Biology subjects

Chung, N.

Publications and source records attributed to Chung, N..

10 recordsLinked to original sources

bioRxiv2026

N-glycoproteomic profiling of the hookworm Ancylostoma ceylanicum reveals heterogeneity and phosphorylcholine-modified glycans in larval and adult parasites.

Glycans of parasitic worms (helminths) have been previously described with roles in immunomodulation and the recognition of helminth proteins by the host immune system. Despite their significance in other systems, little is known about the glycosylation patterns of the hookworm Ancylostoma ceylanicum, an intestinal parasite of animals and humans, due to challenges in sample collection and structural complexity of many nematode glycans. Herein, we describe results of a released N-glycan analysis of adult hookworms, which revealed oligomannose, hybrid, and complex structures, including those with non-mammalian features. Next, we report the results of glycoproteomic analyses to identify hookworm glycoproteins and their associated glycan structures. We compared three enrichment methods and found that strong anion-exchange electrostatic repulsion liquid chromatography (SAX-ERLIC) led to the identification of the most N-glycopeptides. Analyses of larval and adult extracts revealed 82 larval and 157 adult N-glycoproteins, the majority of which were stage-specific. Of particular interest, phosphorylcholine (PC), a known immunomodulatory glycan modification, was detected in both stages, and several unique PC-modified structures were identified in each. Collectively, these results provide the first site-specific characterization of intact N-glycopeptides in A. ceylanicum and, with an adult-stage released glycan analysis, establish a method for glycoproteomic characterizations of other helminths.

microbiology↗

bioRxiv2025

In-depth analysis of the tear fluid glycoproteome reveals diverse lacritin glycosylation and spliceoforms

Tear fluid comprises a diverse group of extracellular glycoproteins which are critical for ocular homeostasis. Within the tear fluid glycoproteome, lacritin is highly expressed and plays a key role in immune response, tear secretion, and antimicrobial activity. Importantly, glycosylation constitutes over 50% of lactritins molecular weight. However, despite this fact, nothing is known about the specific glycan structures on lacritin and how they influence its protein folding, function, or downstream biological processes. Similarly, it remains completely unknown whether alterations to lacritin glycans are correlated with ocular pathologies. To address this gap in knowledge, we harnessed mass spectrometry (MS) to conduct the first O-glycoproteomic study of tear fluid. Here, we report unprecedented coverage of lacritin glycosylation, detailing 19 O-glycosites bearing a myriad of glycan structures. Further, we leveraged Alphafold 3.0 and GlycoShape to visualize the impact of these glycans on its structure, demonstrating that O-glycosylation renders the protein backbone rigid and extended. Surprisingly, we also detected protein-level evidence of two lacritin spliceoforms, representing the first observation of these isoforms by MS. Simultaneously, we describe the most comprehensive characterization of the tear fluid glycoproteome to date, elucidating the glycosylation profile of Immunoglobulin A (IgA), lactoferrin, and other glycoproteins with demonstrated clinical relevance as diagnostic biomarkers. Overall, this study lays critical groundwork for future biochemical investigation of tear fluid glycoproteins and their application as diagnostic or therapeutic tools for ocular diseases.

biochemistry↗

bioRxiv2025

A comparison shopper's guide to forest datasets

Recent advances in remote sensing, data availability, and cloud-based computing have led to a rapid expansion of publicly accessible datasets characterizing forest cover and land use. These datasets are widely used in ecological research, natural resource management, and policymaking. However, the sheer number of available products--and the often-subtle differences among them--pose significant challenges for users seeking the most appropriate dataset for their specific objectives. Here, we evaluate 27 derived products from 12 publicly available sources that quantify tree or forest cover and use across the conterminous United States (CONUS), with temporal coverage ranging from 5 to 30 years. The products include both satellite-based remote sensing data and ground-based national forest inventory data. We ask: How, why, and where do these datasets differ in their estimates of forest extent and change over time? Our analysis reveals that estimates of total forest area at the CONUS scale differ by over 2,000,000 km{superscript 2}, and correlations among forest change estimates vary widely in both direction and statistical significance. To support dataset selection and interpretation, we developed an open-access map comparison tool using Google Earth Engine. Our findings highlight the substantial implications of dataset choice for understanding forest dynamics and underscore the need for careful selection and transparent reporting in forest-related analyses.

ecology↗

bioRxiv2023

Xylosyltransferase Bump-and-hole Engineering to Chemically Manipulate Proteoglycans in Mammalian Cells

Mammalian cells orchestrate signalling through interaction events on their surfaces. Proteoglycans are an intricate part of these interactions, carrying large glycosaminoglycan polysaccharides that recruit signalling molecules. Despite their importance in development, cancer and neurobiology, a relatively small number of proteoglycans have been identified. In addition to the complexity of glycan extension, biosynthetic redundancy in the first protein glycosylation step by two xylosyltransferase isoenzymes XT1 and XT2 complicates annotation of proteoglycans. Here, we develop a chemical genetic strategy that manipulates the glycan attachment site of cellular proteoglycans. By employing a tactic termed bump- and-hole engineering, we engineer the two isoenzymes XT1 and XT2 to specifically transfer a chemically modified xylose analogue to target proteins. The chemical modification contains a bioorthogonal tag, allowing the ability to visualise and profile target proteins modified by both transferases in mammalian cells. The versatility of our approach allows pinpointing glycosylation sites by tandem mass spectrometry, and exploiting the chemical handle to manufacture proteoglycans with defined GAG chains for cellular applications. Engineered XT enzymes permit a view into proteoglycan biology that is orthogonal to conventional techniques in biochemistry.

