Biology subjects

Tong, K.

Publications and source records attributed to Tong, K..

14 recordsLinked to original sources

bioRxiv2026

Synthetic cell-cell adhesion provides benefits of proximity without diffusion-related costs

Multicellularity provides many advantages over unicellularity, but accessing those benefits with synthetic biology approaches is challenging due to potential tradeoffs. The same diffusion limitations that exclude toxins can limit nutrient transport, and small intercellular spaces that facilitate sharing also increase local competition. To isolate when and how cell aggregation, an essential feature of multicellularity, is useful, we compared yeast strains differing only in expression of a single adhesion gene. Unexpectedly, aggregates containing thousands of cells showed no signs of diffusion limitations, growing as well as unicellular strains in both rich and toxic media, likely because reversible cell adhesion allows cells to exchange positions. Aggregating cells also grew better than unicellular strains in the limiting carbon conditions expected to increase diffusion limitations. In mixtures of different genotypes, these benefits were specific to cells expressing the adhesion protein. Together, these results suggest key design principles for engineering aggregates: reformable bonds provide a mechanism to sidestep the fitness costs of increased size, and the benefits within such groups may arise more from nutrient sharing than from diffusion limitations.

synthetic biology↗

bioRxiv2026

Identification of a neural circuit that enables safe, long-term torpor in mice

Torpor, a state of regulated hypothermia and hypometabolism, is a critical survival strategy in certain mammals. Previous studies established the primacy of neuron control in initiating acute torpor. However, whether neuromodulation can safely sustain torpor over the long term, and the specific governing pathways, remain unanswered challenges limiting its translational potential. Here we identify a distinct preoptic neuronal population expressing General control nonderepressible 2 ( G neurons) in mice that is essential and sufficient for torpor induction. Strikingly, sustained and selective activation of G neurons induces a stable, weeks-long torpid state (G neuron-driven long-term torpor, GLT), from which animals arouse without obvious behavioral deficits or tissue pathology. In contrast, long-term torpor driven by pan-neuronal activation of the same brain region (PLT) causes multiple post-recovery damages, underscoring the unique safety profile of selective pathway of G neurons. In a mouse cancer model, GLT directly suppressed tumor proliferation and markedly sensitized tumors to chemotherapy, achieving profound therapeutic outcomes. Together, we delineate a distinct neuronal population safely enabling prolonged torpor. This establishes a specific paradigm for inducing a stable and sustained hypometabolic state, providing a transformative platform for fundamental physiological research and pioneering clinical strategies against chronic pathologies such as cancer.

neuroscience↗

bioRxiv2026

The tongue-brain axis mediates a hidden amino acid appetite

Selecting a diet containing all essential amino acids (EAAs) is critical for health. Following EAA deprivation, animals can select a nutritiously complete food source; however, the underlying mechanisms in vertebrates remain unclear. In mice, we show that leucine deficiency activates hypothalamic agouti-related protein (AgRP) neurons, which project to the paraventricular thalamus (PVT) via {gamma}-aminobutyric acid and are required for EAA deficiency-induced leucine appetite in mice. Furthermore, the peripheral tongue amino acid sensor general control nonderepressive-2 (GCN2) mediates acute EAA appetite via AgRP neurons. Together, these findings identify a tongue-AgRP-PVT circuit underlying EAA appetite, which is important for the rapid and accurate selection of essential nutrients.

