Biology subjects

Rhee, K.

Publications and source records attributed to Rhee, K..

15 recordsLinked to original sources

bioRxiv2026

Visualizing zinc-starved slow-replicating Mycobacterium tuberculosis in neutrophil-rich lung lesions

The months-long treatment for tuberculosis is partially attributed to a minor Mycobacterium tuberculosis (Mtb) subpopulation exhibiting phenotypic drug resistance, although origins of these bacilli in hosts remain unknown. Previously, we observed that mycobacteria respond to growth-restrictive limitations of zinc by accumulating Mrf, and inducing Mpy recruitment to the decoding center of the ribosome, thereby hibernating the ribosome in an inactive state. Mrf accumulation - regulated at both transcriptional and post-translational levels by Zur and Clp, respectively, in a zinc-dependent manner - facilitates the formation of stable Mpy-70S ribosome complex in zinc-starved cells. Here, using a translational fusion of Mrf with a fluorescent-reporter protein we visualized zinc-starved mycobacteria with a physiology conducive for Mpy-dependent ribosome hibernation. About half of zinc-starved mycobacterial population expressed microscopically detectable levels of the reporter and exhibited Mpy-dependent slowdown of growth and isoniazid tolerance in vitro. In chronic, high-burden mouse lung infections, ~3% of Mtb bacilli expressed the reporter, and the frequency increased to ~35% upon isoniazid treatment. Transcriptomics of mouse lungs suggests an important role of S100A8/A9 and ZIP2 in neutrophils in zinc mobilization during Mtb infections. Moreover, depletion of neutrophils, or loss of S100A9 caused significant attenuation of zinc starvation response in Mtb. Altogether, the study offers a new approach for visualizing and manipulating slow-replicating Mtb in host tissues.

microbiology↗

bioRxiv2025

Genes required for Mycobacterium tuberculosis to survive the transition from aerosol to pulmonary alveolar lining fluid and early infection in a model of transmission

Mycobacterium tuberculosis (Mtb) must survive multiple changes in environment for aerosol transmission. Our genome-wide screen for rehydration of modeled aerosols in surrogate pulmonary alveolar lining fluid identified a survival-sustaining role for 22 genes not required for survival during earlier stages of transmission, 20 of which are non-essential in routine culture. Thirteen genes were also needed for Mtb to survive in alveolar macrophages after passing through the earlier stages of transmission. Nine 9 of the 13 were needed for full infectivity of aerosols in mice. Seven of the genes sustaining the intra-alveolar survival of aerosolized Mtb are likely to regulate Mtbs uptake, catabolism or synthesis of lipids as regulated by cAMP. Our results reveal a dynamic form of conditional essentiality that only emerges after bacteria experience a sequence of other pathophysiologically relevant conditions. These findings enlarge Mtbs candidate transmission survival genome with stage-specific genes encoding potential targets for blocking tuberculosis transmission.

microbiology↗

bioRxiv2025

GPCR Antagonism via Antibody-Mediated Rewiring of Receptor Trafficking and Degradation

G-protein coupled receptors (GPCRs) represent one of the most important yet incompletely targeted classes of therapeutic proteins. Here, we report a novel strategy for functional GPCR antagonism through antibody-mediated endocytosis and lysosomal degradation. Our engineered bispecific antibodies, termed GPCR-TfR1 Targeting Chimeras (GTACs), achieve potent and selective downregulation of various GPCRs, including RXFP1 and CCR6, critical cancer and immune drug targets that are difficult to antagonize with conventional approaches. GTACs led to complete inhibition of receptor activity with over 100-fold greater potency than conventional antibody antagonists. Using four-color imaging, we elucidated the trafficking mechanism of both the target protein and TfR1 in living cells. The GTAC platform enables robust antagonism of signaling across diverse GPCR families and establishes induced endocytosis and lysosomal trafficking as a fundamentally new paradigm for therapeutic GPCR modulation.

bioengineering↗

bioRxiv2025

A periplasmic protein complex mediates cell envelope integrity and intrinsic multidrug resistance in Mycobacterium tuberculosis

