Biology subjects

Wang, T.-Y.

Publications and source records attributed to Wang, T.-Y..

At least 37 records · Page 2Linked to original sources

bioRxiv2024

Aromatase-independent estrogenesis: Wood-Ljungdahl pathway likely contributed to the emergence of estrogens in the biosphere

Androgen and estrogen, key sex hormones, were long thought to be exclusively produced by vertebrates. The O2-dependent aromatase that converts androgen to estrogen (estrogenesis) has never been identified in any prokaryotes. Here, we report the discovery of anaerobic estrogenesis in a Peptococcaceae bacterium (strain TUW77) isolated from the gut of the great blue-spotted mudskipper (Boleophthalmus pectinirostris). This strain exhibits unprecedented testosterone fermentation pathways, transforming testosterone into estrogens and androstanediol under anaerobic conditions. Physiological experiments revealed that strain TUW77 grows exclusively on testosterone, utilizing the androgenic C-19 methyl group as both the carbon source and electron donor. The genomic analysis identified three copies of a polycistronic gene cluster, abeABC (anaerobic bacterial estrogenesis), encoding components of a classic cobalamin-dependent methyltransferase system. These genes, highly expressed under testosterone-fed conditions, show up to 57% protein identity to the characterized EmtAB from denitrifying Denitratisoma spp., known for methylating estrogen into androgen (the reverse reaction). Tiered transcriptomic and proteomic analyses suggest that the removed C-19 methyl group is completely oxidized to CO2 via the oxidative Wood-Ljungdahl pathway, while the reducing equivalents (NADH) fully reduce remaining testosterone to androstanediol. Consistently, the addition of anthraquinone-2,6-disulfonate, an extracellular electron acceptor, to testosterone-fed TUW77 cultures enabled complete testosterone conversion into estrogen without androstanediol accumulation (anaerobic testosterone oxidation). This discovery of aromatase-independent estrogenesis in anaerobic bacteria suggests that the ancient Wood-Ljungdahl pathway may have contributed to the emergence of estrogens in the early biosphere. SignificanceUsing a testosterone-grown anaerobic bacterium as a model organism, we characterized this unusual anaerobic estrogenesis at the molecular level. Our findings challenge the long-held belief that estrogen production is exclusive to aromatase-containing vertebrates, expanding our understanding of steroid hormone biosynthesis across domains of life. The involvement of ancient strictly anaerobic Peptococcaceae members and the Wood-Ljungdahl pathway suggests that bacterial estrogenesis may predate O2-dependent estrogenesis in vertebrates. Furthermore, the identification of estrogen-producing bacteria in animal guts opens new avenues for potential microbiome-based hypoestrogenism therapies to supplement estrogen in menopausal or ovariectomized females, offering an innovative alternative to current hormone replacement strategies.

microbiology↗

bioRxiv2024

Chromosome-level genome assembly of the loach goby, Rhyacichthys aspro, reveals mechanisms underlying Gobioidei diversification

The percomorph fish clade Gobioidei are a suborder that comprises over 2,200 species distributed in nearly all aquatic habitats. To understand the genetics underlying their diversification, we sequenced and annotated the genome of the loach goby, Rhyacichthys aspro, the basal most group, and compared it with nine additional Gobioidei species. Within Gobioidei, the loach goby possesses the smallest genome at 607 Mb, and a rise in species diversity from basal to derived lineages is mirrored by enlarged genomes and a higher presence of repeat elements (REs), particularly DNA transposons. These transposons are enriched in coding and regulatory regions and their copy number increase is strongly correlated with mutation rate, suggesting that DNA repair after transposon excision/insertion leads to nearby mutations. Consequently, the proliferation of DNA transposons might be the crucial driver of Gobioidei diversification and adaptability. The loach goby genome also points to mechanisms of ecological adaptation. It contains relatively few genes for lateral line development but an over representation of synaptic function genes, with genes putatively under selection linked to synapse organization and calcium signaling, suggesting a sensory system distinct from other Gobioidei species. We also see an overabundance of genes involved in neurocranium development and renal function, adaptations likely connected to its flat morphology suited for strong currents and an amphidromous life cycle. Comparative analyses with hill-stream loaches and the European eel reveal convergent adaptations in body shape and saltwater balance. These findings shed light on the loach gobys survival mechanisms and the broader evolutionary trends within Gobioidei.

