Biology subjects

Wu, D. C.

Publications and source records attributed to Wu, D. C..

7 recordsLinked to original sources

bioRxiv2026

Structural insights into antibody responses against influenza A virus in its natural reservoir

While influenza A virus undergoes rapid antigenic drift in humans, at least some subtypes, such as H3, have relatively stable antigenicity in natural waterfowl reservoirs, despite the presence of immune pressure. However, the underlying mechanisms remain poorly understood. This study identified and characterized 187 antibodies to H3 hemagglutinin from experimentally infected mallard ducks, 18 of which were further analyzed by cryo-EM. Compared with human H3 antibodies, duck H3 antibodies exhibited higher glycan-binding propensity, more balanced immunodominance hierarchy, and targeted distinct epitopes. Other unique features of duck H3 antibodies included a convergent CDR H3-independent heavy chain-only binding mode and an N-glycosylated CDR H3 as decoy receptor. By annotating duck immunoglobulin germline genes, we also demonstrated the importance of gene conversion in duck H3 antibodies. Overall, our findings provide insights into how millennia of coevolution have shaped the interplay between influenza A virus antigenic drift and antibody responses in the natural reservoir.

immunology↗

bioRxiv2021

Arg-tRNA synthetase links inflammatory metabolism to RNA splicing via nuclear condensates

Cells respond to perturbations like inflammation by sensing changes in metabolite levels. Especially prominent is arginine, which has known connections to the inflammatory response. Here, we found that depletion of arginine during inflammation decreased levels of a nuclear form of arginyl-tRNA synthetase (ArgRS). Surprisingly, we found that nuclear ArgRS interacts with serine/arginine repetitive matrix protein 2 (SRRM2), a spliceosomal protein and nuclear speckle component and that arginine depletion impacted both condensate-like nuclear trafficking of SRRM2 and splice-site usage in certain genes. These splice-site usage changes cumulated in synthesis of different protein isoforms that altered cellular metabolism and peptide presentation to immune cells. Our findings uncover a novel mechanism whereby a tRNA synthetase cognate to a key amino acid that is metabolically controlled during inflammation modulates the splicing machinery.

molecular biology↗

bioRxiv2021

Human organoid systems reveal in vitro correlates of fitness for SARS-CoV-2 B.1.1.7

A new phase of the COVID-19 pandemic has started as several SARS-CoV-2 variants are rapidly emerging globally, raising concerns for increased transmissibility. As animal models and traditional in vitro systems may fail to model key aspects of the SARS-CoV-2 replication cycle, representative in vitro systems to assess variants phenotypically are urgently needed. We found that the British variant (clade B.1.1.7), compared to an ancestral SARS-CoV-2 clade B virus, produced higher levels of infectious virus late in infection and had a higher replicative fitness in human airway, alveolar and intestinal organoid models. Our findings unveil human organoids as powerful tools to phenotype viral variants and suggest extended shedding as a correlate of fitness for SARS-CoV-2. One-Sentence SummaryBritish SARS-CoV-2 variant (clade B.1.1.7) infects organoids for extended time and has a higher fitness in vitro.

microbiology↗

bioRxiv2021

Human airway cells prevent SARS-CoV-2 multibasic cleavage site cell culture adaptation

Virus propagation methods generally use transformed cell lines to grow viruses from clinical specimens, which may force viruses to rapidly adapt to cell culture conditions, a process facilitated by high viral mutation rates. Upon propagation in VeroE6 cells, SARS-CoV-2 may mutate or delete the multibasic cleavage site (MBCS) in the spike protein that facilitates serine protease-mediated entry into human airway cells. We report that propagating SARS-CoV-2 on the human airway cell line Calu-3 - that expresses serine proteases - prevents MBCS mutations. Similar results were obtained using a human airway organoid-based culture system for SARS-CoV-2 propagation. Thus, in-depth knowledge on the biology of a virus can be used to establish methods to prevent cell culture adaptation.

microbiology↗

bioRxiv2020

Human cells contain myriad excised linear intron RNAs with potential functions in gene regulation and utility as disease biomarkers