The intrinsic drug resistance of Mycobacterium tuberculosis (Mtb) is a major barrier to effective tuberculosis (TB) treatment, largely due to its complex, impermeable cell envelope. We identified a periplasmic protein complex comprising FecB and Rv3035 that is essential for maintaining envelope integrity and mediating intrinsic multidrug resistance in Mtb. FecB interacts with Rv3035, forming a stable heterodimer that associates with the cell envelope biosynthesis protein AftB. We report the structures of Rv3035 alone and in complex with FecB and identify critical residues for complex formation and function. Co-essentiality and genetic interaction analyses support a functional link between FecB, Rv3035 and AftB, an arabinofuranosyltransferase which synthesizes arabinogalactan and lipoarabinomannan. Loss of FecB or Rv3035 disrupted AftB-mediated arabinan synthesis, suggesting that these proteins support AftBs enzymatic activity. Importantly, FecB is required for Mtb virulence in mice, underscoring its physiological relevance. These findings highlight FecB, Rv3035 and AftB as promising therapeutic targets.

microbiology↗

bioRxiv2023

Transferrin Receptor Targeting Chimeras (TransTACs) for Membrane Protein Degradation

Cancer cells require high levels of iron for rapid proliferation, leading to a significant upregulation of the iron carrier protein Transferrin Receptor (TfR) on their cell surface. Leveraging this phenomenon and the exceptionally fast endocytosis rate of TfR, we introduce Transferrin Receptor TArgeting Chimeras (TransTAC), a novel molecular archetype for membrane protein degradation in cancers and other cell types. TransTACs repurpose the naturally recycling receptor TfR1 for protein degradation. To accomplish this, we utilized a combination of protein engineering strategies to redirect the target protein from recycling-endosome trafficking to lysosomal degradation. We show that TransTACs can highly efficiently degrade a diverse range of single-pass, multi-pass, native, or synthetic membrane proteins, establishing new possibilities for targeted cancer therapy.

bioengineering↗

bioRxiv2023

Colorectal cancer detection and treatment with engineered probiotics

Bioengineered probiotics enable new opportunities to improve colorectal cancer (CRC) screening, prevention and treatment strategies. Here, we demonstrate the phenomenon of selective, long-term colonization of colorectal adenomas after oral delivery of probiotic E. coli Nissle 1917 (EcN) to a genetically-engineered murine model of CRC predisposition. We show that, after oral administration, adenomas can be monitored over time by recovering EcN from stool. We also demonstrate specific colonization of EcN to solitary neoplastic lesions in an orthotopic murine model of CRC. We then exploit this neoplasia-homing property of EcN to develop early CRC intervention strategies. To detect lesions, we engineer EcN to produce a small molecule, salicylate, and demonstrate that oral delivery of this strain results in significantly increased levels of salicylate in the urine of adenoma-bearing mice, in comparison to healthy controls. We also assess EcN engineered to locally release immunotherapeutics at the neoplastic site. Oral delivery to mice bearing adenomas, reduced adenoma burden by [~]50%, with notable differences in the spatial distribution of T cell populations within diseased and healthy intestinal tissue, suggesting local induction of robust anti-tumor immunity. Together, these results support the use of EcN as an orally-delivered platform to detect disease and treat CRC through its production of screening and therapeutic molecules.

bioengineering↗

bioRxiv2022

Cyclic AMP is a critical mediator of intrinsic drug resistance and fatty acid metabolism in M. tuberculosis

Cyclic AMP (cAMP) is a ubiquitous second messenger that transduces signals from cellular receptors to downstream effectors. Mycobacterium tuberculosis (Mtb), the etiological agent of tuberculosis, devotes a considerable amount of coding capacity to produce, sense, and degrade cAMP. Despite this fact, our understanding of how cAMP regulates Mtb physiology remains limited. Here, we took a genetic approach to investigate the function of the sole essential adenylate cyclase in Mtb H37Rv, Rv3645. We found that lack of rv3645 resulted in increased sensitivity to numerous antibiotics by a mechanism independent of substantial increases in envelope permeability. We made the unexpected observation that rv3645 is conditionally essential for Mtb growth only in the presence of long-chain fatty acids, a host-relevant carbon source. A suppressor screen further identified mutations in the atypical cAMP phosphodiesterase rv1339 that suppress both fatty-acid and drug sensitivity phenotypes in strains lacking rv3645. Using mass spectrometry, we found that Rv3645 is the dominant source of cAMP under standard laboratory growth conditions, that cAMP production is the essential function of Rv3645 in the presence of long-chain fatty acids, and that reduced levels of cAMP result in increased antibiotic susceptibility. Our work defines rv3645 and cAMP as central mediators of intrinsic multidrug resistance and fatty acid metabolism in Mtb and highlights the potential utility of small molecule modulators of cAMP signaling.