biochemistry↗

bioRxiv2023

A genome-wide CRISPR screen supported by human genetics identifies the TNRC18 gene locus as a novel regulator of inflammatory signaling

Interleukin-1{beta} (IL-1{beta}) is dysregulated in many chronic inflammatory diseases, yet the genetic factors influencing IL-1{beta} production and signaling remain largely unknown. Myeloid-derived cells are the primary producers of IL-1{beta}, prompting a genome-wide CRISPR knockout screen in the human myeloid-derived U937 cell model, treated with lipopolysaccharide (LPS) to mimic inflammatory conditions, and sorted for high and low intracellular IL-1{beta} levels. A total of 295 genes were identified as regulators of IL-1{beta} production, including known mediators, such as TLR4, JAK-STAT, IL-10 receptor, and the Cullin ring finger ligase complex. Notably, 57 out of the 295 genes overlapped with loci associated with human inflammatory diseases, including the TNRC18 gene on chromosome 7p22.1 associated with multiple diseases in the Finnish population. U937 cells engineered with the homozygous rs748670681 risk allele associated with inflammatory bowel disease, demonstrated decreased levels of mRNA for TNRC18 and an adjacent gene WIPI2, reduction in LPS-dependent gene activation and cytokine production, but elevation of interferon-responsive gene programs. Transcriptomic profiles for individual knockouts of TNRC18 and WIPI2 attributed the loss of LPS-dependent signaling primarily to TNRC18 while the exacerbation of interferon signaling is a hallmark of loss of WIPI2. Collectively, these findings delineate the global regulatory mechanisms of IL-1{beta} production and provide molecular insights to the role of the rs748670681 variant as a pleiotropic risk factor for inflammatory diseases.

genomics↗

bioRxiv2020

Accurate Microbiome Sequencing with Synthetic Long Read Sequencing

The microbiome plays a central role in biochemical cycling and nutrient turnover of most ecosystems. Because it can comprise myriad microbial prokaryotes, eukaryotes and viruses, microbiome characterization requires high-throughput sequencing to attain an accurate identification and quantification of such co-existing microbial populations. Short-read next-generation-sequencing (srNGS) revolutionized the study of microbiomes and remains the most widely used approach, yet read lengths spanning only a few of the nine hypervariable regions of the 16S rRNA gene limit phylogenetic resolution leading to misclassification or failure to classify in a high percentage of cases. Here we evaluate a synthetic long-read (SLR) NGS approach for full-length 16S rRNA gene sequencing that is high-throughput, highly accurate and low-cost. The sequencing approach is amenable to highly multiplexed sequencing and provides microbiome sequence data that surpasses existing short and long-read modalities in terms of accuracy and phylogenetic resolution. We validated this commercially-available technology, termed LoopSeq, by characterizing the microbial composition of well-established mock microbiome communities and diverse real-world samples. SLR sequencing revealed differences in aquatic community complexity associated with environmental gradients, resolved species-level community composition of uterine lavage from subjects with histories of misconception and accurately detected strain differences, multiple copies of the 16S rRNA in a single strains genome, as well as low-level contamination in soil cyanobacterial cultures. This approach has implications for widespread adoption of high-resolution, accurate long-read microbiome sequencing as it is generated on popular short read sequencing platforms without the need for additional infrastructure.

microbiology↗

bioRxiv2020

Adaptation of codon and amino acid use for translational functions in highly expressed cricket genes

BackgroundFor multicellular organisms, much remains unknown about the dynamics of synonymous codon and amino acid use in highly expressed genes, including whether their use varies with expression in different tissue types and sexes. Moreover, specific codons and amino acids may have translational functions in highly transcribed genes, that largely depend on their relationships to tRNA gene copies in the genome. However, these relationships and putative functions are poorly understood, particularly in multicellular systems. ResultsHere, we rigorously studied codon and amino acid use in highly expressed genes from reproductive and nervous system tissues (male and female gonad, somatic reproductive system, brain, ventral nerve cord, and male accessory glands) in the cricket Gryllus bimaculatus. We report an optimal codon, defined as the codon preferentially used in highly expressed genes, for each of the 18 amino acids with synonymous codons in this organism. The optimal codons were largely shaped by selection, and their identities were mostly shared among tissue types and both sexes. However, the frequency of optimal codons was highest in gonadal genes. Concordant with translational selection, a majority of the optimal codons had abundant matching tRNA gene copies in the genome, but sometimes obligately required wobble tRNAs. We suggest the latter may comprise a mechanism for slowing translation of abundant transcripts, particularly for cell-cycle genes. Non-optimal codons, defined as those least commonly used in highly transcribed genes, intriguingly often had abundant tRNAs, and had elevated use in a subset of genes with specialized functions (gametic and apoptosis genes), suggesting their use promotes the upregulation of particular mRNAs. In terms of amino acids, we found evidence suggesting that amino acid frequency, tRNA gene copy number, and amino acid biosynthetic costs (size/complexity) had all interdependently evolved in this insect model, potentially for translational optimization. ConclusionsCollectively, the results strongly suggest that codon use in highly expressed genes, including optimal, wobble, and non-optimal codons, and their tRNAs abundances, as well as amino acid use, have been adapted for various functional roles in translation within this cricket. The effects of expression in different tissue types and the two sexes are discussed.

