Biology subjects

Urano, T.

Publications and source records attributed to Urano, T..

4 recordsLinked to original sources

bioRxiv2025

Locations of consecutive G·C base-pairs direct genomic nucleosome positioning

The nucleosome is a structural and functional subunit of chromatin, and its positioning and dynamics in the eukaryotic genomes serves as a key platform for gene regulation. Here, we determine the positions of fully wrapped nucleosomes across the yeast genome by chemical mapping through the histone H2A-A122C residue, which cleaves near the DNA entry/exit sites. This approach reveals the most refined sequence-dependent profile of nucleosomes reported to date. Comparison of H2A-A122C with the H3-Q85C and H4-S47C methods clearly shows a sequence preference for the chemical cleavage site, and mapping results suggest that the nucleosome locations are dynamic in vivo. More importantly, we find that the depletion of CC and GG dinucleotides at nucleotide positions -11 to -9 and +9 to +11 bp, respectively, from the nucleosome dyad position (0) is inextricably associated with the enrichment of AA/AT/TA/TT dinucleotides in both yeast and mouse genomes. Introducing consecutive C*G base pairs to the corresponding sites in the 601 sequence shifts the nucleosomes to less frequent positions without altering thermal stability, implying a structural constraint imposed by DNA sequence. Thus, CC and GG dinucleotides in the major groove blocks at superhelix locations (SHL) -1.0 and +1.0, respectively, destabilize histone-DNA interactions, serving as intrinsic determinants of nucleosome positioning in eukaryotic genomes.

molecular biology↗

bioRxiv2024

Microthrombi-growth in ADAMTS13 deficiency exacerbated ulcerative colitis via mucosal and endothelial dysfunction

BackgroundUlcerative colitis (UC) is a chronic inflammatory bowel disease characterized by mucosal inflammation and ulceration, with systemic immune dysregulation exacerbating disease progression. While the accumulation of von Willebrand factor (VWF), normally digested by a disintegrin-like and metalloproteinase with thrombospondin type 1 motif 13 (ADAMTS13) from ultra-large multimers to small molecules, has been implicated in UC pathogenesis, the association between aberrant microthrombus formation and colitis remains unclear. MethodsPlasma and inflamed colonic tissues from UC patients were analyzed. In the experimental colitis model, we employed an intravital imaging technique to reveal real-time structural dynamics of the mucus layer and the mucosal vasculature. Dextran sulfate sodium (DSS)-induced colitis in wild-type and ADAMTS13-deficient mice was evaluated for disease progression, mucus layer disruption, leukocyte recruitment, and thrombus formation in mucosal vessels using intravital multiphoton excitation microscopy within single-cell spatial resolution. ResultsUC patients exhibited significantly reduced plasma ADAMTS13 activity correlating with disease severity and excessive VWF deposition in inflamed colonic tissues. In DSS-induced colitis mice, ADAMTS13 deficiency showed heightened disease activity and increased mucosal erosion in histochemical analysis compared to wild-type mice. A novel methodology established in this study using intravital microscopy successfully appraised colonic mucus barrier integrity by visualizing fluorescent dextran penetration from the colonic lumen to the crypts. ADAMTS13 deficiency accelerated mucus layer disruption, leukocyte adhesion, and microthrombi formation, particularly close to crypt epithelium regions. Vessel-specific analyses demonstrated that obstructive microthrombi were most prominent in the mucosal layer, contributing to local ischemia and mucosal erosion. Therapeutical usage of recombinant human ADAMTS13 alleviated microthrombus formation, improved mucosal integrity, and mitigated colitis severity in wild-type and ADAMTS13-deficient mice. ConclusionsObstructive thrombi formed in mucosal vessels due to impaired ADAMTS13 activity appeared essential in the disease progression of UC. Advanced intravital imaging provided novel insights into single-cell resolution UC pathogenesis.

physiology↗

bioRxiv2023

Asymmetric patterns of nucleosome positioning sequences in protein-coding regions

Although nucleosome positioning is key to epigenetic regulation, how the DNA sequence contributes to positioning remains elusive, especially in the context of transcription direction. Analysis of nucleotide bases with respect to the nucleosomal DNA coordinates requires precise nucleosomal mapping information on the genome. However, currently available base-pair-resolution nucleosome maps based on cysteine-mediated chemical cleavage do not fully satisfy the requirement due to method-specific cleavage biases. Here, we generated a chimeric nucleosomal DNA model to achieve less-biased prediction. The model revealed that yeast protein-coding sequences have higher affinity for the promoter-proximal half of nucleosomes than for the distal half. Strikingly, peaks of calculated affinity scores for the promoter-proximal half periodically matched the first few nucleosome positions. Detailed analysis of nucleotide bases revealed that the AA dinucleotide in the left side of the top strand contributes to nucleosome detection frequency in intergenic regions, while the complementary dinucleotide TT is preferred in the other side. In contrast, the sense strand is AA-rich throughout the nucleosome coordinate in protein-coding regions, which is consistent with asymmetric affinity. These data suggest that eukaryotes have evolved DNA sequences with asymmetric affinity for nucleosome formation to maintain epigenetic integrity of protein-coding regions.

genomics↗

bioRxiv2019

Chemical map-based prediction of nucleosome positioning using the Bioconductor package nuCpos

BackgroundAssessing the nucleosome-forming potential of specific DNA sequences is important for understanding complex chromatin organization. Methods for predicting nucleosome positioning include bioinformatics and biophysical approaches. An advantage of bioinformatics methods, which are based on in vivo nucleosome maps, is the use of natural sequences that may contain previously unknown elements involved in nucleosome positioning in vivo. The accuracy of such prediction attempts reflects the genomic coordinate resolution of the nucleosome maps applied. Nucleosome maps are constructed using micrococcal nuclease digestion followed by high-throughput sequencing (MNase-seq). However, as MNase has a strong preference for A/T-rich sequences, MNase-seq may not be appropriate for this purpose. In addition to MNase-seq-based maps, base pair-resolution chemical maps of in vivo nucleosomes from three different species (budding and fission yeasts, and mice) are currently available. However, these chemical maps have yet to be integrated into publicly available computational methods. ResultsWe developed a Bioconductor package (named nuCpos) to demonstrate the superiority of chemical maps in predicting nucleosome positioning. The accuracy of chemical map-based prediction in rotational settings was higher than that of the previously developed MNase-seq-based approach. With our method, predicted nucleosome occupancy reasonably matched in vivo observations and was not affected by A/T nucleotide frequency. Effects of genetic alterations on nucleosome positioning that had been observed in living yeast cells could also be predicted. nuCpos calculates individual histone binding affinity (HBA) scores for given 147-bp sequences to examine their suitability for nucleosome formation. We also established local HBA as a new parameter to predict nucleosome formation, which was calculated for 13 overlapping nucleosomal DNA subsequences. HBA and local HBA scores for various sequences agreed well with previous in vitro and in vivo studies. Furthermore, our results suggest that nucleosomal subsegments that are disfavored in different rotational settings contribute to the defined positioning of nucleosomes. ConclusionsOur results demonstrate that chemical map-based statistical models are beneficial for studying nucleosomal DNA features. Studies employing nuCpos software can enhance understanding of chromatin regulation and the interpretation of genetic alterations and facilitate the design of artificial sequences.

bioinformatics↗