Biology subjects

Deek, J.

Publications and source records attributed to Deek, J..

5 recordsLinked to original sources

bioRxiv2026

Beyond Bisulfite Sequencing: Resolving 5-hmC with Nanopore Sequencing Unmasks the True-5mC Methylation Entropy Landscape

DNA methylation dynamically regulates cellular function and phenotype. At the tissue level, stochasticity in methylation patterns, as measured by methylation entropy, drives plasticity, development, cancer, and aging. Modulation of methylation patterns is facilitated by erasure of 5-methylcytosine (5mC) via the oxidized intermediate 5-hydroxymethylcytosine (5hmC). Bisulfite sequencing cannot distinguish the two modifications, labeling them both as 5-mC. We quantitatively analyze the effect of this historical conflation on the genome-wide distribution of methylation levels and methylation entropy. Using nanopore sequencing with direct 5mC and 5hmC calling, we compare True-mC to bisulfite-like analysis in two model systems, kidney cancer and the mouse medial prefrontal cortex. We show that bisulfite sequencing introduces systematic, tissue-specific shifts in methylation distribution that affect the mechanistic interpretation of the underlying biology. The distortion scaled with endogenous 5hmC content: substantial in brain, where over half of the entropy-associated gene ontology terms recovered under bisulfite-like analysis were absent under True-mC, and minimal in the low-5hmC kidney cancer sample, which delimits the regime in which bisulfite-derived entropy remains interpretable. Together, these findings establish that True-5mC-based methylation entropy redefines the physical mapping of certain epigenomes, demonstrating that in some contexts, what has previously been interpreted as stochastic maintenance failure is frequently the structured signature of distinct and mechanistically interpretable cytosine biochemistry.

bioinformatics↗

bioRxiv2025

Write and read: Harnessing Synthetic DNA Modifications for Nanopore Sequencing.

An exciting feature of nanopore sequencing is its ability to record multi-omic information on the same sequenced DNA molecule. Well-trained models allow the detection of nucleotide-specific molecular signatures through changes in ionic current as DNA molecules translocate through the nanopore. Thus, naturally occurring DNA modifications, such as DNA methylation and hydroxymethylation, may be recorded simultaneously with the genetic sequence. Additional genomic information, such as chromatin state or the locations of bound transcription factors, may also be recorded if their locations are chemically encoded into the DNA. Here, we present a versatile "write-and-read" framework, where chemo-enzymatic DNA labeling with unnatural synthetic tags results in predictable electrical fingerprints in nanopore sequencing. As a proof-of-concept, we explore a DNA glucosylation approach that selectively modifies 5-hydroxymethylcytosine (5hmC) with glucose or glucose-azide adducts. We demonstrate that these modifications generate distinct and reproducible electrical shifts, enabling the direct detection of chemically altered nucleotides. We further demonstrate that enzymatic alkylation, such as the enzymatic transfer of azide residues to the N6 position of adenines, also produces characteristic nanopore signal shifts relative to the native adenine and 6-methyladenine. Beyond direct nucleotide detection, this approach introduces new possibilities for bio-orthogonal DNA labeling, enabling an extended alphabet of sequence-specific detectable moieties. The future use of programmable chemical modifications for simultaneous analysis of multiple omics features on individual molecules opens new avenues for genetic research and discovery.

bioengineering↗

bioRxiv2024

Long-Read Structural and Epigenetic Profiling of a Kidney Tumor-matched Sample with Nanopore Sequencing and Optical Genome Mapping

Carcinogenesis often involves significant alterations in the cancer genome architecture, marked by large structural and copy number variations (SVs and CNVs) that are difficult to capture with short-read sequencing. Traditionally, cytogenetic techniques are applied to detect such aberrations, but they are limited in resolution and do not cover features smaller than several hundred kilobases. Optical genome mapping and nanopore sequencing are attractive technologies that bridge this resolution gap and offer enhanced performance for cytogenetic applications. These methods profile native, individual DNA molecules, thus capturing epigenetic information. We applied both techniques to characterize a clear cell renal cell carcinoma (ccRCC) tumors structural and copy number landscape, highlighting the relative strengths of each method in the context of variant size and average read length. Additionally, we assessed their utility for methylome and hydroxymethylome profiling, emphasizing differences in epigenetic analysis applicability.

genomics↗

bioRxiv2022

Optical Genome and Epigenome Mapping of Clear Cell Renal Cell Carcinoma

Cancer cells display complex genomic aberrations that include large-scale genetic rearrangements and epigenetic modulation that are not easily characterized by short-read sequencing. We present a method for simultaneous profiling of long-range genetic/epigenetic changes in matched cancer samples. Clear cell renal cell carcinoma (ccRCC) is the most common subtype of kidney cancer. Most ccRCC cases demonstrate somatic genomic alterations involving the short arm of chromosome 3 (3p), most often targeting the von Hippel-Lindau (VHL) gene. Aiming to identify somatic alterations that characterize early stage ccRCC, we performed comprehensive genetic, cytogenetic and epigenetic analyses comparing ccRCC tumor to adjacent non-tumorous tissue. Optical genome mapping identified genomic aberrations such as structural and copy number variations, complementing exome-sequencing results. Single-molecule methylome and hydroxymethylome mapping revealed multiple differential regions, some of them known to be associated with ccRCC pathogenesis. Among them, metabolic pathways were significantly enriched. Moreover, significant global epigenetic differences were detected between the tumor and the adjacent non-tumorous tissue, and a correlation between epigenetic signals and gene expression was found. This is the first reported comparison of a human tumor and a matched tissue by optical genome/epigenome mapping, revealing well-established and novel somatic aberrations.

genomics↗

bioRxiv2021

Genome sequences of Aegilops species of section Sitopsis reveal phylogenetic relationships and provide resources for wheat improvement

Aegilops is a close relative of wheat (Triticum spp.), and Aegilops species in the section Sitopsis represent a rich reservoir of genetic diversity for improvement of wheat. To understand their diversity and advance their utilization, we produced whole-genome assemblies of Ae. longissima and Ae. speltoides. Whole-genome comparative analysis, along with the recently sequenced Ae. sharonensis genome, showed that the Ae. longissima and Ae. sharonensis genomes are highly similar and most closely related to the wheat D subgenome. By contrast, the Ae. speltoides genome is more closely related to the B subgenome. Haplotype block analysis supported the idea that Ae. speltoides is the closest ancestor of the wheat B subgenome and highlighted variable and similar genomic regions between the three Aegilops species and wheat. Genome-wide analysis of nucleotide-binding site leucine-rich repeat (NLR) genes revealed species-specific and lineage-specific NLR genes and variants, demonstrating the potential of Aegilops genomes for wheat improvement. TeaserGenome sequences of Aegilops species provides a key for efficient exploitation of this rich genetic resource in wheat improvement.

plant biology↗