neuroscience↗

bioRxiv2025

The MTL200: a surface-based, probabilistic atlas of the medial temporal lobe

The human medial temporal lobe (MTL) includes a number of cortical regions surrounding the hippocampus implicated in long-term memory and high-level perception, including temporopolar cortex (TPC), entorhinal cortex (ERC), perirhinal cortex (PRC), and parahippocampal cortex (PHC). A critical prerequisite to studying the function of these areas is to define their anatomical boundaries. While several atlases or segmentations methods for automated boundary definition exist, they have limitations: 1) they are derived from small sample sizes; 2) most do not include TPC; and 3) they are volumetric rather than surface-based and are therefore not straightforward to use for surface-based analysis of MRI data. Here we present the MTL200: a surface-based, probabilistic atlas of MTL regions TPC, ERC, PRC, and PHC. The atlas is derived from hand drawings of regions based on gross anatomical features corresponding to cytoarchitectonic boundaries, using anatomical MRI data from 200 human participants. This work provides a valuable tool for researchers studying the anatomy and function of MTL regions, particularly the poorly characterized area TPC.

neuroscience↗

bioRxiv2025

Dynamic Reprogramming of Stromal Pdgfra-expressing cells during WNT-Mediated Transformation of the Intestinal Epithelium

Stromal fibroblasts regulate critical signaling gradients along the intestinal crypt-villus axis1 and provide a niche that supports adjacent epithelial stem cells. Here we report that Pdgfra-expressing fibroblasts secrete ligands that promote a regenerative-like state in the intestinal mucosa during early WNT-mediated tumorigenesis. Using a mouse model of WNT-driven oncogenesis and single-cell RNA sequencing (RNA-seq) of mesenchyme cell populations, we revealed a dynamic reprogramming of Pdgfra+ fibroblasts that facilitates WNT-mediated tissue transformation. Functional assays of potential mediators of cell-to-cell communication between these fibroblasts and the oncogenic epithelium revealed that TGFB signaling is notably induced in Pdgfra+ fibroblasts in the presence of oncogenic epithelium, and TGFB was essential to sustain regenerative-like growth of organoids ex vivo. Genetic reduction of Cdx2 in the {beta}-catenin mutant epithelium elevated the fetal-like/regenerative transcriptome and accelerated WNT-dependent onset of oncogenic transformation of the tissue in vivo. These results demonstrate that Pdgfra+ fibroblasts are activated during WNT-driven oncogenesis to promote a regenerative state in the epithelium that precedes and facilitates formation of tumors.

cancer biology↗

bioRxiv2024

Whole-genome duplication in the Multicellularity Long Term Evolution Experiment

Whole-genome duplication (WGD) is widespread across eukaryotes and can promote adaptive evolution1-4. However, given the instability of newly-formed polyploid genomes5-7, understanding how WGDs arise in a population, persist, and underpin adaptations remains a challenge. Using our ongoing Multicellularity Long Term Evolution Experiment (MuLTEE)8, we show that diploid snowflake yeast (Saccharomyces cerevisiae) under selection for larger multicellular size rapidly undergo spontaneous WGD. From its origin within the first 50 days of the experiment, tetraploids persist for the next 950 days (nearly 5,000 generations, the current leading edge of our experiment) in ten replicate populations, despite being genomically unstable. Using synthetic reconstruction, biophysical modeling, and counter-selection experiments, we found that tetraploidy evolved because it confers immediate fitness benefits in this environment, by producing larger, longer cells that yield larger clusters. The same selective benefit also maintained tetraploidy over long evolutionary timescales, inhibiting the reversion to diploidy that is typically seen in laboratory evolution experiments. Once established, tetraploidy facilitated novel genetic routes for adaptation, playing a key role in the evolution of macroscopic multicellular size via the origin of evolutionarily conserved aneuploidy. These results provide unique empirical insights into the evolutionary dynamics and impacts of WGD, showing how it can initially arise due to its immediate adaptive benefits, be maintained by selection, and fuel long-term innovations by creating additional dimensions of heritable genetic variation.

