Biology subjects

Lee, D.-H.

Publications and source records attributed to Lee, D.-H..

9 recordsLinked to original sources

bioRxiv2026

The Impact of TOR-S6K-eIF5A Signaling on Translational Control is Key to Plant Root Development

Nutrient-responsive Target of Rapamycin (TOR)-S6 kinase (S6K) signaling coordinates plant growth with protein synthesis, but how it interfaces with translation elongation to produce specific developmental outputs remains unclear. Here, we identify eukaryotic translation factor 5A (eIF5A) as an S6K-associated phosphoprotein in Arabidopsis thaliana. S6K1 and S6K2 associate with all three eIF5A isoforms and phosphorylate them in vitro, with Ser2 emerging as the major S6K1-responsive site in eIF5A-2. The corresponding N-terminal serine is invariant across the analyzed Archaeplastida eIF5A proteins. Conditional depletion of TOR, S6K1/2, or eIF5A produces overlapping reductions in primary root length and root hair coverage. eIF5A depletion leaves bulk protein synthesis and polysome profiles largely unchanged while selectively decreasing output from deca-proline reporters. Reporter activity is restored by amiRNA-resistant wild-type and phosphomimetic S2D eIF5A-2, whereas S2A does not restore activity, demonstrating the functional importance of the Ser2 state. Proteomic profiling identifies a restricted set of eIF5A-responsive proteins, and five of six tested insertion mutants display altered primary root growth, root hair coverage, or both. Together, these findings uncover a regulatory connection between S6K and eIF5A and establish eIF5A-dependent selective translation as a mechanism contributing to root development in A. thaliana.

molecular biology↗

bioRxiv2026

Infant gut microbiomes contribute to metabolic states that impact brain function

Alterations in the gut microbiome are associated with neurodevelopmental disorders, but causal mechanisms and therapeutic strategies remain undefined. Here, we demonstrate that human infant microbiomes isolated during the first six months of life drive behavioral impairments in mice and that microbiota-based interventions restore mice to normal behavior. Early-life microbiomes from twelve infants who later exhibited cognitive deficits at 2 years old (low-scoring) transferred adverse metabolic, brain, and behavioral phenotypes to mice, in contrast to microbiomes from twenty-three cognitively typical or high-scoring infants. Deficits in mice were rescued by fecal microbiota transplant from high-scoring infants or a rationally designed consortium that promoted amino acid levels. We confirmed lower fecal amino acid concentrations in low-scoring infants and replicated the association between early-life microbiome composition and cognitive outcomes in a second geographically independent infant cohort. Altogether, we discovered an early-life microbiome-mediated metabolic state causally linked to cognitive deficits and amenable to microbial intervention.

microbiology↗

bioRxiv2025

Structural basis of cyclic phytocytokine recognition by the HSL3/NUT receptor

Plant receptor kinases perceive diverse peptide signals to coordinate stress responses and developmental programs. The HAESA-LIKE 3 (HSL3/NUT) receptor recognizes CTNIP/SCREW phytocytokines--disulfide-stabilized cyclic peptides that regulate immune signaling and stress adaptation. However, how HSL3 distinguishes these structurally constrained cyclic peptides from linear signaling molecules remains unknown. Here we report near-atomic resolution cryo-EM structures of HSL3 in apo and CTNIP448-70-bound states at [~]2.6 [A], using Arabidopsis CTNIP4 as a representative family member, revealing distinct mechanisms for cyclic peptide recognition. The conserved CTNIP motif occupies a negatively charged pocket in HSL3s C-terminal region through a combination of polar contacts, hydrogen bonds, salt bridges, and van der Waals interactions. The receptor employs a two-step recognition mechanism--electrostatic steering followed by motif anchoring--that enables rapid ligand capture and release, consistent with the transient nature of stress signaling. Notably, an N-glycan at Asn449 directly contacts the CTNIP4 peptide, establishing glycosylation as an active participant in ligand recognition. Structure-guided mutagenesis combined with reactive oxygen species (ROS) burst assays confirmed the functional importance of key binding interfaces. N-terminal truncation experiments revealed a minimal active fragment: CTNIP451-70 supported both rapid ROS production and sustained seedling growth inhibition, whereas the shorter CTNIP454-70 variant retained ROS activity but failed to trigger long-term seedling growth inhibition. Structure-guided coevolutionary analysis across plant lineages reveals patterns of both conserved and variable receptor-ligand interfaces, highlighting evolutionary flexibility while preserving core features of recognition. These conserved recognition principles, mediated by receptor glycosylation and evolutionary plasticity, enable specificity in peptide signaling with implications for engineering stress-resilient crops.

