Biology subjects

Wang, H. H.

Publications and source records attributed to Wang, H. H..

At least 19 recordsLinked to original sources

bioRxiv2026

Directed evolution of the Drosophila microbiome improves intestinal health and extends lifespan

The gut microbiome and its bacterially derived metabolites are known to affect many aspects of the host organisms health, including metabolism, immune response, intestinal inflammation, oxidative stress, and even lifespan. Because pathological changes in the gut microbiome and these functions are associated with aging, many have hypothesized that we could protect against aging by generating beneficial changes to the gut microbiome. Here, we directed evolution outside of the host (ex vivo) and generated a Drosophila gut microbiome resistant to paraquat, a toxin that causes oxidative stress. Compared to a control microbiome, this paraquat-resistant (PQR) microbiome transplanted back into the Drosophila gut endowed the host with multiple health benefits: increased resistance to dietary paraquat, reduced age-related pathologies in the gut, and extended lifespan. We identified the beneficial species of the PQR microbiome as Lactiplantibacillus plantarum and further identified mutations specific to lifespan-extending isolates linked to greater production of acetate. Directly feeding this short-chain fatty acid, acetate, to Drosophila was sufficient to recapitulate an extended lifespan, similar to that induced by gut colonization of PQR bacteria in the gut. These results serve as a proof of principle that increasing the resistance of the microbiome to oxidative stress via directed ex vivo evolution could serve as a therapeutic strategy to protect against aging.

physiology↗

bioRxiv2025

Colibactin-producing E. coli promote carcinogenesis of gastroesophageal adenocarcinoma and simultaneously induce autophagy and differentiation

Background & AimsGastroesophageal adenocarcinoma (GEAC) is a malignancy of the gastroesophageal junction (GEJ) and is associated with reflux of gastroduodenal contents and Barretts Esophagus (BE). A shift towards gram-negative bacteria in the microbiota of the GEJ additionally promotes inflammation and likely carcinogenesis. Enterobacteriaceae are enriched in advanced stages of GEAC development, and members of this family can produce colibactin, a genotoxin implicated in DNA damage and tumor progression. We aimed to validate these observations and investigate the effect of E. coli with or without colibactin production on GEAC-carcinogenesis. MethodsBacteria were profiled in human biopsies with imaging and 16S rRNA gene sequencing. Organoids of our L2-IL1B mouse model of GEAC were exposed to E. coli with colibactin (CoPEC) and without colibactin production (noCoPEC) via organoid microinjection. The phenotypic and transcriptomic changes in the organoids after the coculture with E. coli were evaluated via histology and single-cell RNA sequencing. ResultsIn human specimens, we observed an infiltration of bacteria in GEJ-tissue upon tumor formation and detected Enterobacteriaceae in one third of BE-patients. CoPEC-injected organoids exhibited high rates of proliferation and DNA damage, and an upregulation of cancer-associated genes and pathways. Furthermore, genes and pathways associated with immune activation, defense mechanisms, metabolic reprogramming, autophagy and differentiation were upregulated in CoPEC-injected organoids. ConclusionIn addition to the enrichment of Enterobacteriaceae in the GEJ-tissue of patients at late stages of GEAC, we show that the exposure of colibactin-producing E. coli to murine BE-organoids promotes genetic instability and proliferation, and the activation of cancer-associated pathways, while also activating autophagy and enhancing intercellular homeostasis. This indicates that colibactin-producing E. coli have a dual effect on early stages of GEAC-carcinogenesis. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=120 SRC="FIGDIR/small/687579v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@1281a14org.highwire.dtl.DTLVardef@1c83c91org.highwire.dtl.DTLVardef@10088d2org.highwire.dtl.DTLVardef@1698a2_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗

bioRxiv2025

Genetic interaction of Specc1l and Thm1 reveals cytoskeletal - ciliary crosstalk

Cilium formation and actin cytoskeleton dynamics are interconnected, with evidence showing that elevated filamentous actin (F-actin) negatively regulates primary cilia length. Loss of the cytoskeletal protein SPECC1L, which itself does not localize to cilia, leads to increased F-actin and shortened cilia. Depolymerizing F-actin in Specc1l mutant cells restored cilia lengths, substantiating this inverse relationship. In cells harboring a Specc1l allele lacking only the coiled-coil domain 2, intracellular regions with both elevated and reduced F-actin were observed together with cilia shortening. Notably, F-actin was decreased at the ciliary base, suggesting that a different F-actin subpopulation contributes to the inverse relationship. We also identified a genetic interaction between Specc1l and Thm1, which encodes an intraflagellar transport-A (IFT-A) protein. Double or compound heterozygotes for Specc1l and Thm1 exhibited a higher penetrance of cleft palate compared to Specc1l heterozygotes alone. Together, these findings reveal a role for SPECC1L in cytoskeletal regulation of ciliogenesis affecting palate development.

cell biology↗

bioRxiv2025

Functional integrity of the SEL1L-HRD1 complex is critical for ERAD and organismal viability

The SEL1L-HRD1 complex represents the most evolutionarily conserved branch of endoplasmic reticulum-associated degradation (ERAD), with SEL1L acting as a key cofactor for the E3 ubiquitin ligase HRD1. While the physiological relevance of this complex has been increasingly recognized, whether SEL1L is strictly required for HRD1 function in mammals has remained unclear. Here, using complementary in vivo and in vitro approaches, we define the architecture and physiological significance of the mammalian SEL1L-HRD1 ERAD complex. Our data demonstrate that direct binding between SEL1L and HRD1 is essential for ERAD function and neonatal survival in mice. In three knock-in mouse models harboring targeted mutations at the SEL1L-HRD1 interface, we show that the L709P variant - unlike the benign P699T mutation - results in complete neonatal lethality within 30 hours of birth, a phenotype more severe than that of the partially lethal S658P variant. Mechanistically, the L709P mutation abolishes SEL1L-HRD1 interaction, disrupting substrate engagement and impairing recruitment of the E2 enzyme UBE2J1, leading to the accumulation and aggregation of misfolded proteins in the ER. Notably, these defects can be partially rescued by HRD1 overexpression, echoing findings from yeast. Together, our results provide definitive evidence that the SEL1L-HRD1 interaction is essential for ERAD activity and neonatal viability in mammals, resolving a long-standing question in ERAD biology and identifying a new therapeutic strategy for modulating ERAD activity in humans. SIGNIFICANCE STATEMENTThe endoplasmic reticulum-associated degradation (ERAD) pathway is essential for maintaining cellular proteostasis and organismal health, yet its molecular regulation in mammals remains incompletely defined. The SEL1L-HRD1 complex constitutes the central axis of this conserved pathway, but whether SEL1L is required for HRD1 function in vivo has remained unresolved. Here, using genetic mouse models and biochemical analyses, we demonstrate that the physical interaction between SEL1L and HRD1 is essential for ERAD activity and neonatal viability. These findings resolve a long-standing question in ERAD biology, provide new insights into the structural and functional organization of the mammalian ERAD machinery, and highlight the SEL1L-HRD1 interface as a potential therapeutic target for modulating ERAD activity in diseases.

cell biology↗

bioRxiv2024

UTX Epigenetically Imposes a Cytolytic Effector Program in Autoreactive Stem-like CD8+ T cell Progenitors

Type 1 Diabetes Mellitus (T1D) is a chronic disease caused by an unremitting autoimmune attack on pancreatic beta cells. This autoimmune chronicity is mediated by stem-like progenitor CD8+ T cells that continually repopulate the pool of beta cell-specific cytolytic effectors. Factors governing the conversion of progenitors to effectors, however, remain unclear. T1D has been linked to a chromosomal region (Xp13-p11) that contains the epigenetic regulator UTX, which suggests a key role for UTX in T1D pathogenesis. Here, we show that T cell-specific UTX deletion in NOD mice protects against T1D development. In T cells of NOD mice and T1D patients, UTX ablation resulted in the accumulation of CD8+ progenitor cells with concomitant deficiency of effectors, suggesting a key role for UTX in poising progenitors for transition to effectors. Mechanistically, UTXs role in T1D was independent of its inherent histone demethylase activity but instead relied on binding with transcription factors (TCF1 and STAT3) to co-regulate genes important in the maintenance and differentiation of progenitor CD8+ T cells. Together, these findings identify a critical role for UTX in T1D and the UTX:TCF1:STAT3 complex as a therapeutic target for terminating the long-lived autoimmune response.