evolutionary biology↗

bioRxiv2023

Proteostatic tuning underpins the evolution of novel multicellular traits

The evolution of multicellularity paved the way for the origin of complex life on Earth, but little is known about the mechanistic basis of early multicellular evolution. Here, we examine the molecular basis of multicellular adaptation in the Multicellularity Long Term Evolution Experiment (MuLTEE). We demonstrate that cellular elongation, a key adaptation underpinning increased biophysical toughness and organismal size, is convergently driven by downregulation of the chaperone Hsp90. Mechanistically, Hsp90-mediated morphogenesis operates by destabilizing the cyclin-dependent kinase Cdc28, resulting in delayed mitosis and prolonged polarized growth. Reinstatement of Hsp90 or Cdc28 expression resulted in shortened cells that formed smaller groups with reduced multicellular fitness. Together, our results show how ancient protein folding systems can be tuned to drive rapid evolution at a new level of biological individuality by revealing novel developmental phenotypes. TeaserDownregulation of Hsp90 decouples cell cycle progression and growth to drive the evolution of macroscopic multicellularity.

evolutionary biology↗

bioRxiv2023

Global analysis of aging-related protein structural changes uncovers enzyme polymerization-based control of longevity

Aging is associated with progressive phenotypic changes over time. Virtually all cellular phenotypes are produced by proteins and structural alterations in proteins can lead to age-related diseases. Nonetheless, comprehensive knowledge of proteins undergoing structural-functional changes during cellular aging and their contribution to age-related phenotypes is lacking. Here, we conducted proteome-wide analysis of early age-related protein structural changes in budding yeast using limited proteolysis-mass spectrometry. The results, compiled in online ProtAge-catalog, unravelled age-related functional changes in regulators of translation, protein folding and amino acid metabolism. Mechanistically, we found that folded glutamate synthase Glt1 polymerizes into supramolecular self-assemblies during aging causing breakdown of cellular amino acid homeostasis. Inhibiting Glt1 polymerization by mutating the polymerization interface restored amino acid levels in aged cells, attenuated mitochondrial dysfunction and led to life span extension. Altogether, this comprehensive map of protein structural changes enables identifying novel mechanisms of age-related phenotypes and offers opportunities for their reversal.

cell biology↗

bioRxiv2021

Variation of female pronucleus reveal oocyte or embryo abnormality: An expert experience deep learning of non-dark box analysis

BackgroundPronuclear assessment appears to have the ability to distinguish good and bad embryos in the zygote stage, but paradoxical results were obtained in clinical studies. This situation might be caused by the robust qualitative detection of the development of dynamic pronuclei. Here, we aim to establish a quantitative pronuclear measurement method by applying expert experience deep learning from large annotated datasets. MethodsConvinced handle-annotated 2PN images (13419) were used for deep learning then corresponded errors were recorded through handle check for subsequent parameters adjusting. We used 790 embryos with 52479 PN images from 155 patients for analysis the area of pronuclei and the pre-implantation genetic test results. Establishment of the exponential fitting equation and the key coefficient {beta} 1was extracted from the model for quantitative analysis for pronuclear(PN) annotation and automatic recognition. FindingsBased on the female original PN coefficient {beta}1, the chromosome-normal rate in the blastocyst with biggest PN area is much higher than that of the blastocyst with smallest PN area (58.06% vs. 45.16%, OR=1.68 [1.07-2.64]; P=0.031). After adjusting coefficient {beta}1 by the first three frames which high variance of outlier PN areas was removed, coefficient {beta}1 at 12 hours and at 14 hours post-insemination, similar but stronger evidence was obtained. All these discrepancies resulted from the female propositus in the PGT-SR subgroup and smaller chromosomal errors. Conclusion(s)The results suggest that detailed analysis of the images of embryos could improve our understanding of developmental biology. FundingNone