plant biology↗

bioRxiv2025

Heartbeat-like dynamics drives oxygen activation in methane monooxygenase

Soluble methane monooxygenase (sMMO) is an enzyme that hydroxylates methane (CH4), a potent greenhouse gas, at non-heme di-iron active sites under atmospheric conditions. The regulatory component (MMOB) is essential for the catalytic activity of hydroxylase (MMOH) as it induces conformational changes in the active site and facilitating substrate ingress. Recent advances in cryogenic electron microscopy (cryo-EM) have enabled us to elucidate the high resolution picture of sMMO catalytic mechanism. We describe the 2.85 [A] cryo-EM structure of MMOH-MMOB, with one equivalent of MMOB bound to MMOH (H-1B), which is in contrast with previously solved crystal structures. MMOB allosterically regulates the MMOH protomer ({beta}{gamma}) and induces conformational changes that propagate from the surface to the di-iron coordination site. The N-terminal region of the MMOH {beta}-subunit (NT-H{beta}) stabilizes helices essential for iron coordination and oxygen activation. The MMOB-bound protomer (HBA, {beta}{gamma}B) presents the first structural report of a 2.7 [A] Fe{middle dot}{middle dot}{middle dot}Fe distance, while the non-MMOB-bound protomer (HBB, {beta}{gamma}) and MMOH display a 3.1 [A] distance. The coordination of Fe-ligands is maintained by the structural stabilization provided by the {beta}- and {gamma}-subunits of MMOH. This novel cryo-EM structure reveals new coordination environments, offering crucial mechanistic insights into sMMO catalysis.

biochemistry↗

bioRxiv2023

Insect immune resolution with EpOME/DiHOME and its dysregulation by their analogs leading to pathogen hypersensitivity