immunology↗

bioRxiv2024

MCSPACE: inferring microbiome spatiotemporal dynamics from high-throughput co-localization data

Recent advances in high-throughput approaches for estimating co-localization of microbes, such as SAMPL-seq, allow characterization of the biogeography of the gut microbiome longitudinally and at unprecedented scale. However, these high-dimensional data are complex and have unique noise properties. To address these challenges, we developed MCSPACE, a probabilistic AI method that infers from microbiome co-localization data spatially coherent assemblages of taxa, their dynamics over time, and their responses to perturbations. To evaluate MCSPACEs capabilities, we generated the largest longitudinal microbiome co-localization dataset to date, profiling spatial relationships of microbes in the guts of mice subjected to serial dietary perturbations over 76 days. Analyses of these data and an existing human longitudinal dataset demonstrated superior benchmarking performance of MCSPACE over existing methods, and moreover yielded insights into spatiotemporal structuring of the gut microbiome, including identifying temporally persistent and dynamic microbial assemblages in the human gut, and shifts in assemblages in the murine gut induced by specific dietary components. Our results highlight the utility of our method, which we make available to the community as an open-source software tool, for elucidating dynamics of microbiome biogeography and gaining insights into the role of spatial relationships in host-microbial ecosystem function.

microbiology↗

bioRxiv2024

SAMPL-seq reveals micron-scale spatial hubs in the human gut microbiome

The local arrangement of microbes can profoundly impact community assembly, function, and stability. To date, little is known about the spatial organization of the human gut microbiome. Here, we describe a high-throughput and streamlined method, dubbed SAMPL-seq, that samples microbial composition of micron-scale sub-communities with split-and-pool barcoding to capture spatial colocalization in a complex consortium. SAMPL-seq analysis of the gut microbiome of healthy humans identified bacterial taxa pairs that consistently co-occurred both over time and across multiple individuals. These colocalized microbes organize into spatially distinct groups or "spatial hubs" dominated by Bacteroideceae, Ruminococceae, and Lachnospiraceae families. From a dietary perturbation using inulin, we observed reversible spatial rearrangement of the gut microbiome, where specific taxa form new local partnerships. Spatial metagenomics using SAMPL-seq can unlock new insights to improve the study of microbial communities. One Sentence SummaryHigh throughput micron-scale subcommunity sampling and sequencing identifies distinct spatial associations of gut bacteria within and across individuals.

systems biology↗

bioRxiv2024

Purkinje Cell-specific Deficiency in SEL1L-HRD1 Endoplasmic Reticulum-Associated Degradation Causes Progressive Cerebellar Ataxia in Mice

Recent studies have identified multiple genetic variants of SEL1L-HRD1 ER-associated degradation (ERAD) in humans with neurodevelopmental disorders and locomotor dysfunctions, including ataxia. However, the relevance and importance of SEL1L-HRD1 ERAD in the pathogenesis of ataxia remain unexplored. Here we show that SEL1L deficiency in Purkinje cells leads to early-onset progressive cerebellar ataxia with progressive loss of Purkinje cells with age. Mice with Purkinje cell-specific deletion of SEL1L (Sel1LPcp2Cre) exhibit motor dysfunction beginning around 9 weeks of age. Transmission electron microscopy (TEM) analysis reveals dilated ER and fragmented nuclei in Purkinje cells of adult Sel1LPcp2Cre mice, indicative of altered ER homeostasis and cell death. Lastly, loss of Purkinje cells is associated with a secondary neurodegeneration of granular cells, as well as robust activation of astrocytes and proliferation of microglia, in the cerebellum of Sel1LPcp2Cre mice. These data demonstrate the pathophysiological importance of SEL1L-HRD1 ERAD in Purkinje cells in the pathogenesis of cerebellar ataxia. One-sentence summarySEL1L-HRD1 ERAD is indispensable for Purkinje cell function and cerebellar ataxia pathogenesis in mice.

neuroscience↗

bioRxiv2024

SARS-CoV-2 Omicron BA.2.87.1 Exhibits Higher Susceptibility to Serum Neutralization Than EG.5.1 and JN.1