genetics↗

bioRxiv2021

De novo evolution of macroscopic multicellularity

While early multicellular lineages necessarily started out as relatively simple groups of cells, little is known about how they became Darwinian entities capable of open-ended multicellular adaptation1,2. To explore this, we initiated the Multicellularity Long Term Evolution Experiment (MuLTEE), selecting for larger group size in the snowflake yeast (Saccharomyces cerevisiae) model system. Given the historical importance of oxygen limitation3, our ongoing experiment consists of three metabolic treatments4: anaerobic, obligately aerobic, and mixotrophic yeast. After 600 rounds of selection, snowflake yeast in the anaerobic treatment evolved to be macroscopic, becoming ~2{middle dot}104 times larger (~mm scale) and ~104-fold more biophysically tough, while retaining a clonal multicellular life cycle. They accomplished this through sustained biophysical adaptation, evolving increasingly elongate cells that initially reduced the strain of cellular packing, then facilitated branch entanglements that enabled groups of cells to stay together even after many cellular bonds fracture. In contrast, snowflake yeast competing for low oxygen remained microscopic, evolving to be just ~6-fold larger, underscoring the critical role of oxygen levels in the evolution of multicellular size. Taken together, this work provides unique insight into an ongoing evolutionary transition in individuality, showing how simple groups of cells overcome fundamental biophysical limitations via gradual, yet sustained, multicellular adaptation.

evolutionary biology↗

bioRxiv2021

A combination of RBD and NTD neutralizing antibodies limits the generation of SARS-CoV-2 spike neutralization-escape mutants

Most known SARS-CoV-2 neutralizing antibodies (nAbs), including those approved by the FDA for emergency use, inhibit viral infection by targeting the receptor-binding domain (RBD) of the spike (S) protein. Variants of concern (VOC) carrying mutations in the RBD or other regions of S reduce the effectiveness of many nAbs and vaccines by evading neutralization. Therefore, therapies that are less susceptible to resistance are urgently needed. Here, we characterized the memory B-cell repertoire of COVID-19 convalescent donors and analyzed their RBD and non-RBD nAbs. We found that many of the non-RBD-targeting nAbs were specific to the N-terminal domain (NTD). Using neutralization assays with authentic SARS-CoV-2 and a recombinant vesicular stomatitis virus carrying SARS-CoV-2 S protein (rVSV-SARS2), we defined a panel of potent RBD and NTD nAbs. Next, we used a combination of neutralization-escape rVSV-SARS2 mutants and a yeast display library of RBD mutants to map their epitopes. The most potent RBD nAb competed with hACE2 binding and targeted an epitope that includes residue F490. The most potent NTD nAb epitope included Y145, K150 and W152. As seen with some of the natural VOC, the neutralization potencies of COVID-19 convalescent sera were reduced by 4-16-fold against rVSV-SARS2 bearing Y145D, K150E or W152R spike mutations. Moreover, we found that combining RBD and NTD nAbs modestly enhanced their neutralization potential. Notably, the same combination of RBD and NTD nAbs limited the development of neutralization-escape mutants in vitro, suggesting such a strategy may have higher efficacy and utility for mitigating the emergence of VOC. ImportanceThe US FDA has issued emergency use authorizations (EUAs) for multiple investigational monoclonal antibody (mAb) therapies for the treatment of mild to moderate COVID-19. These mAb therapeutics are solely targeting the receptor binding domain of the SARS-CoV-2 spike protein. However, the N-terminal domain of the spike protein also carries crucial neutralizing epitopes. Here, we show that key mutations in the N-terminal domain can reduce the neutralizing capacity of convalescent COVID-19 sera. We report that a combination of two neutralizing antibodies targeting the receptor binding and N-terminal domains may have higher efficacy and is beneficial to combat the emergence of virus variants.

microbiology↗

bioRxiv2020

Characterization of the SARS-CoV-2 S Protein: Biophysical, Biochemical, Structural, and Antigenic Analysis