Epoxyoctadecamonoenoic acids (EpOMEs) are epoxide derivatives of linoleic acid (9,12-octadecadienoic acid: LA). They are metabolized into dihydroxyoctadecamonoenoic acids (DiHOMEs) in mammals. Unlike in mammals where they act as adipokines or lipokines, EpOMEs act as immunosuppressants in insects. However, the functional link between EpOMEs and pro-immune mediators such as PGE2 is not known. In addition, the physiological significance of DiHOMEs is not clear in insects. This study analyzed the physiological role of these C18 oxylipins using a lepidopteran insect pest, Spodoptera exigua. Immune challenge of S. exigua rapidly upregulated the expression of the phospholipase A2 gene to trigger C20 oxylipin biosynthesis, followed by the upregulation of genes encoding EpOME synthase (SE51385) and a soluble epoxide hydrolase (Se-sEH). The sequential gene expression resulted in the upregulations of the corresponding gene products such as PGE2, EpOMEs, and DiHOMEs. Interestingly, only PGE2 injection without the immune challenge significantly upregulated the gene expression of SE51825 and Se-sEH. The elevated levels of EpOMEs acted as immunosuppressants by inhibiting cellular and humoral immune responses induced by the bacterial challenge, in which 12,13-EpOME was more potent than 9,10-EpOME. However, DiHOMEs did not inhibit the cellular immune responses but upregulated the expression of antimicrobial peptides selectively suppressed by EpOMEs. The negative regulation of insect immunity by EpOMEs and their inactive DiHOMEs were further validated by synthetic analogs of the linoleate epoxide and corresponding diol. Furthermore, inhibitors specific to Se-sEH used to prevent EpOME degradation significantly suppressed the immune responses. The data suggest a physiological role of C18 oxylipins in resolving insect immune response. Any immune dysregulation induced by EpOME analogs or sEH inhibitors significantly enhanced insect susceptibility to the entomopathogen, Bacillus thuringiensis. Author summaryUpon immune challenge, recognition signal triggers insect immunity to remove the pathogens by cellular and humoral responses. Various immune mediators propagate the immune signals to nearby tissues, in which polyunsaturated fatty acid (PUFA) derivatives play crucial roles. However, little was known on how the insects terminate the activated immune responses after pathogen neutralization. Interestingly, C20 PUFA was detected at the early infection stage and later C18 PUFAs were induced in a lepidopteran insect, Spodoptera exigua. This study showed the role of epoxyoctadecamonoenoic acids (EpOMEs) in the immune resolution at the late infection stage to quench the excessive and unnecessary immune responses. In contrast, dihydroxy-octadecamonoenoates (DiHOMEs) were the hydrolyzed and inactive forms of EpOMEs. The hydrolysis is catalyzed by soluble epoxide hydrolase (sEH). Inhibitors specific to sEH mimicked the immunosuppression induced by EpOMEs. Furthermore, the inhibitor treatments significantly enhanced the bacterial virulence of Bacillus thuringiensis against S. exigua. This study proposes a negative control of the immune responses using EpOME/DiHOME in insects.

immunology↗

bioRxiv2022

Three consecutive glycolysis enzymes are involved in autophagic flux regulation through monitoring nutrient availability

Autophagy serves as an important recycling route for growth and survival of eukaryotic organisms in nutrient-deficient conditions. When confronted with starvation, metabolic flux is coordinated by individual metabolic enzymes. Given that the metabolic diversity of carbon in eukaryotes is related to their lifestyle, autophagy may be modulated by metabolic enzymes by monitoring carbon flux. Here, we attempted to identify carbon metabolic genes that modulate autophagy using VIGS screening of 45 glycolysis- and the Calvin-Benson cycle-related genes. We report here that three consecutive triose-phosphate-processing enzymes involved in the cytosolic glycolysis, TPI (triose-phosphate-isomerase), GAPC (glyceraldehyde-3-phosphate dehydrogenase), and PGK (phosphoglycerate kinase), designated TGP, negatively regulate autophagy. Depletion of TGP enzymes result in spontaneous autophagy induction and increases ATG1 kinase activity. TGP enzymes interact with ATG101, a regulatory component of the ATG1 kinase complex. Spontaneous autophagy induction and abnormal growth under insufficient sugar in the TGP mutants is suppressed by crossing with the atg101 mutant. Considering that triose-phosphates are photosynthates transported to the cytosol from active chloroplasts, the TGP enzymes may be strategically positioned to monitor the flow of photosynthetic sugars and modulate autophagy accordingly. Collectively, these results suggest that TGP enzymes negatively control autophagy acting upstream of the ATG1 complex, which is critical for seedling development.