As SARS-CoV-2 continues to spread and mutate, tracking the viral evolutionary trajectory and understanding the functional consequences of its mutations remain crucial. Here, we characterized the antibody evasion, ACE2 receptor engagement, and viral infectivity of the highly mutated SARS-CoV-2 Omicron subvariant BA.2.87.1. Compared with other Omicron subvariants, including EG.5.1 and the current predominant JN.1, BA.2.87.1 exhibits less immune evasion, reduced viral receptor engagement, and comparable infectivity in Calu-3 lung cells. Intriguingly, two large deletions ({Delta}15-26 and {Delta}136-146) in the N-terminal domain (NTD) of the spike protein facilitate subtly increased antibody evasion but significantly diminish viral infectivity. Collectively, our data support the announcement by the USA CDC that the public health risk posed by BA.2.87.1 appears to be low.

microbiology↗

bioRxiv2023

Transcriptomic reprogramming screen identifies SRSF1 as rejuvenation factor

Aging is a complex process that manifests through the time-dependent functional decline of a biological system. Age-related changes in epigenetic and transcriptomic profiles have been successfully used to measure the aging process1,2. Moreover, modulating gene regulatory networks through interventions such as the induction of the Yamanaka factors has been shown to reverse aging signatures and improve cell function3,4. However, this intervention has safety and efficacy limitations for in vivo rejuvenation5,6, underscoring the need for identifying novel age reversal factors. Here, we discovered SRSF1 as a new rejuvenation factor that can improve cellular function in vitro and in vivo. Using a cDNA overexpression screen with a transcriptomic readout we identified that SRSF1 induction reprograms the cell transcriptome towards a younger state. Furthermore, we observed beneficial changes in senescence, proteasome function, collagen production, and ROS stress upon SRSF1 overexpression. Lastly, we showed that SRSF1 can improve wound healing in vitro and in vivo and is linked to organismal longevity. Our study provides a proof of concept for using transcriptomic reprogramming screens in the discovery of age reversal interventions and identifies SRSF1 as a promising target for cellular rejuvenation.

genomics↗

bioRxiv2023

GENTANGLE: integrated computational design of gene entanglements

SummaryThe design of two overlapping genes in a microbial genome is an emerging technique for adding more reliable control mechanisms in engineered organisms for increased safety. The design of functional gene pairs is a challenging procedure and computational design tools are used to improve the efficiency to deploy successful designs in genetically engineered systems. GENTANGLE (Gene Tuples ArraNGed in overLapping Elements) is a high performance containerized pipeline for the computational design of two overlapping genes translated in different reading frames of the genome. This new software package can be used to design and test gene entanglements for microbial engineering projects using arbitrary sets of user specified gene pairs. Availability and ImplementationThe GENTANGLE source code and its submodules are freely available on GitHub at https://github.com/BiosecSFA/gentangle. The DATANGLE (DATA for genTANGLE) repository contains related data and results, and is freely available on GitHub at https://github.com/BiosecSFA/datangle. The GENTANGLE repository wiki contains detailed instructions on how to use the container and the different components of software and data, including reproducing the results. The code is licensed under the GNU Affero General Public License version 3 (https://www.gnu.org/licenses/agpl.html). Contactmartimartine1@llnl.gov and allen99@llnl.gov

bioinformatics↗

bioRxiv2023

Antigenicity and receptor affinity of SARS-CoV-2 BA.2.86 spike

Although the COVID-19 pandemic has officially ended1, SARS-CoV-2 continues to spread and evolve. Recent infections have been dominated by XBB.1.5 and EG.5.1 subvariants2. A new subvariant designated BA.2.86 has just emerged, spreading to 21 countries in 5 continents3. This virus contains 34 spike mutations compared to its BA.2 predecessor, thereby raising concerns about its propensity to evade existing antibodies. We examined its antigenicity using human sera and monoclonal antibodies (mAbs). Reassuringly, BA.2.86 was not more resistant to human sera than XBB.1.5 and EG.5.1, indicating that the new subvariant would not have a growth advantage in this regard. Importantly, sera from patients who had XBB breakthrough infection exhibited robust neutralizing activity against all viruses tested, suggesting that upcoming XBB.1.5 monovalent vaccines could confer added protection. The finding that the longer genetic distance of BA.2.86 did not yield a larger antigenic distance was partially explained by the mAb data. While BA.2.86 showed greater resistance to mAbs to subdomain 1 (SD1) and receptor-binding domain (RBD) class 2 and 3 epitopes, it was more sensitive to mAbs to class 1 and 4/1 epitopes in the "inner face" of RBD that is exposed only when this domain is in the "up" position. We also identified six new spike mutations that mediate antibody resistance, including E554K that threatens SD1 mAbs in clinical development. The BA.2.86 spike also had a remarkably high receptor affinity. The ultimate trajectory of this new SARS-CoV-2 variant will soon be revealed by continuing surveillance, but its worldwide spread is worrisome.

