Biology subjects

Lee, H.

Publications and source records attributed to Lee, H..

At least 217 records · Page 12Linked to original sources

bioRxiv2024

Netrin G1 Ligand is a new stromal immunomodulator that promotes pancreatic cancer

Understanding pancreatic cancer biology is fundamental for identifying new targets and for developing more effective therapies. In particular, the contribution of the stromal microenvironment to pancreatic cancer tumorigenesis requires further exploration. Here, we report the stromal roles of the synaptic protein Netrin G1 Ligand (NGL-1) in pancreatic cancer, uncovering its pro-tumor functions in cancer-associated fibroblasts and in immune cells. We observed that the stromal expression of NGL-1 inversely correlated with patients overall survival. Moreover, germline knockout (KO) mice for NGL-1 presented decreased tumor burden, with a microenvironment that is less supportive of tumor growth. Of note, tumors from NGL-1 KO mice produced less immunosuppressive cytokines and displayed an increased percentage of CD8+ T cells than those from control mice, while preserving the physical structure of the tumor microenvironment. These effects were shown to be mediated by NGL-1 in both immune cells and in the local stroma, in a TGF-{beta}-dependent manner. While myeloid cells lacking NGL-1 decreased the production of immunosuppressive cytokines, NGL-1 KO T cells showed increased proliferation rates and overall polyfunctionality compared to control T cells. CAFs lacking NGL-1 were less immunosuppressive than controls, with overall decreased production of pro-tumor cytokines and compromised ability to inhibit CD8+ T cells activation. Mechanistically, these CAFs downregulated components of the TGF-{beta} pathway, AP-1 and NFAT transcription factor families, resulting in a less tumor-supportive phenotype. Finally, targeting NGL-1 genetically or using a functionally antagonistic small peptide phenocopied the effects of chemotherapy, while modulating the immunosuppressive tumor microenvironment (TME), rather than eliminating it. We propose NGL-1 as a new local stroma and immunomodulatory molecule, with pro-tumor roles in pancreatic cancer. Statement of SignificanceHere we uncovered the pro-tumor roles of the synaptic protein NGL-1 in the tumor microenvironment of pancreatic cancer, defining a new target that simultaneously modulates tumor cell, fibroblast, and immune cell functions. This study reports a new pathway where NGL-1 controls TGF-{beta}, AP-1 transcription factor members and NFAT1, modulating the immunosuppressive microenvironment in pancreatic cancer. Our findings highlight NGL-1 as a new stromal immunomodulator in pancreatic cancer.

cancer biology↗

bioRxiv2024

Brain-wide functional connectome analysis of 40,000 individuals reveals brain networks that show aging effects in older adults

The functional connectome changes with aging. We systematically evaluated aging related alterations in the functional connectome using a whole-brain connectome network analysis in 39,675 participants in UK Biobank project. We used adaptive dense network discovery tools to identify networks directly associated with aging from resting-state fMRI data. We replicated our findings in 499 participants from the Lifespan Human Connectome Project in Aging study. The results consistently revealed two motor-related subnetworks (both permutation test p-values <0.001) that showed a decline in resting-state functional connectivity (rsFC) with increasing age. The first network primarily comprises sensorimotor and dorsal/ventral attention regions from precentral gyrus, postcentral gyrus, superior temporal gyrus, and insular gyrus, while the second network is exclusively composed of basal ganglia regions, namely the caudate, putamen, and globus pallidus. Path analysis indicates that white matter fractional anisotropy mediates 19.6% (p<0.001, 95% CI [7.6% 36.0%]) and 11.5% (p<0.001, 95% CI [6.3% 17.0%]) of the age-related decrease in both networks, respectively. The total volume of white matter hyperintensity mediates 32.1% (p<0.001, 95% CI [16.8% 53.0%]) of the aging-related effect on rsFC in the first subnetwork.

neuroscience↗

bioRxiv2024

Acoustic target strengths and swimbladder morphology of Chub mackerel Scomber japonicus in the Northwest Pacific Ocean

