Biology subjects

Rosas, R.

Publications and source records attributed to Rosas, R..

4 recordsLinked to original sources

bioRxiv2025

G34R cancer mutation alters the conformational ensemble and dynamics of the histone H3.3 tails

In recent years, mutations in the histone variant H3.3 have been discovered in pediatric and adult gliomas and osteosarcomas. One of these is the G34R mutation in the H3.3 N-terminal tail. While this mutation is known to disrupt epigenomic pathways, the effects on nucleosome structure itself have not been explored. In light of recent studies, that have demonstrated that the interaction of the H3 tail with nucleosomal and linker DNA is driven in large part by arginine residues, we sought to determine if the G34R cancer mutation and adjacent G33R mutation, not observed in cancer, directly alter nucleosome structural dynamics. Using NMR spectroscopy and molecular dynamics simulations, we investigate the effects of these mutations on the H3 tail in the context of the nucleosome. Our results show that both of these mutations enhance the association of the H3 tails with DNA and decrease the conformational dynamics around the site of mutation. Moreover, our results reveal that both mutations lead to changes in the conformational ensemble of the entire tail, re-positioning it on the nucleosomal DNA as well as promoting intra-tail interactions. This has implications for the accessibility of the H3 tail to effector proteins and for changes in higher-order chromatin structure resulting from a shift in intra-versus inter-nucleosome contacts mediated by the H3 tail.

biophysics↗

bioRxiv2022

A novel Single Alpha-Helix-DNA-binding domain in CAF-1 promotes gene silencing and DNA damage survival through tetrasome-length DNA selectivity and spacer function

The histone chaperone chromatin assembly factor 1 (CAF-1) deposits nascent histone H3/H4 dimers onto newly replicated DNA forming the central core of the nucleosome known as the tetrasome. How CAF-1 ensures there is sufficient space for the assembly of tetrasomes remains unknown. Structural and biophysical characterization of the lysine/glutamic acid/arginine-rich (KER) region of CAF-1 revealed a 128 [A] single alpha helix (SAH) motif with unprecedented DNA binding properties. Distinct KER sequence features and length of the SAH drive the selectivity of CAF-1 for tetrasome-length DNA and facilitate function in budding yeast. In vivo, the KER cooperates with the DNA-binding winged helix domain in CAF-1 to overcome DNA damage sensitivity and maintain silencing of gene expression. We propose that the KER SAH links functional domains within CAF-1 with structural precision, acting as a DNA binding spacer element during chromatin assembly.

biochemistry↗

bioRxiv2020

The SEQC2 Epigenomics Quality Control (EpiQC) Study: Comprehensive Characterization of Epigenetic Methods, Reproducibility, and Quantification

Cytosine modifications in DNA such as 5-methylcytosine (5mC) underlie a broad range of developmental processes, maintain cellular lineage specification, and can define or stratify cancer and other diseases. However, the wide variety of approaches available to interrogate these modifications has created a need for harmonized materials, methods, and rigorous benchmarking to improve genome-wide methylome sequencing applications in clinical and basic research. Here, we present a multi-platform assessment and a global resource for epigenetics research from the FDAs Epigenomics Quality Control (EpiQC) Group. The study design leverages seven human cell lines that are designated as reference materials and publicly available from the National Institute of Standards and Technology (NIST) and Genome in a Bottle (GIAB) consortium. These samples were subject to a variety of genome-wide methylation interrogation approaches across six independent laboratories, with a primary focus was on 5-methylcytosine modifications. Each sample was processed in two or more technical replicates by three whole-genome bisulfite sequencing (WGBS) protocols (TruSeq DNA methylation, Accel-NGS MethylSeq, and SPLAT), oxidative bisulfite sequencing (TrueMethyl), one enzymatic deamination method (EMseq), targeted methylation sequencing (Illumina Methyl Capture EPIC), and single-molecule long-read nanopore sequencing from Oxford Nanopore Technologies. After rigorous quality assessment and comparison to Illumina EPIC methylation microarrays and testing on a range of algorithms (Bismark, BitmapperBS, BWAMeth, and GemBS), we found overall high concordance between assays (R=0.87-R0.93), differences in efficency of read mapping and CpG capture and coverage, and platform performance. The data provided herein can guide continued used of these reference materials in epigenomics assays, as well as provide best practices for epigenomics research and experimental design in future studies.

genomics↗

bioRxiv2019

A novel role for the tumor suppressor gene ITF2 in lung tumorigenesis through the action of Wnt signaling pathway

Despite often leading to platinum resistance, platinum-based chemotherapy continues to be the standard treatment for many epithelial tumors. In this study we analyze the cytogenetic alterations that arise after cisplatin treatment providing novel insights into the molecular biology and the cellular mechanisms involved in the acquired resistance in these tumor types. MethodsIn this study, we used 1 million array-CGH and qRT-PCR methodologies to identify and validate cytogenetic alterations that arise after cisplatin treatment in four lung and ovarian paired cisplatin-sensitive/resistant cell lines. We used whole transcriptome sequencing (RNA-seq), functional transfection assays and gene-pathway activity analysis in our experimental cellular models and in fresh frozen primary NSCLC tumors to identify genes with a potential role in the development of this malignancy. Results were further explored in 55 lung and ovarian primary tumors and control samples and in two extensive in silico databases (TCGA and KMplotter) with 1,926 NSCLC and 1,425 additional epithelial tumors. ResultsLong-term cell exposure to platinum induces the frequent deletion of ITF2 gene. Restoration of ITF2 expression re-sensitizes tumor cells to platinum and recovers the levels of Wnt/{beta}-catenin transcriptional activity. ITF2 expression was also frequently downregulated in NSCLC, ovarian and other epithelial tumors, predicting a worse overall survival. We also identified an inverse correlation in expression between ITF2 and HOXD9, revealing that NSCLC patients with lower expression of HOXD9 have a better overall survival rate that was independent of the tumor histology. ConclusionWe have defined the implication of ITF2 as a molecular mechanism behind the development of cisplatin resistance probably through the activation of the Wnt-signaling pathway. Our translational data suggest that ITF2 could be used as a general epithelial tumor platinum-predictive marker and have identified HOXD9 as a potential prognostic biomarker in NSCLC, a gene which expression is induced by Wnt signaling. Furthermore, this data highlights the possible role of ITF2 and HOXD9 as a novel therapeutic target for platinum resistant tumors.

cancer biology↗