microbiology↗

bioRxiv2023

SEL1L-HRD1 ER-associated degradation is a new ataxia gene

The SEL1L-HRD1 protein complex represents the most conserved branch of endoplasmic reticulum (ER)-associated degradation (ERAD); however, definitive evidence for the importance of SEL1L in HRD1 ERAD is lacking. Here we report that attenuation of the interaction between SEL1L and HRD1 impairs HRD1 ERAD function and has pathological consequences in mice. Our data show that SEL1L variant p.Ser658Pro (SEL1LS658P) previously identified in Finnish Hound suffering cerebellar ataxia is a recessive hypomorphic mutation, causing partial embryonic lethality, developmental delay, and early-onset cerebellar ataxia in homozygous mice carrying the bi-allelic variant. Mechanistically, SEL1LS658P variant attenuates the SEL1L-HRD1 interaction and causes HRD1 dysfunction by generating electrostatic repulsion between SEL1L F668 and HRD1 Y30 residues. Proteomic screens of SEL1L and HRD1 interactomes revealed that the SEL1L-HRD1 interaction is prerequisite for the formation of a functional HRD1 ERAD complex, as SEL1L recruits not only the lectins OS9 and ERLEC1, but the E2 UBE2J1 and retrotranslocon DERLIN, to HRD1. These data underscore the pathophysiological importance and disease relevance of the SEL1L-HRD1 complex, and identify a key step in organizing the HRD1 ERAD complex.

cell biology↗

bioRxiv2023

Bacterial genome engineering using CRISPR RNA-guided transposases

CRISPR-associated transposons (CASTs) have the potential to transform the technology landscape for kilobase-scale genome engineering, by virtue of their ability to integrate large genetic payloads with high accuracy, easy programmability, and no requirement for homologous recombination machinery. These transposons encode efficient, CRISPR RNA-guided transposases that execute genomic insertions in E. coli at efficiencies approaching [~]100%, generate multiplexed edits when programmed with multiple guides, and function robustly in diverse Gram-negative bacterial species. Here we present a detailed protocol for engineering bacterial genomes using CAST systems, including guidelines on the available homologs and vectors, customization of guide RNAs and DNA payloads, selection of common delivery methods, and genotypic analysis of integration events. We further describe a computational crRNA design algorithm to avoid potential off-targets and CRISPR array cloning pipeline for DNA insertion multiplexing. Starting from available plasmid constructs, the isolation of clonal strains containing a novel genomic integration event-of-interest can be achieved in 1 week using standard molecular biology techniques.

bioengineering↗

bioRxiv2022

Alarming antibody evasion properties of rising SARS-CoV-2 BQ and XBB subvariants

The SARS-CoV-2 Omicron variant continues to evolve, with new BQ and XBB subvariants now rapidly expanding in Europe/US and Asia, respectively. As these new subvariants have additional spike mutations, they may possess altered antibody evasion properties. Here, we report that neutralization of BQ.1, BQ.1.1, XBB, and XBB.1 by sera from vaccinees and infected persons was markedly impaired, including sera from individuals who were boosted with a WA1/BA.5 bivalent mRNA vaccine. Compared to the ancestral strain D614G, serum neutralizing titers against BQ and XBB subvariants were lower by 13-81-fold and 66-155-fold, respectively, far beyond what had been observed to date. A panel of monoclonal antibodies capable of neutralizing the original Omicron variant, including those with Emergency Use Authorization, were largely inactive against these new subvariants. The spike mutations that conferred antibody resistance were individually studied and structurally explained. Finally, the ACE2-binding affinities of the spike proteins of these novel subvariants were found to be similar to those of their predecessors. Taken together, our findings indicate that BQ and XBB subvariants present serious threats to the efficacy of current COVID-19 vaccines, render inactive all authorized monoclonal antibodies, and may have gained dominance in the population because of their advantage in evading antibodies.