The Northwest Pacific chub mackerel (Scomber japonicus) is one of the most productive, economically important fishery resources worldwide. Due to fluctuations in their abundance and distribution, there is a pressing need to accurately assess this species and to ensure total allowable catch limits are followed. Acoustic target strength (TS; dB) measurements of Scomber japonicus were conducted at 38, 70, and 120 kHz using a split-beam echosounder of individuals from nine size groups (mean fork length, 10.8[~]28.3 cm) swimming freely in a net cage within a seawater tank. An underwater camera was utilized to simultaneously measure the swimming angle. A least-squares regression analysis revealed that when the slope was constrained to 20, as per the generally applicable morphometric equation, the resulting values for the constant term (b20) were -67.7, -66.6, and - 67.3 dB at 38, 70, and 120 kHz, respectively. The mean swimming angle of S. japonicus across the groups was -10.5[~]9.6{degrees} (standard deviation (SD), 16.3[~]33.3{degrees}). In addition, the ratio of swimbladder height to swimbladder length, swimbladder length to fork length, and tilt angle of the swimbladder (mean {+/-} SD) were 0.191 {+/-} 0.060, 0.245 {+/-} 0.055, and 9.6 {+/-} 3.0{degrees}, respectively. These results can be used for the acoustic stock assessment of S. japonicus in the Northwest Pacific Ocean.

animal behavior and cognition↗

bioRxiv2024

Targeting phosphatase DUSP22 ameliorates skeletal muscle wasting via Akt independent JNK-FOXO3a repression

Skeletal muscle wasting results from numerous conditions, such as sarcopenia, glucocorticoid therapy or intensive care. It prevents independent living in the elderly, predisposes to secondary diseases, and ultimately reduces lifespan. There is no approved drug therapy and the major causative mechanisms are not fully understood. Dual specificity phosphatase 22 (DUSP22) is a pleiotropic signaling molecule that plays important roles in immunity and cancer. However, the role of DUSP22 in skeletal muscle wasting is unknown. In this study, DUSP22 was found to be upregulated in sarcopenia patients and models of skeletal muscle wasting. DUSP22 knockdown or pharmacological inhibition prevented multiple forms of muscle wasting. Mechanistically, targeting DUSP22 suppressed FOXO3a, a master regulator of skeletal muscle wasting, via downregulation of the stress-activated kinase JNK, which occurred independently of aberrant Akt activation. DUSP22 targeting was also effective in human skeletal muscle cells undergoing atrophy. In conclusion, phosphatase DUSP22 is a novel target for preventing skeletal muscle wasting. The DUSP22-JNK-FOXO3a axis could be exploited to treat sarcopenia or related aging disorders.

cell biology↗

bioRxiv2024

Dynamic Tracking of Native Polyclonal Hematopoiesis in Adult Mice

Hematopoietic dysfunction has been associated with a reduction in the number of active precursors. However, precursor quantification at homeostasis and under diseased conditions is constrained by the scarcity of available methods. To address this issue, we optimized a method for quantifying a wide range of hematopoietic precursors. Assuming the random induction of a stable label in precursors following a binomial distribution, estimates depend on the inverse correlation between precursor numbers and the variance of precursor labeling among independent samples. Experimentally validated to cover the full dynamic range of hematopoietic precursors in mice (1 to 105), we utilized this approach to demonstrate that thousands of precursors, which emerge after modest expansion during fetal-to-adult transition, contribute to native and perturbed hematopoiesis. We further estimated the number of precursors in a mouse model of Fanconi Anemia, showcasing how repopulation deficits can be classified as autologous (cell proliferation) and non-autologous (lack of precursor). Our results support an accessible and reliable approach for precursor quantification, emphasizing the contemporary perspective that native hematopoiesis is highly polyclonal.

