible tomato plants vsiRNAs of the 21-nt size class were predominant, their amount was drastically reduced in tomato containing Ty-1. The latter, instead, revealed elevated levels of vsiRNAs of the 22- and 24-nt size classes. In addition, the genomic distribution profiles of the vsiRNAs were changed in Ty-1 plants compared with those from susceptible MM. In MM three clear hotspots were seen, but these were less pronounced in Ty-1 plants, likely due to enhanced transitive silencing to neighboring viral genomic sequences. The largest increase in the amount of vsiRNAs was observed in the intergenic region and the V1 viral gene. The results suggest that Ty-1 enhances an antiviral TGS response. Whether the elevated levels of 22 nts vsiRNAs contribute to an enhanced PTGS response or an additional TGS response involving a noncanonical pathway of RNA dependent DNA methylation remains to be investigated.

2021 ◽  
Arindam Sikdar ◽  
Umanath Sharma ◽  
Rajesh Barua ◽  
Abir U. Igamberdiev ◽  
Samir C. Debnath

Abstract Epigenetic variation plays a role in developmental gene regulation and responses to the environment. An efficient interaction of zeatin induced cytosine methylation and secondary compounds has been displayed for the first time in tissue-culture shoots of lingonberry (Vaccinium vitis-idaea) in vitro, in vivo and its cutting-cultivar Erntedank. Through MSAP assay, we observed highest methylated sites in leaf regenerants (LC1) from all primer combinations (108 bands), with their highest variation in secondary metabolites. We measured that four tissue-culture plants showed higher methylation bands than cutting propagated donor plants (ED) which exhibited 79 bands of methylation, which is comparatively low. On the other hand, we observed the highest total phenolic content in node culture-derived greenhouse grown plants, NC3 but leaf culture-derived greenhouse grown plants, LC1 represented low phenolic content. Our study showed more methylation in micropropagated plants (NC1, NC2, NC3, LC1) than those derived from cutting propagated ED plants, where methylation was not present. On the contrary, we observed higher secondary metabolites in ED plants but comparatively less in micropropagated shoots (NC1, NC2) and plants (NC3, LC1). Our study displayed that higher methylation sites observed in micropropagated plants possessed less amount of secondary metabolites.

2021 ◽  
Vol 12 ◽  
Cun Rui ◽  
Yuexin Zhang ◽  
Yapeng Fan ◽  
Mingge Han ◽  
Maohua Dai ◽  

Gossypium barbadense is a cultivated cotton not only known for producing superior fiber but also for its salt and alkaline resistance. Here, we used Whole Genome Bisulfite Sequencing (WGBS) technology to map the cytosine methylation of the whole genome of the G. barbadense hypocotyl at single base resolution. The methylation sequencing results showed that the mapping rates of the three samples were 75.32, 77.54, and 77.94%, respectively. In addition, the Bisulfite Sequence (BS) conversion rate was 99.78%. Approximately 71.03, 53.87, and 6.26% of the cytosine were methylated at CG, CHG, and CHH sequence contexts, respectively. A comprehensive analysis of DNA methylation and transcriptome data showed that the methylation level of the promoter region was a positive correlation in the CHH context. Saline-alkaline stress was related to the methylation changes of many genes, transcription factors (TFs) and transposable elements (TEs), respectively. We explored the regulatory mechanism of DNA methylation in response to salt and alkaline stress during cotton hypocotyl elongation. Our data shed light into the relationship of methylation regulation at the germination stage of G. barbadense hypocotyl cell elongation and salt-alkali treatment. The results of this research help understand the early growth regulation mechanism of G. barbadense in response to abiotic stress.

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