Coronavirus disease 2019 (COVID-19) is a global health crisis caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), and there is a critical need to produce large quantities of high-quality SARS-CoV-2 Spike (S) protein for use in both clinical and basic science settings. To address this need, we have evaluated the expression and purification of two previously reported S protein constructs in Expi293F and ExpiCHO-S cells, two different cell lines selected for increased expression of secreted glycoproteins. We show that ExpiCHO-S cells produce enhanced yields of both SARS-CoV-2 S proteins. Biochemical, biophysical, and structural (cryo-EM) characterization of the SARS-CoV-2 S proteins produced in both cell lines demonstrate that the reported purification strategy yields high quality S protein (non-aggregated, uniform material with appropriate biochemical and biophysical properties). Importantly, we show that multiple preparations of these two recombinant S proteins from either cell line exhibit identical behavior in two different serology assays. We also evaluate the specificity of S protein-mediated host cell binding by examining interactions with proposed binding partners in the human secretome. In addition, the antigenicity of these proteins is demonstrated by standard ELISAs, and in a flexible protein microarray format. Collectively, we establish an array of metrics for ensuring the production of high-quality S protein to support clinical, biological, biochemical, structural and mechanistic studies to combat the global pandemic caused by SARS-CoV-2.

biochemistry↗

bioRxiv2020

Polarized endosome dynamics engage cytosolic Par-3 and dynein during asymmetric division

Asymmetric cell division (ACD), which produces two daughters with different fates, is fundamental for generating cellular diversity. In the developing embryos of both invertebrates and vertebrates, asymmetrically dividing progenitors generate daughter cells with differential activity of Notch signaling1-7, a key regulator of cell fate decisions8,9. The cell polarity regulator Par-3 is critical for establishing this Notch asymmetry1,4,6, but the underlying mechanisms are not understood. Here, employing in vivo time-lapse imaging in the developing zebrafish forebrain during the mitotic cycle of radial glia, the principal vertebrate neural stem cells10,11, we show that during ACD, endosomes containing the Notch ligand Delta D (Dld) undergo convergent movement toward the cleavage plane, followed by preferential segregation into the posterior (and subsequently basal) Notchhi daughter. This asymmetric segregation requires the activity of Par-3 and the dynein motor complex. Employing label-retention expansion microscopy, we further detect Par-3 in the cytosol in association with the dynein light intermediate chain 1 (DLIC1) on Dld endosomes, suggesting a direct involvement of Par-3 in dynein-mediated polarized transport of Notch signaling endosomes. Our data reveal an unanticipated mechanism by which Par-3 regulates cell fate decision by directly polarizing Notch signaling components during ACD.

cell biology↗

bioRxiv2019

Negative plant-microbiome feedback limits productivity in aquaponics

The demand for food will outpace productivity of conventional agriculture due to projected growth of the human population, concomitant with shrinkage of arable land, increasing scarcity of freshwater, and a rapidly changing climate. Efforts to increase conventional agricultural output come with significant environmental impacts stemming from deforestation and excessive use of chemicals, including soil salinization, erosion, and nutrient runoffs. While aquaponics has potential to sustainably supplement food production with minimal environmental impact, there is a need to better characterize the complex interplay between the various components (fish, plant, microbiome) of these systems to optimize scale up and productivity. For instance, much of our knowledge of beneficial and detrimental microbial communities vis-a-vis crop productivity comes from studies on plant-microbiome interactions in soil. Here, we investigated how the practice of continued transfer of microbial communities from pre-existing systems might promote or impede productivity of aquaponics. Specifically, we monitored plant growth phenotypes, water chemistry, and microbiome composition of rhizospheres, biofilters, and fish feces over 61-days of lettuce (Lactuca sativa) growth in aquaponic systems inoculated with bacteria that were either commercially sourced or originating from a pre-existing aquaponic system. Strikingly, L. sativa plant and root growth was significantly reduced across all replicates inoculated with the established microbiome. Further analyses revealed the reduced productivity was potentially a consequence of plant-specific pathogen enrichment, including Pseudomonas, through transfer of microbiomes from pre-existing systems - a phenomenon consistent with negative feedbacks in soil ecology. These findings underscore the need for diagnostic tools to monitor microbiome composition, detect negative feedbacks early, and minimize pathogen accumulation in aquaponic systems.

microbiology↗