cell biology↗

bioRxiv2024

Selective regulation of corticostriatal synapses by astrocytic phagocytosis

In the adult brain, neural circuit homeostasis depends on the constant turnover of synapses via astrocytic phagocytosis mechanisms. However, it remains unclear whether this process occurs in a circuit-specific manner. Here, we reveal that astrocytes target and reorganize excitatory synapses in the striatum. Using model mice lacking astrocytic phagocytosis receptors in the dorsal striatum, we found that astrocytes constantly remove corticostriatal synapses rather than thalamostriatal synapses. This preferential elimination suggests that astrocytes play a selective role in modulating corticostriatal plasticity and functions via phagocytosis mechanisms. Supporting this notion, corticostriatal long-term potentiation (LTP) and the early phase of motor sequence learning are dependent on astrocytic phagocytic receptors. Together, our findings demonstrate that astrocytes contribute to the connectivity and plasticity of the striatal circuit by preferentially engulfing a specific subset of excitatory synapses within brain regions innervated by multiple excitatory sources.

neuroscience↗

bioRxiv2024

Engineering of highly active and diverse nuclease enzymes by combining machine learning and ultra-high-throughput screening

Optimizing enzymes to function in novel chemical environments is a central goal of synthetic biology, but optimization is often hindered by a rugged, expansive protein search space and costly experiments. In this work, we present TeleProt, an ML framework that blends evolutionary and experimental data to design diverse protein variant libraries, and employ it to improve the catalytic activity of a nuclease enzyme that degrades biofilms that accumulate on chronic wounds. After multiple rounds of high-throughput experiments using both TeleProt and standard directed evolution (DE) approaches in parallel, we find that our approach found a significantly better top-performing enzyme variant than DE, had a better hit rate at finding diverse, high-activity variants, and was even able to design a high-performance initial library using no prior experimental data. We have released a dataset of 55K nuclease variants, one of the most extensive genotype-phenotype enzyme activity landscapes to date, to drive further progress in ML-guided design.

bioinformatics↗

bioRxiv2024

Dimeric R25CPTH(1-34) Activates the Parathyroid Hormone-1 Receptor in vitro and Stimulates Bone Formation in Osteoporotic Female Mice

Osteoporosis, characterized by reduced bone density and strength, increases fracture risk, pain, and limits mobility. Established therapies of Parathyroid hormone (PTH) analogs effectively promote bone formation and reduce fractures in severe osteoporosis, but their use is limited by potential adverse effects. In the pursuit of safer osteoporosis treatments, we investigated R25CPTH, a PTH variant wherein the native arginine at position 25 is substituted by cysteine. These studies were prompted by our finding of high bone mineral density in a hypoparathyroidism patient with the R25C homozygous mutation, and we explored its effects on PTH type-1 receptor (PTH1R) signaling in cells and bone metabolism in mice. Our findings indicate that R25CPTH(1-84) forms dimers both intracellularly and extracellularly, and the synthetic dimeric peptide, R25CPTH(1-34), exhibits altered activity in PTH1R-mediated cAMP response. Upon a single injection in mice, dimeric R25CPTH(1-34) induced acute calcemic and phosphaturic responses comparable to PTH(1-34). Furthermore, repeated daily injections increased calvarial bone thickness in intact mice and improved trabecular and cortical bone parameters in ovariectomized (OVX) mice, akin to PTH(1-34). The overall results reveal a capacity of a dimeric PTH peptide ligand to activate the PTH1R in vitro and in vivo as PTH, suggesting a potential path of therapeutic PTH analog development.

genomics↗

bioRxiv2024

Neuroprotective role of Hippo signaling by microtubule stability control in C. elegans