microbiology↗

bioRxiv2022

Longitudinal assessment of brain structure and behavior in youth with rapid weight gain: Potential contributing causes and consequences

ObjectiveIndependent of weight status, rapid weight gain has been associated with underlying brain structure variation in regions associated with food intake and impulsivity among pre-adolescents. Yet, we lack clarity on how developmental maturation coincides with rapid weight gain and weight stability. MethodsWe identified brain predictors of two-year rapid weight gain and its longitudinal effects on brain structure and impulsivity in the Adolescent Brain Cognitive DevelopmentSM Study(R). Youth were categorized as Healthy Weight/Weight Stable (WSHW, n=225) or Weight Gainers (WG, n=221, >38lbs); 63% of the WG group were healthy weight at 9-to-10-years-old. ResultsA five-fold cross-validated logistic elastic-net regression revealed that rapid weight gain was associated with structural variation amongst 39 brain features at 9-to-10-years-old in regions involved with executive functioning, appetitive control, and reward sensitivity. Two years later, WG youth showed differences in change over time in several of these regions and performed worse on measures of impulsivity. ConclusionsThese findings suggest that brain structure in pre-adolescence may predispose some to rapid weight gain and that weight gain itself may alter maturational brain change in regions important for food intake and impulsivity. Behavioral interventions that target inhibitory control may improve trajectories of brain maturation and facilitate healthier behaviors.

neuroscience↗

bioRxiv2021

Predictive modeling reveals that higher-order cooperativity drives transcriptional repression in a synthetic developmental enhancer

A challenge in quantitative biology is to predict output patterns of gene expression from knowledge of input transcription factor patterns and from the arrangement of binding sites for these transcription factors on regulatory DNA. We tested whether widespread thermodynamic models could be used to infer parameters describing simple regulatory architectures that inform parameter-free predictions of more complex enhancers in the context of transcriptional repression by Runt in the early fruit 2y embryo. By modulating the number and placement of Runt binding sites within an enhancer, and quantifying the resulting transcriptional activity using live imaging, we discovered that thermodynamic models call for higher-order cooperativity between multiple molecular players. This higher-order cooperativity capture the combinatorial complexity underlying eukaryotic transcriptional regulation and cannot be determined from simpler regulatory architectures, highlighting the challenges in reaching a predictive understanding of transcriptional regulation in eukaryotes and calling for approaches that quantitatively dissect their molecular nature.

biophysics↗

bioRxiv2021

A Comprehensive Overview of the Physical Health of the Adolescent Brain Cognitive Development Study (ABCD) Cohort at Baseline

Physical health in childhood is crucial for neurobiological as well as overall development, and can shape long-term outcomes into adulthood. The landmark, longitudinal Adolescent Brain Cognitive Development StudySM (ABCD study(R)), was designed to investigate brain development and health in almost 12,000 youth who were recruited when they were 9-10 years old and will be followed through adolescence and early adulthood. The overall goal of this paper is to provide descriptive analyses of physical health measures in the ABCD study at baseline, including but not limited to sleep, physical activity and sports involvement, and body mass index, and how these measures vary across demographic groups. This paper outlines how the physical health of the ABCD sample corresponds with that of the US population and highlights important avenues for health disparity research. This manuscript will provide important information for ABCD users and help guide analyses investigating physical health as it pertains to adolescent and young adult development.

developmental biology↗

bioRxiv2021

Diet Potentially Drives the Differentiation of Eating Behaviours via Alterations to the Gut Microbiome in Infants

Certain infant eating behaviours are associated with adverse health outcomes such as obesity. While a diet consisting of infant formula has been linked to higher-risk eating behaviours and changes in the gut microbiome, little is known about what role the gut microbiome plays in mediating eating behaviours. Using 16S rRNA sequences extracted from 96 fecal samples collected from 58 infants, we identified a subset of bacterial taxa that were more abundant in formula-fed infants, primarily composed of the phylum Firmicutes. The presence of these taxa correlated with a lower drive to eat (i.e., lower food responsiveness). Furthermore, short-chain fatty acid production pathways were significantly more abundant in formula-fed infants, negatively correlated with food responsiveness, and positively associated with relative abundance of the Firmicutes subset. Our results suggest that higher abundances of Firmicutes in formula-fed infants may decrease their food responsiveness through short-chain fatty acid production in the first four months of life. Taken together, these findings suggest a potential role for the infants diet in impacting eating behaviour via changes to the gut microbiome, which may lead to the development of novel interventions for the prevention of childhood obesity.