genomics↗

bioRxiv2024

The Fanconi anemia pathway repairs colibactin-induced DNA interstrand cross-links

Colibactin is a secondary metabolite produced by bacteria present in the human gut and is implicated in the progression of colorectal cancer and inflammatory bowel disease. This genotoxin alkylates deoxyadenosines on opposite strands of host cell DNA to produce DNA interstrand cross-links (ICLs) that block DNA replication. While cells have evolved multiple mechanisms to resolve ("unhook") ICLs encountered by the replication machinery, little is known about which of these pathways promote resistance to colibactin-induced ICLs. Here, we use Xenopus egg extracts to investigate replication-coupled repair of plasmids engineered to contain site-specific colibactin-ICLs. We show that replication fork stalling at a colibactin-ICL leads to replisome disassembly and activation of the Fanconi anemia ICL repair pathway, which unhooks the colibactin-ICL through nucleolytic incisions. These incisions generate a DNA double-strand break intermediate in one sister chromatid, which can be repaired by homologous recombination, and a monoadduct ("ICL remnant") in the other. Our data indicate that translesion synthesis past the colibactin-ICL remnant depends on Pol{eta} and a Pol{kappa}-REV1-Pol{zeta} polymerase complex. Although translesion synthesis past colibactin-induced DNA damage is frequently error-free, it can introduce T>N point mutations that partially recapitulate the mutation signature associated with colibactin exposure in vivo. Taken together, our work provides a biochemical framework for understanding how cells tolerate a naturally-occurring and clinically-relevant ICL.

molecular biology↗

bioRxiv2024

Human cell surface-AAV interactomes identify LRP6 as blood-brain-barrier transcytosis receptor and immune cytokine IL3 as AAV9 binder

Adeno-associated viruses (AAVs) are foundational gene delivery tools for basic science and clinical therapeutics. However, lack of mechanistic insight, especially for engineered vectors created by directed evolution, can hamper their application. Here, we adapted an unbiased human cell microarray platform to determine the extracellular and cell surface interactomes of natural and engineered AAVs. We identified a naturally-evolved and serotype-specific interaction between the AAV9 capsid and human interleukin 3 (IL3), with possible roles in host immune modulation, as well as lab-evolved low-density-lipoprotein-receptor-related-protein 6 (LRP6) interactions specific to engineered capsids that cross the blood-brain barrier in non-human primates after intravenous administration. The unbiased cell microarray screening approach also allowed us to identify off-target tissue binding interactions of engineered brain-enriched AAV capsids that may inform vectors peripheral organ tropism and side effects. These results allow confident application of engineered AAVs in diverse organisms and unlock future target-informed engineering of improved viral and non-viral vectors for non-invasive therapeutic delivery to the brain.

bioengineering↗

bioRxiv2023

Single tissue proteomics in Caenorhabditis elegans reveals proteins resident in intestinal lysosome-related organelles

The nematode intestine is the primary site for nutrient uptake and storage as well as the synthesis of biomolecules; lysosome-related organelles known as gut granules are important for many of these functions. Aspects of intestine biology are not well understood, including the export of the nutrients it imports and the molecules it synthesizes, as well as the complete functions and protein content of the gut granules. Here, we report a mass spectrometry-based proteomic analysis of the intestine of the Caenorhabditis elegans and of its gut granules. Overall, we identified approximately 5,000 proteins each in the intestine and the gonad and showed that most of these proteins can be detected in samples extracted from a single worm, suggesting the feasibility of individual-level genetic analysis using proteomes. Comparing proteomes and published transcriptomes of the intestine and the gonad, we identified proteins that appear to be synthesized in the intestine and then transferred to the gonad. To identify gut granule proteins, we compared the proteome of individual intestines deficient in gut granules to the wild-type. The identified gut granule proteome includes proteins known to be exclusively localized to the granules and additional putative novel gut granule proteins. We selected two of these putative gut granule proteins for validation via immunohistochemistry, and our successful confirmation of both suggests that our strategy was effective in identifying the gut granule proteome. Our results demonstrate the practicability of single tissue mass-spectrometry- based proteomic analysis in small organisms and in its utility for making discoveries. Significance statementWe show that tissue-specific proteomic analysis is achievable and can be done efficiently at an individual level in a small nematode, with resolution sufficient for genetic analysis on a single animal basis. With data collected from single animals, we produced high-quality sets of proteins that described the proteomes of the gonad and the intestine. Comparison of these proteomes with the organs transcriptomes improved our understanding of interorgan protein transport. We applied single-tissue proteomic to describe the proteome of the gut granules in the nematode intestine, a specialized lysosome-related organelle with important functions but which is not well characterized, identifying proteins not previously known to be associated with LROs and verifying two by subcellular localization.