genomics↗

bioRxiv2019

Validation of a CRISPR/cas9-based technology platform for examining specific immune gene functions in an experimental murine model of IBD

Inflammatory bowel diseases (IBD) are complex, multifactorial disorders characterized by chronic relapsing intestinal inflammation. Association studies have identified hundreds of genes that are linked to IBD and potentially regulate its pathology. The further dissection of the genetic network underlining IBD pathogenesis and pathophysiology is hindered by the limited capacity to investigate the role of each GWAS association through functional studies, including the generation of knockout animal models for each of the associated genes. The CRISPR/Cas9 system represents a cutting edge technology which has the potential to transform the field of IBD research by facilitating the introduction of genetic alterations in an efficient and effective manner. Using the CD40-mediated-colitis model, our results demonstrate the validity of a CRISPR/Cas9-based platform as a tool for the validation of target genes or interference strategies in experimental IBD. The utilization of this discovery strategy will allow for the timely in vivo validation of therapeutic targets as the rapidly emerge from current genetic and genomics efforts with human disease tissue. As such, the CRISPR/Cas9-based platform can significantly shorten the time span between target identification and generation of proof of principle experiments for drug discovery.

immunology↗

bioRxiv2019

VHL Synthetic Lethality Signatures Uncovered by Genotype-specific CRISPR-Cas9 Screens

BackgroundGenome-wide CRISPR-Cas9 essentiality screening represents a powerful approach to identify genetic vulnerabilities in cancer cells. Here, we applied this technology and designed a strategy to identify target genes that are synthetic lethal (SL) with von Hippel-Lindau (VHL) tumor suppressor gene. Inactivation of VHL has been frequently found in clear cell renal cell carcinoma (ccRCC). Its SL partners serve as potential drug targets for the development of targeted cancer therapies.\n\nResultsWe performed parallel genome-wide CRISPR screens in two pairs of isogenic ccRCC cell lines that differ only in the VHL status. Comparative analyses of screening results not only confirmed a well-known role for mTOR signaling in renal carcinoma, but also identified DNA damage response and selenocysteine biosynthesis pathways as major SL targets in VHL-inactivated cancer cells. Follow-up studies provided cellular and mechanistic insights into SL interactions of these pathway genes with the VHL gene.\n\nConclusionsUsing isogenic CRISPR screening approach, we uncovered novel biological processes that are SL with VHL, which can be exploited for drug development for ccRCC. Our CRISPR and RNA-seq datasets provide a rich resource for future investigation of the function of the VHL tumor suppressor protein. Our work demonstrates the efficiency of CRISPR-based synthetic lethality screening in human isogenic cell pairs. Similar strategies could be employed to unveil SL partners with other oncogenic drivers.

cancer biology↗

bioRxiv2018

Transcriptome analysis reveals negative correlation between transcripts of mitochondrial genes and L1HS, and positive correlation between KRAB-ZFPs and older LINE elements in normal somatic tissue

BackgroundDespite the long-held assumption that transposons are normally only expressed in the germ-line, recent evidence shows that transcripts of transposable element (TE) sequences are frequently found in the somatic cells. However, the extent of variation in TE transcript levels across different tissues and different individuals are unknown, and the co-expression between TEs and host gene mRNAs have not been examined.\n\nResultsHere we report the variation in TE derived transcript levels across tissues and between individuals observed in the non-tumorous tissues collected for The Cancer Genome Atlas. We found core TE co-expression modules consisting mainly of transposons, showing correlated expression across broad classes of TEs. Despite this co-expression within tissues, there are individual TE loci that exhibit tissue-specific expression patterns, when compared across tissues. The core TE modules were negatively correlated with other gene modules that consisted of immune response genes in interferon signaling. KRAB Zinc Finger Proteins (KZFPs) were over-represented gene members of the TE modules, showing positive correlation across multiple tissues. But we did not find overlap between TE-KZFP pairs that are co-expressed and TE-KZFP pairs that are bound in published ChIP-seq studies.\n\nConclusionsWe find unexpected variation in TE derived transcripts, within and across non-tumorous tissues. We describe a broad view of the RNA state for non-tumorous tissues exhibiting higher level of TE transcripts. Tissues with higher level of TE transcripts have a broad range of TEs co-expressed, with high expression of a large number of KZFPs, and lower RNA levels of immune genes.

genomics↗