The evolutionarily conserved Hippo (Hpo) pathway has been shown to impact early development and tumorigenesis by governing cell proliferation and apoptosis. However, its post-developmental roles are relatively unexplored. Here, we demonstrate its roles in post-mitotic cells by showing that defective Hpo signaling accelerates age-associated structural and functional decline of neurons in C. elegans. Loss of wts-1/LATS, the core kinase of the Hpo pathway, resulted in premature deformation of touch neurons and impaired touch responses in a yap-1/YAP-dependent manner, the downstream transcriptional co-activator of LATS. Decreased movement as well as microtubule destabilization by treatment with colchicine or disruption of microtubule stabilizing genes alleviated the neuronal deformation of wts-1 mutants. Colchicine exerted neuroprotective effects even during normal aging. In addition, the deficiency of a microtubule-severing enzyme spas-1 also led to precocious structural deformation. These results consistently suggest that hyper-stabilized microtubules in both wts-1-deficient neurons and normally aged neurons are detrimental to the maintenance of neuronal structural integrity. In summary, Hpo pathway governs the structural and functional maintenance of differentiated neurons by modulating microtubule stability, raising the possibility that the microtubule stability of fully developed neurons could be a promising target to delay neuronal aging. Our study provides potential therapeutic approaches to combat age- or disease-related neurodegeneration.

developmental biology↗

bioRxiv2024

Agrobacterium-mediated Cuscuta campestris transformation as a tool for understanding plant-plant interactions

Cuscuta campestris, a stem parasitic plant, has served as a valuable model plant for the exploration of plant-plant interactions and molecular trafficking. However, a major barrier to C. campestris research is that a method to generate stable transgenic plants has not yet been developed. Here, we describe the development of a Cuscuta transformation protocol using various reporter genes (GFP, GUS, or RUBY) and morphogenic genes (CcWUS2 and CcGRF/GIF), ultimately leading to a robust protocol for Agrobacterium-mediated C. campestris transformation. The stably transformed and regenerated RUBY C. campestris plants produced haustoria, the signature organ of parasitic plants, and these were functional in forming host attachments. The locations of T-DNA integration in the parasite genome were confirmed through TAIL-PCR. Transformed C. campestris also produced flowers and transgenic seeds exhibiting betalain pigment, providing proof of germline transmission of the RUBY transgene. Furthermore, the RUBY reporter is not only a useful selectable marker for the Agrobacterium-mediated transformation, but also provides insight into the movement of molecules from C. campestris to the host during parasitism. Thus, the protocol for transformation of C. campestris reported here overcomes a major obstacle to Cuscuta research and opens new possibilities for studying parasitic plants and their interactions with hosts.

plant biology↗

bioRxiv2024

Modeling heterogeneity in single-cell perturbation states enhances detection of response eQTLs

Identifying response expression quantitative trait loci (reQTLs) can help to elucidate mechanisms of disease associations. Typically, such studies model the effect of perturbation as discrete conditions. However, perturbation experiments usually affect perturbed cells heterogeneously. We demonstrated that modeling of per-cell perturbation state enhances power to detect reQTLs. We use public single-cell peripheral blood mononuclear cell (PBMC) data, to study the effect of perturbations with Influenza A virus (IAV), Candida albicans (CA), Pseudomonas aeruginosa (PA), and Mycobacterium tuberculosis (MTB) on gene regulation. We found on average 36.9% more reQTLs by accounting for single cell heterogeneity compared to the standard discrete reQTL model. For example, we detected a decrease in the eQTL effect of rs11721168 for PXK in IAV. Furthermore, we found that on average of 25% reQTLs have cell-type-specific effects. For example, in IAV the increase of the eQTL effect of rs10774671 for OAS1 was stronger in CD4+T and B cells. Similarly, in all four perturbation experiments, the reQTL effect for RPS26 was stronger in B cells. Our work provides a general model for more accurate reQTL identification and underscores the value of modeling cell-level variation.

genetics↗

bioRxiv2024

Identification of B cell subsets based on antigen receptor sequences using deep learning