cell biology↗

bioRxiv2023

Role of a holo-insertase complex in the biogenesis of biophysically diverse ER membrane proteins

Mammalian membrane proteins perform essential physiologic functions that rely on their accurate insertion and folding at the endoplasmic reticulum (ER). Using forward and arrayed genetic screens, we systematically studied the biogenesis of a panel of membrane proteins, including several G-protein coupled receptors (GPCRs). We observed a central role for the insertase, the ER membrane protein complex (EMC), and developed a dual-guide approach to identify genetic modifiers of the EMC. We found that the back of sec61 (BOS) complex, a component of the multipass translocon, was a physical and genetic interactor of the EMC. Functional and structural analysis of the EMC*BOS holocomplex showed that characteristics of a GPCRs soluble domain determine its biogenesis pathway. In contrast to prevailing models, no single insertase handles all substrates. We instead propose a unifying model for coordination between the EMC, multipass translocon, and Sec61 for biogenesis of diverse membrane proteins in human cells.

cell biology↗

bioRxiv2023

Lesion-specific suppression of YAP/TAZ by biomimetic nanodrug ameliorates atherosclerosis development

Atherosclerosis, characterized by the buildup of lipid-rich plaque on the vessel wall, is the primary cause of myocardial infarction and ischemic stroke. Recent studies have demonstrated that dysregulation of yes-associated protein 1 (YAP) and transcriptional coactivator with PDZ-binding domain (TAZ) contributes to plaque development, making YAP/TAZ potential therapeutic targets. However, systemic modulation of YAP/TAZ expression or activities risks serious off-target effects, limiting clinical applicability. To address the challenge, this study develops monocyte membrane-coated nanoparticles (MoNP) as a drug delivery vehicle targeting activated endothelium lining the plaque surface and utilizes MoNP to deliver verteporfin (VP), a potent YAP/TAZ inhibitor, for lesion-specific treatment of atherosclerosis. The results reveal that MoNP significantly enhance payload delivery to inflamed endothelial cells (EC) while avoiding phagocytic cells, and preferentially accumulate in atherosclerotic regions. MoNP-mediated delivery of VP substantially reduces YAP/TAZ expression, suppressing inflammatory gene expression and macrophage infiltration in cultured EC and mouse arteries exposed to atherogenic stimuli. Importantly, this lesion-targeted VP nanodrug effectively decreases plaque development in mice without causing noticeable histopathological changes in major organs. Collectively, these findings demonstrate a plaque-targeted and pathway-specific biomimetic nanodrug, potentially leading to safer and more effective treatments for atherosclerosis.

bioengineering↗

bioRxiv2023

Self-Healable Spider Dragline Silk Materials

Developing materials with structural flexibility that permits self-repair in response to external disturbances remains challenging. Spider silk, which combines an exceptional blend of strength and pliability in nature, serves as an ideal dynamic model for adaptive performance design. In this work, a novel self-healing material is generated using spider silk. Dragline silk from spider Nephila pilipes is demonstrated with extraordinary in situ self-repair property through a constructed thin film format, surpassing that of two other silks from spider Cyrtophora moluccensis and silkworm Bombyx mori. Subsequently, R2, a key spidroin associated with self-healing, is biosynthesized, with validated cohesiveness. R2 is further programmed with tunable healability (permanent and reversible) and conductivity (graphene doping; R2G) for electronics applications. In the first demonstration, film strips from R2 and R2G are woven manually into multidimensional (1D-3D) conductive fabrics for creating repairable logic gate circuits. In the second example, a reversibly-healable R2/R2G strip is fabricated as a re-configurable wearable ring probe to fit fingertips of varying widths while retaining its detecting capabilities. Such prototype displays a unique conformable wearable technology. Last, the remarkable finding of self-healing in spider silk could offer a new material paradigm for developing future adaptive biomaterials with tailored performance and environmental sustainability.