microbiology↗

bioRxiv2022

Spatiotemporal dynamics during niche remodeling by super-colonizing microbiota in the mammalian gut

While fecal microbiota transplantation (FMT) has been shown to be effective in reversing gut dysbiosis, we lack an understanding for the fundamental processes underlying microbial engraftment in the mammalian gut. Here, we explored a murine gut colonization model leveraging natural inter-individual variations in gut microbiomes to elucidate the spatiotemporal dynamics of FMT. We identified a natural super-donor consortium that universally engrafts into diverse recipients and resists reciprocal colonization. Temporal profiling of the gut microbiome showed an ordered succession of rapid engraftment by early colonizers within 72 hours followed by a slower emergence of late colonizers over 15-30 days. Moreover, engraftment was localized to distinct compartments of the gastrointestinal tract in a species-specific manner. Spatial metagenomic characterization suggested engraftment was mediated by simultaneous transfer of spatially co-localizing species from the super-donor consortia. These results offer a mechanism of super-donor colonization by which nutritional niches are expanded in a spatiotemporally- dependent manner.

microbiology↗

bioRxiv2022

Antibody Evasion Properties of SARS-CoV-2 Omicron Sublineages

The identification of the Omicron variant (B.1.1.529.1 or BA.1) of SARS-CoV-2 (severe acute respiratory syndrome coronavirus 2) in Botswana in November 20211 immediately raised alarms due to the sheer number of mutations in the spike glycoprotein that could lead to striking antibody evasion. We2 and others3-6 recently reported results in this Journal confirming such a concern. Continuing surveillance of Omicron evolution has since revealed the rise in prevalence of two sublineages, BA.1 with an R346K mutation (BA.1+R346K) and B.1.1.529.2 (BA.2), with the latter containing 8 unique spike mutations while lacking 13 spike mutations found in BA.1. We therefore extended our studies to include antigenic characterization of these new sublineages. Polyclonal sera from patients infected by wild-type SARS-CoV-2 or recipients of current mRNA vaccines showed a substantial loss in neutralizing activity against both BA.1+R346K and BA.2, with drops comparable to that already reported for BA.12,3,5,6. These findings indicate that these three sublineages of Omicron are antigenically equidistant from the wild-type SARS-CoV-2 and thus similarly threaten the efficacies of current vaccines. BA.2 also exhibited marked resistance to 17 of 19 neutralizing monoclonal antibodies tested, including S309 (sotrovimab)7, which had retained appreciable activity against BA.1 and BA.1+R346K2-4,6. This new finding shows that no presently approved or authorized monoclonal antibody therapy could adequately cover all sublineages of the Omicron variant.

microbiology↗

bioRxiv2021

Structural basis for antibody resistance to SARS-CoV-2 omicron variant

The recently reported B.1.1.529 Omicron variant of SARS-CoV-2 includes 34 mutations in the spike protein relative to the Wuhan strain that initiated the COVID-19 pandemic, including 15 mutations in the receptor binding domain (RBD). Functional studies have shown omicron to substantially escape the activity of many SARS-CoV-2-neutralizing antibodies. Here we report a 3.1 [A] resolution cryo-electron microscopy (cryo-EM) structure of the Omicron spike protein ectodomain. The structure depicts a spike that is exclusively in the 1-RBD-up conformation with increased mobility and inter-protomer asymmetry. Many mutations cause steric clashes and/or altered interactions at antibody binding surfaces, whereas others mediate changes of the spike structure in local regions to interfere with antibody recognition. Overall, the structure of the omicron spike reveals how mutations alter its conformation and explains its extraordinary ability to evade neutralizing antibodies. HighlightsO_LISARS-CoV-2 omicron spike exclusively adopts 1-RBD-up conformation C_LIO_LIOmicron substitutions alter conformation and mobility of RBD C_LIO_LIA subset of omicron mutations change the local conformation of spike C_LIO_LIThe structure reveals the basis of antibody neutralization escape C_LI

immunology↗