B cell receptors (BCRs) denote antigen specificity, while corresponding cell subsets indicate B cell functionality. Since each B cell uniquely encodes this combination, physical isolation and subsequent processing of individual B cells become indispensable to identify both attributes. However, this approach accompanies high costs and inevitable information loss, hindering high-throughput investigation of B cell populations. Here, we present BCR-SORT, a deep learning model that predicts cell subsets from their corresponding BCR sequences by leveraging B cell activation and maturation signatures encoded within BCR sequences. Subsequently, BCR-SORT is demonstrated to improve reconstruction of BCR phylogenetic trees, and reproduce results consistent with those verified using physical isolation-based methods or prior knowledge. Notably, when applied to BCR sequences from COVID-19 vaccine recipients, it revealed inter-individual heterogeneity of evolutionary trajectories towards Omicron-binding memory B cells. Overall, BCR-SORT offers great potential to improve our understanding of B cell responses.

bioinformatics↗

bioRxiv2024

Gene Regulatory Network topology governs resistance and treatment escape in glioma stem-like cells

Poor prognosis and drug resistance in glioblastoma (GBM) manifests from heterogeneity and treatment-induced shifts in phenotypic states of tumor cells, including dedifferentiation to glioma stem-like cells (GSCs). This rare tumorigenic cell subpopulation is inherently resistant to temozolomide, undergoes proneural-to-mesenchymal transition (PMT) to evade therapy, and thereby drives recurrence. Through inference of transcriptional regulatory networks (TRNs) of patient-derived GSCs (PD-GSCs) at single-cell resolution, we demonstrate how topology of transcription factor interactions drives distinct trajectories of cell state transitions of susceptible and resistant PD-GSCs in response to cytotoxic drug treatment. By experimentally testing TRN simulation-based predictions, we show that drug treatment drives surviving cells of a PD-GSC along a trajectory of intermediate states, akin to a bottleneck in gene expression space, exposing vulnerability to potentiated killing by sequential addition of siRNA or a second drug targeting transcriptional programs governing non-genetic plasticity of a PD-GSC. Thus, our findings demonstrate an approach to uncover and use TRN topology of a PD-GSC to rationally predict combinatorial and sequential treatments that block treatment escape and acquired resistance in GBM.

systems biology↗

bioRxiv2024

Multiplexing of temporal and spatial information in the lateral entorhinal cortex

Episodic memory involves the processing of spatial and temporal aspects of personal experiences. The lateral entorhinal cortex (LEC) plays an essential role in subserving memory. However, the specific mechanism by which LEC integrates spatial and temporal information remains elusive. Here, we recorded LEC neurons while rats performed foraging and shuttling behaviors on one-dimensional, linear or circular tracks. Unlike open-field foraging tasks, many LEC cells displayed spatial firing fields in these tasks and demonstrated selectivity for traveling directions. Furthermore, some LEC neurons displayed changes in the firing rates of their spatial rate maps during a session, a phenomenon referred to as rate remapping. Importantly, this temporal modulation was consistent across sessions, even when the spatial environment was altered. Notably, the strength of temporal modulation was found to be greater in LEC compared to other brain regions, such as the medial entorhinal cortex (MEC), CA1, and CA3. Thus, the spatial rate mapping observed in LEC neurons may serve as a coding mechanism for temporal context, allowing for flexible multiplexing of spatial and temporal information.

neuroscience↗

bioRxiv2024

Repulsion from slow-diffusing nutrients improves chemotaxis towards moving sources

Chemotaxis, or the following of chemical concentration gradients, is essential for microbes to locate nutrients. However, microbes often display paradoxical behaviors, such as Escherichia coli being repelled by several amino acids. Here, we explore chemotaxis towards a moving source and demonstrate that when multiple nutrients are released from the source repulsion from certain nutrients actually improves chemotaxis towards the source. Because a moving source leaves most of the nutrient plume behind it, simply following the concentration gradient results in aiming behind the source and failing to intercept it. However, when attraction to a fast-diffusing nutrient and repulsion from a slow-diffusing nutrient are combined, motion in a new direction emerges and the chance of intercepting the source is increased up to six-fold. We demonstrate that this "differential strategy" is robust against numerous variations, including order-of-magnitude increases in the repellent release rate. Finally, we leverage existing data to show that E. coli is attracted to fast-diffusing amino acids and repelled by slow-diffusing ones, suggesting it may utilize a differential strategy and providing an explanation for its repulsion from these amino acids. Our results thus illuminate new possibilities in how microbes can integrate signals from multiple gradients to accomplish the difficult chemotactic tasks.