bioengineering↗

bioRxiv2022

Region- and variance-based DNA methylation analyses reveal novel disease genes and pathways for systemic lupus erythematosus

BackgroundSystemic lupus erythematosus (SLE) is a prototype autoimmune disease with unclear pathogenesis. DNA methylation is an important regulatory mechanism on gene expression, providing a key angle to understand disease mechanisms. To understand the pathways involved in SLE, and to develop biomarkers for its diagnosis and treatment, we analyzed DNA methylation profiles on blood cells from SLE patients and healthy controls. ResultsWe identified most differentially methylated regions (DMRs) in T cells, while majority of differentially variable sites (DVSs) were found in B cells, featuring hypervariability in enhancers. We observed a prominent T cell receptor (TCR) signaling cluster with consistent hypermethylation and a B cell receptor (BCR) cluster with highly increased variability in SLE. Genes involved in innate immunity were often found hypomethylated, while adaptive immunity genes were featured with hypermethylation. Using a machine learning approach, we identified 60 genes that accurately distinguished SLE patients from healthy individuals, which also showed correlation with disease activities. ConclusionsThis study highlights the role of lymphocyte receptor aberrations in the disease and identified a list of genes showing great potential as biomarkers and shedding new light on disease mechanisms, through novel analyses of methylation data.

bioinformatics↗

bioRxiv2022

Spatial and temporal origin of the third SARS-CoV-2 Outbreak in Taiwan

Since the first report of SARS-CoV-2 in December 2019, Taiwan has gone through three local outbreaks. Unlike the first two outbreaks, the spatial and temporal origin of the third outbreak (April 20 to November 5, 2021) is still unclear. We assembled and analyzed a data set of more than 6,000 SARS-CoV-2 genomes, including 300 from Taiwan and 5812 related sequences downloaded from GISAID as of 2021/12/08. We found that the third outbreak in Taiwan was caused by a single virus lineage belonging to Alpha (B.1.1.7) strain. This lineage, T-III (the third outbreak in Taiwan), carries a distinct genetic fingerprint, consisting of spike M1237I (S-M1237I) and three silent mutations, C5812T, C15895T, and T27869C. The T-III is closest to the sequences derived from Turkey on February 8, 2021. The estimated age of the most recent common ancestor (TMRCA) of T-III is March 23, 2021 (95% highest posterior density [HPD] February 24 - April 13, 2021), almost one month before the first three confirmed cases on April 20, 2021. The effective population size of the T-III showed approximately 20-fold increase after the onset of the outbreak and reached a plateau in early June 2021. Our results reconcile several unresolved observations, including the occurrence of two infection clusters at the same time without traceable connection and several airline pilots who were PCR negative but serum IgM-/IgG+ for SARS-CoV-2 in late April. Therefore, in contrast to the general notion that the third SARS-CoV-2 outbreak in Taiwan was sparked by two imported cases from USA on April 20, 2021, which, in turn, was caused by the partial relaxation of entry quarantine measures in early April 2021, our comprehensive analyses demonstrated that the outbreak was most likely originated from Europe in February 2021.

evolutionary biology↗

bioRxiv2022

Structure of Anabaena flos-aquae gas vesicles revealed by cryo-ET

Gas vesicles (GVs) are gas-filled protein nanostructures employed by several species of bacteria and archaea as flotation devices to enable access to optimal light and nutrients. The unique physical properties of GVs have led to their use as genetically-encodable contrast agents for ultrasound and MRI. Currently, however, the structure and assembly mechanism of GVs remain unknown. Here we employ cryo-electron tomography to reveal how the GV shell is formed by a helical filament of highly conserved GvpA subunits. This filament changes polarity at the center of the GV cylinder--a site that may act as an elongation center. High-resolution subtomogram averaging reveals a corrugated pattern of the shell arising from polymerization of GvpA into a {beta}-sheet. The accessory protein GvpC forms a helical cage around the GvpA shell, providing structural reinforcement. Together, our results help explain the remarkable mechanical properties of GVs and their ability to adopt different diameters and shapes.