Previous Thermostable Group II Intron Reverse Transcriptase sequencing (TGIRT-seq) found that human plasma contains short ([≤]300 nt) structured full-length excised linear intron RNAs (FLEXIs) with potential utility as blood-based biomarkers. Here, TGIRT-seq identified >9,000 different FLEXIs in human cells, including relatively abundant FLEXIs with cell-type-specific expression patterns. Analysis of published CLIP-seq datasets identified 126 RNA-binding proteins (RBPs) that bind different FLEXIs, including 53 RBPs that bind [≥]30 FLEXIs. These RBPs included splicing factors, transcription factors, a chromatin remodeling protein, cellular growth regulators, and proteins with cytoplasmic functions. Analysis of published datasets identified subsets of these RBPs that bind at FLEXI splice sites and impact alternative splicing or FLEXI host gene mRNA levels. Hierarchical clustering identified 6 subsets of RBPs whose binding sites were co-enriched in different subsets of FLEXIs: AGO1-4 and DICER, including but not limited to annotated agotrons and mirtron pre-miRNAs; DKC1, NOLC1, SMNDC1, and AATF (Apoptosis Antagonizing Transcription Factor), including but not limited to FLEXIs encoding snoRNAs; two sets of alternative splicing factors; and two sets that included RBPs with cytoplasmic functions (e.g., LARP4, PABPC4, METAP2, and ZNF622) together with nuclear regulatory proteins. The subsets of host genes encoding FLEXIs that bind these RBPs were enriched with non-FLEXI other short and long introns that bind the same RBPs, and TGIRT-seq of nuclear and cytoplasmic fractions from 4 cell lines showed cytoplasmic enrichment of FLEXIs with binding sites for RBPs that function in the cytoplasm. Collectively, our findings suggest that different subsets of RBPs bind FLEXIs and other introns to coordinately regulate the expression of functionally related host genes and then co-localize with stably bound FLEXIs to different intracellular locations. The cell-type specific expression of relatively abundant FLEXIs suggests utility as cellular as well as blood-based RNA biomarkers.

genomics↗

bioRxiv2020

Identification of protein-protected mRNA fragments and structured excised intron RNAs in human plasma by TGIRT-seq peak calling

Human plasma contains >40,000 different coding and non-coding RNAs that are potential biomarkers for human diseases. Here, we used thermostable group II intron reverse transcriptase sequencing (TGIRT-seq) combined with peak calling to simultaneously profile all RNA biotypes in apheresis-prepared human plasma pooled from healthy individuals. Extending previous TGIRT-seq analysis, we found that human plasma contains largely fragmented mRNAs from >19,000 protein-coding genes, abundant full-length, mature tRNAs and other structured small non-coding RNAs, and less abundant tRNA fragments and mature and pre-miRNAs. Many of the mRNA fragments identified by peak calling correspond to annotated protein-binding sites and/or have stable predicted secondary structures that could afford protection from plasma nucleases. Peak calling also identified novel repeat RNAs, miRNA-sized RNAs, and putatively structured intron RNAs of potential biological, evolutionary, and biomarker significance, including a family of full-length excised introns RNAs, subsets of which correspond to mirtron pre-miRNAs or agotrons.

genomics↗

bioRxiv2018

Limitation of alignment-free tools in total RNA-seq quantification

BackgroundAlignment-free RNA quantification tools have significantly increased the speed of RNA-seq analysis. However, it is unclear whether these state-of-the-art RNA-seq analysis pipelines can quantify small RNAs as accurately as they do with long RNAs in the context of total RNA quantification.\n\nResultWe comprehensively tested and compared four RNA-seq pipelines on the accuracies of gene quantification and fold-change estimation on a novel total RNA benchmarking dataset, in which small non-coding RNAs are highly represented along with other long RNAs. The four RNA-seq pipelines were of two commonly-used alignment-free pipelines and two variants of alignment-based pipelines. We found that all pipelines showed high accuracies for quantifying the expressions of long and highly-abundant genes. However, alignment-free pipelines showed systematically poorer performances in quantifying lowly-abundant and small RNAs.\n\nConclusionWe have shown that alignment-free and traditional alignment-based quantification methods performed similarly for common gene targets, such as protein-coding genes. However, we identified a potential pitfall in analyzing and quantifying lowly-expressed genes and small RNAs with alignment-free pipelines, especially when these small RNAs contain mutations.

bioinformatics↗