ecology↗

bioRxiv2024

Recruitment of the m6A/Am demethylase FTO to target RNAs by the telomeric zinc finger protein ZBTB48

N6-methyladenosine (m6A), the most abundant internal modification on eukaryotic mRNA, and N6, 2'-O-dimethyladenosine (m6Am), are epitranscriptomic marks that function in multiple aspects of posttranscriptional regulation. Fat mass and obesity-associated protein (FTO) can remove both m6A and m6Am; however, little is known about how FTO achieves its substrate selectivity. Here, we demonstrate that ZBTB48, a C2H2-zinc finger protein that functions in telomere maintenance, associates with FTO and binds both mRNA and the telomere-associated regulatory RNA TERRA to regulate the functional interactions of FTO with target transcripts. Specifically, depletion of ZBTB48 affects targeting of FTO to sites of m6A/m6Am modification, changes cellular m6A/m6Am levels and, consequently, alters decay rates of target RNAs. ZBTB48 ablation also accelerates growth of HCT-116 colorectal cancer cells and modulates FTO- dependent regulation of Metastasis-associated protein 1 (MTA1) transcripts by controlling the binding to MTA1 mRNA of the m6A reader IGF2BP2. Our findings thus uncover a previously unknown mechanism of posttranscriptional regulation in which ZBTB48 co-ordinates RNA- binding of the m6A/m6Am demethylase FTO to control expression of its target RNAs.

molecular biology↗

bioRxiv2023

A diverse proteome is present and enzymatically active in metabolite extracts

Metabolomics, a foundational tool in metabolism research, relies on the accurate transmittal of biochemical profiles underlying biological phenotypes. Over the years, workflows used in metabolomics have been assumed to remove enzymes to preserve metabolite levels during processing. Here, we uncover a diverse landscape of over 1,000 proteins, strongly enriched for metabolic enzymes, within metabolite extracts generated using common extraction workflows. Moreover, by combining in-extract stable isotope additions and enzyme inhibitors, we demonstrate transaminase activity, which is preventable by protein removal by 3 kDa filtration. We extend these findings to untargeted metabolomics, revealing that both post-extraction formation of glutamate dipeptide and depletion of total glutathione can also be prevented by removing proteins. Finally, we present a simple yet novel workflow that integrates passive filtration for protein removal of crude metabolite extracts as a superior method for broad-coverage metabolomics. Our findings have broad-reaching experimental implications across all fields that use metabolomics and molecular metabolism, especially cancer, immunology, and diabetes research.

biochemistry↗

bioRxiv2023

Sequential genetic, epigenetic alterations in human pluripotent stem cells for recurrent abnormality

Human embryonic stem cells (hESCs) are naturally equipped to maintain genome integrity to minimize genetic mutations during early embryo development. However, genetic aberration risks and subsequent cellular changes in hESCs during in vitro culture pose a significant threat to stem cell therapy. While a few studies have reported specific somatic mutations and copy number variations (CNVs), the molecular mechanisms underlying culture-adapted phenotype acquisitions of hESCs are largely unknown. Therefore, we conducted comprehensive genomic, single-cell transcriptomic, and single-cell ATAC-seq analyses of an isogenic hESC model displaying definitive culture-adapted phenotypes. Notably, hPSCs with a copy number gain of 20q11.21 during early passage did not present culture-adapted phenotypes nor BCL2L1 induction. Single-cell RNAseq and ATACseq analyses revealed active transcriptional regulation at 20q11.21 loci at late-passaged hESCs with the induced BCL2L1 and TPX2 to trigger culture-adapted phenotypes was associated with epigenetic changes facilitating TEA domain (TEAD) binding. These results suggest that copy number 20q11.21 gain and additional epigenetic changes are necessary for expressing culture-adapted phenotypes by activating gene transcription at this specific locus.

cell biology↗