molecular biology↗

bioRxiv2022

Utilisation of Oxford Nanopore sequencing to generate six complete gastropod mitochondrial genomes as part of a biodiversity curriculum

High-throughput sequencing has enabled genome skimming approaches to produce complete mitochondrial genomes (mitogenomes) for species identification and phylogenomics purposes. In particular, the portable sequencing device from Oxford Nanopore Technologies (ONT) has the potential to facilitate hands-on training from sampling to sequencing and interpretation of mitogenomes. In this study, we present the results from sampling and sequencing six gastropod mitogenomes (Aplysia argus, Cellana orientalis, Cellana toreuma, Conus ebraeus, Conus miles and Tylothais aculeata) from a graduate level biodiversity course. The students were able to produce mitochondrial genomes from sampling to annotation using existing protocols and programs. Approximately 4Gb of sequence was produced from 15 Flongle and two R9.4 Flowcells, averaging 235Mb and N50=4.4kb per Flowcell. Five of the six 14.1-18kb mitogenomes were circlised containing all 13 core protein coding genes. Additional Illumina sequencing reveal that the ONT assemblies were able to span over highly AT rich sequence in the control region that was otherwise missing in Illumina-assembled mitogenomes, but still contained a base error of one every 70.8-346.7bp with the majority occurring at homopolymer regions. Our findings suggest that ONT are portable and can be used to rapidly produce mitogenomes at low cost and tailored to genomics-based training in biodiversity research.

ecology↗

bioRxiv2021

Handcuffing intrinsically disordered regions in Mlh1-Pms1 disrupts mismatch repair

The DNA mismatch repair (MMR) factor Mlh1-Pms1 contains long intrinsically disordered regions (IDRs). While essential for MMR, their exact functions remain elusive. We performed cross-linking mass spectrometry to identify the major interactions within the Mlh1-Pms1 heterodimer and used this information to insert FRB and FKBP dimerization domains into the IDRs of Mlh1 and Pms1. Yeast bearing these constructs were grown with rapamycin to induce dimerization. Strains containing FRB and FKBP domains in the Mlh1 IDR displayed complete MMR defects when grown with rapamycin, but removing rapamycin restored MMR functions. Furthermore, linking the Mlh1 and Pms1 IDRs through FRB-FKBP dimerization disrupted Mlh1-Pms1 binding to DNA, inappropriately activated Mlh1-Pms1, and caused MMR defects in vivo. We conclude that dynamic and coordinated rearrangements of the MLH IDRs regulate how the complex clamps DNA to catalyze MMR. The application of the FRB-FKBP dimerization system to interrogate in vivo functions of a critical repair complex will be useful for probing IDRs in diverse enzymes and to probe transient loss of MMR on demand.

genetics↗

bioRxiv2021

Targeting CTP Synthetase 1 to Restore Interferon Induction and Impede Nucleotide Synthesis in SARS-CoV-2 Infection

The newly emerged SARS-CoV-2 caused a global pandemic with astonishing mortality and morbidity. The mechanisms underpinning its highly infectious nature remain poorly understood. We report here that SARS-CoV-2 exploits cellular CTP synthetase 1 (CTPS1) to promote CTP synthesis and suppress interferon (IFN) induction. Screening a SARS-CoV-2 expression library identified ORF7b and ORF8 that suppressed IFN induction via inducing the deamidation of interferon regulatory factor 3 (IRF3). Deamidated IRF3 fails to bind the promoters of classic IRF3-responsible genes, thus muting IFN induction. Conversely, a shRNA-mediated screen focused on cellular glutamine amidotransferases corroborated that CTPS1 deamidates IRF3 to inhibit IFN induction. Functionally, ORF7b and ORF8 activate CTPS1 to promote de novo CTP synthesis while shutting down IFN induction. De novo synthesis of small-molecule inhibitors of CTPS1 enabled CTP depletion and IFN induction in SARS-CoV-2 infection, thus impeding SARS-CoV-2 replication. Our work uncovers a strategy that a viral pathogen couples immune evasion to metabolic activation to fuel viral replication. Inhibition of the cellular CTPS1 offers an attractive means for developing antiviral therapy that would be resistant to SARS-CoV-2 mutation.