plant biology↗

bioRxiv2021

Impact of sulfamethoxazole on a riverine microbiome

The continued emergence of bacterial pathogens presenting antimicrobial resistance is widely recognised as a global health threat and recent attention focused on potential environmental reservoirs of antibiotic resistance genes (ARGs). Freshwater environments such as rivers represent a potential hotspot for ARGs and antibiotic resistant bacteria as they are receiving systems for effluent discharges from wastewater treatment plants (WWTPs). Effluent also contains low levels of different antimicrobials including antibiotics and biocides. Sulfonamides are antibacterial chemicals widely used in clinical, veterinary and agricultural settings and are frequently detected in sewage sludge and manure in addition to riverine ecosystems. The impact of such exposure on ARG prevalence and diversity is unknown, so the aim of this study was to investigate the release of a sub-lethal concentration of the sulfonamide compound sulfamethoxazole (SMX) on the river bacterial microbiome using a flume system. This system was a semi-natural in vitro flume using river water (30 L) and sediment with circulation to mimic river flow. A combination of omics approaches were conducted to study the impact of SMX exposure on the microbiomes within the flumes. Metagenomic analysis showed that the addition of low concentrations of SMX (<4 g L-1) had a limited effect on the bacterial resistome in the water fraction only, with no impact observed in the sediment. Metaproteomics did not show differences in ARGs expression with SMX exposure in water. Overall, the river bacterial community was resilient to short term exposure to sub-lethal concentrations of SMX which mimics the exposure such communities experience downstream of WWTPs throughout the year.

microbiology↗

bioRxiv2021

High-throughput selection of microalgae based on biomass accumulation rates in production environments using PicoShell Particles

Production of high-energy lipids by microalgae may provide a sustainable, renewable energy source that can help tackle climate change. However, microalgae engineered to produce more lipids usually grow slowly, leading to reduced overall yields. Unfortunately, tools that enable the selection of cells based on growth while maintaining high biomass production, such as well-plates, water-in-oil droplet emulsions, and nanowell arrays do not provide production-relevant environments that cells experience in scaled-up cultures (e.g. bioreactors or outdoor cultivation farms). As a result, strains that are developed in the lab often do not exhibit the same beneficial phenotypic behavior when transferred to industrial production. Here we introduce PicoShells, picoliter-scale porous hydrogel compartments, that can enable >100,000 individual cells to be compartmentalized, cultured in production-relevant environments, and selected based on growth and biomass accumulation traits using standard flow cytometers. PicoShells consist of a hollow inner cavity where cells are encapsulated, and a porous outer shell that allows for continuous solution exchange with the external environment so that nutrients, cell-communication factors, and cytotoxic cellular byproducts can transport freely in and out of the inner cavity. PicoShells can also be placed directly into shaking flasks, bioreactors, or other production-relevant environments. We experimentally demonstrate that Chlorella sp. and Saccharomyces cerevisiae grow to significantly larger colony sizes in PicoShells than in water-in-oil droplet emulsions (P < 0.05). We have also demonstrated that PicoShells containing faster biomass accumulating Chlorella clonal colonies can be selected using a fluorescence-activated cell sorter and re-grown. Using the PicoShell process, we select a Chlorella population that accumulates biomass 8% faster than does an un-selected population after a single selection cycle.

bioengineering↗

bioRxiv2016

Parkin knockout inhibits neuronal development via regulation of proteasomal degradation of p21

PARK2 encodes for the E3 ubiquitin ligase parkin and iimplicates in the development of Parkinsons disease (PD). Although the neuroprotective role of parkin is well known, the mechanism of parkins function in neural stem differentiation is not clear. Co-expressions network analysis showed that SNAP25 and BDNF were positively correlated with parkin, but negatively correlated with p21 in human patient brain. Therefore, we investigated a link between the ubiquitin E3 ligase parkin and proteasomal degradation of p21 for the control of neural stem cell differentiation. We discovered that p21 directly binds with parkin and is ubiquitinated by parkin resulting in the loss of cell differentiation ability. Tranfection of p21 shRNA in PARK2 KO mice significantly rescued the differentiation efficacy as well as SNAP25 and BDNF expression. We also defined the decreased p21 ubiquitination and differentiation ability were reversed after treatment with JNK inhibitor, SP600125 in PARK2 KO mice derived neural stem cells. Thus, the present study indicated that parkin knockout inhibits neural stem cell differentiation by JNK-dependent proteasomal degradation of p21.\n\nSummary statementThe present study indicated that parkin knockout inhibits neural stem cell differentiation by JNK-dependent proteasomal degradation of p21.

cell biology↗