microbiology↗

bioRxiv2020

Fate of the M-phase-assembled centrioles during the cell cycle in the TP53;PCNT;CEP215-deleted cells

Cancer cells frequently include supernumerary centrioles. Here, we generated TP53;PCNT;CEP215 triple knockout cell lines and observed precocious separation and amplification of the centrioles at M phase. Many of the triple KO cells maintained supernumerary centrioles throughout the cell cycle. The M-phase-assembled centrioles lack an ability to function as templates for centriole assembly during S phase. They also lack an ability to organize microtubules in interphase. However, we found that a fraction of them acquired an ability to organize microtubules during M phase. Our works provide an example how supernumerary centrioles behave in dividing cells.

cell biology↗

bioRxiv2020

Metabolic homeostasis controls the diversity of cooperative swarming in pathogenic bacteria

Microbes have disproportionate impacts on the macroscopic world. This is in part due to their ability to grow to large groups and cooperatively secrete massive amounts of secondary metabolites that impact their environment. Yet, the conditions enabling secondary metabolism without compromising primary needs remain unclear. Here we investigated the biosynthesis of rhamnolipids, a secondary metabolite that Pseudomonas aeruginosa makes to decrease the surface tension of surrounding liquid. Using a combination of genomics, metabolomics, transcriptomics, and mathematical modeling we show that biosynthesis of rhamnolipids from glycerol varies inconsistently across the phylogenetic tree; instead, non-producer lineages are also those worse at reducing the oxidative stress of primary glycerol metabolism. The link to oxidative stress explains the inconsistent distribution across the P. aeruginosa tree, adding a new layer to the regulation of rhamnolipids--a microbial secondary metabolite important for fitness in natural and clinical settings. SignificanceThe bacterium Pseudomonas aeruginosa is a major source of hospital-acquired infections. This pathogens knack for virulence relies on its ability to multiply and secrete massive amounts of substances that overwhelm microbial competitors and weaken host defenses. It remains unclear how the bacteria conciliate their need to grow and multiply--a need at the individual-level-- with their ability to secrete products--a need of the population. Here we combined genomics, metabolomics and mathematical modeling to study the biosynthesis of rhamnolipids, a surfactant that P. aeruginosa makes to reduce the surface tension of surrounding liquids. Our study reveals a new link between oxidative stress and rhamnolipid synthesis, which helps explain how this important bacterial product has evolved and how it persists in many lineages of pathogens.

systems biology↗

bioRxiv2019

Ankylosis homologue (ANKH) controls extracellular citrate and pyrophosphate homeostasis and affects bone mechanical performance

The membrane protein Ankylosis homologue (ANKH, mouse orthologue: ANK) prevents mineralization of joint-space and articular cartilage. The accepted view is that ANKH mediates cellular release of inorganic pyrophosphate (PPi), a strong physiological inhibitor of mineralization. Using global metabolite profiling, we identified citrate as the most prominent metabolite leaving HEK293 cells in an ANKH-dependent manner. Although PPi levels were increased in culture medium of HEK293-ANKH cells, PPi was formed extracellularly after release of ATP and other nucleoside triphosphates. Ankank/ank mice, which lack functional ANK, had substantially reduced concentrations of citrate in plasma and urine, while citrate was undetectable in urine of a human patient lacking functional ANKH. Bone hydroxyapatite of Ankank/ank mice also contained markedly reduced levels of citrate and PPi and displayed diminished strength. Together, our data show that ANKH is a crucial factor in extracellular citrate and PPi homeostasis that is essential for normal bone development.

developmental biology↗

bioRxiv2019

Rv3722c governs aspartate-dependent nitrogen metabolism in Mycobacterium tuberculosis

Organisms are defined by their genomes, yet many distinguishing features of a given organism are encoded by genes that are functionally unannotated. Mycobacterium tuberculosis (Mtb), the leading cause of death due to a single microbe, co-evolved with humans as its only known natural reservoir, yet the factors mediating Mtbs pathogenicity remain incompletely defined. rv3722c is a gene of unknown function predicted to encode a pyridoxal phosphate binding protein and to be essential for in vitro growth of Mtb. Using metabolomic, genetic and structural approaches, we show that Rv3722c is the primary aspartate aminotransferase of Mtb and mediates an essential but underrecognized role in metabolism: nitrogen distribution. Together with the attenuation of Rv3722c-deficient Mtb in macrophages and mice, these results identify aspartate biosynthesis and nitrogen distribution as potential species-selective drug targets in Mtb.

microbiology↗