microbiology↗

bioRxiv2020

Identification of 38 novel loci for systemic lupus erythematosus and genetic heterogeneity that may underly population disparities in this disease

Systemic lupus erythematosus (SLE), a worldwide autoimmune disease with high heritability, shows differences in prevalence, severity and age of onset among different ancestral groups. Previous genetic studies have focused more on European populations, which appear to be the least affected. Consequently, the genetic variations that underly the commonalities, differences and treatment options in SLE among ancestral groups have not been well elucidated. To address this, we undertook a genome-wide association study, increasing the sample size of Chinese populations to the level of existing European studies. Thirty-eight novel SLE-associated loci and incomplete sharing of genetic architecture were identified. Nine disease loci showed clear ancestral group heterogeneity and implicated antibody production as a potential mechanism for differences in disease manifestation. Polygenic risk scores performed significantly better when trained on matched ancestral data sets. These analyses help to reveal the genetic bases for disparities in SLE among ancestral groups.

genetics↗

bioRxiv2019

Control of adaptive action selection by secondary motor cortex during flexible visual categorization

Adaptive action selection during stimulus categorization is an important feature of flexible behavior. To examine neural mechanism underlying this process, we trained mice to categorize the spatial frequencies of visual stimuli according to a boundary that changed between blocks of trials in a session. Using a model with a dynamic decision criterion, we found that sensory history was important for adaptive action selection after the switch of boundary. Bilateral inactivation of the secondary motor cortex (M2) impaired adaptive action selection by reducing the behavioral influence of sensory history. Electrophysiological recordings showed that M2 neurons carried more information about upcoming choice and previous sensory stimuli when sensorimotor association was being remapped than when it was stable. Thus, M2 causally contributes to flexible action selection during stimulus categorization, with the representations of upcoming choice and sensory history regulated by the demand to remap stimulus-action association.

neuroscience↗

bioRxiv2019

Parallel Concerted Evolution of Ribosomal Protein Genes in Fungi and Its Adaptive Significance

Ribosomal proteins (RPs) genes encode structure components of ribosomes, the cellular machinery for protein synthesis. A single functional copy has been maintained in most of 78-80 RP families in animals due to evolutionary constraints imposed by gene dosage balance. Some fungal species have maintained duplicate copies in most RP families. How the RP genes were duplicated and maintained in these fungal species, and their functional significance remains unresolved. To address these questions, we identified all RP genes from 295 fungi and inferred the timing and nature of gene duplication for all RP families. We found that massive duplications of RP genes have independently occurred by different mechanisms in three distantly related lineages. The RP duplicates in two of them, budding yeast and Mucoromycota, were mainly created by whole genome duplication (WGD) events. However, in fission yeasts, duplicate RP genes were likely generated by retroposition, which is unexpected considering their dosage sensitivity. The sequences of most RP paralogs in each species have been homogenized by repeated gene conversion, demonstrating parallel concerted evolution, which might have facilitated the retention of their duplicates. Transcriptomic data suggest that the duplication and retention of RP genes increased RP transcription abundance. Physiological data indicate that increased ribosome biogenesis allowed these organisms to rapidly consuming sugars through fermentation while maintaining high growth rates, providing selective advantages to these species in sugar-rich environments.

evolutionary biology↗

bioRxiv2019

Structure-based validation can drastically under-estimate error rate in proteome-wide cross-linking mass spectrometry studies

Recent, rapid advances in cross-linking mass spectrometry (XL-MS) has enabled detection of novel protein-protein interactions and their structural dynamics at the proteome scale. Given the importance and scale of the novel interactions identified in these proteome-wide XL-MS studies, thorough quality assessment is critical. Almost all current XL-MS studies validate cross-links against known 3D structures of representative protein complexes. However, current structure validation approach only includes cross-links where both peptides mapped to the 3D structures. Here we provide theoretical and experimental evidence demonstrating this approach can drastically underestimate error rates for proteome-wide XL-MS datasets. Addressing current shortcomings, we propose and demonstrate a comprehensive set of four metrics, including orthogonal experimental validation to thoroughly assess quality of proteome-wide XL-MS datasets.

systems biology↗