Biology subjects

Lee, S.-J.

Publications and source records attributed to Lee, S.-J..

27 records · Page 2Linked to original sources

bioRxiv2022

Characterization of altered molecular mechanisms in Parkinson disease through cell type-resolved multi-omics analyses

Parkinsons disease (PD) is a progressive neurodegenerative disorder. However, cell type-dependent transcriptional regulatory programs responsible for PD pathogenesis remain elusive. Here, we establish transcriptomic and epigenomic landscapes of the substantia nigra (SN) by profiling 87,733 nuclei obtained from healthy controls and PD patients. Our multi-omic data integration provides functional annotation of 128,724 cis-regulatory elements (cREs) and uncovers cell-type specific dysregulated cREs with a strong transcriptional influence on genes implicated in PD. The establishment of high-resolution three-dimensional chromatin contact maps identifies 656 target genes of dysregulated cREs and genetic risk loci, including both novel candidates and known PD risk genes. Notably, these new PD candidate genes exhibit modular gene expression patterns with unique molecular signatures in distinct cell types. Thus, our single-cell transcriptome and epigenome uncover cell type-specific disrupted transcriptional regulations in PD. TeaserSingle-cell transcriptome and epigenome uncover cell type-specific disrupted transcriptional regulations in Parkinsons disease.

genomics↗

bioRxiv2021

Macrophages break interneuromast cell quiescence by intervening the inhibition of Schwann cells in zebrafish lateral line

In the zebrafish lateral line system, interneuromast cells (INCs) between neuromasts are normally kept quiescent by underlying Schwann cells (SWCs). Upon severe injuries that cause the complete loss of an entire neuromast, INCs can occasionally differentiate into neuromasts but how they escape from the inhibition by SWCs is still unclear. Using a genetic/chemical method to specifically ablate a neuromast, we found a small portion of larvae can regenerate a new neuromast, but the regeneration was hindered by inhibiting macrophages. By in toto imaging, we further discovered heterogeneities in macrophage behavior and distribution along lateral line. We witnessed the crawling of macrophages in between injured lateral line and SWCs during regeneration and also in between the second primordium and the first mature lateral line during development. It implies that macrophages may physically separate and alleviate the inhibition from pLLn and SWCs to break the quiescence of INCs during regeneration and development in the zebrafish lateral line.

developmental biology↗

bioRxiv2021

Unraveling the roles of Mast4 in amelogenesis via regulating DLX3 and stem cell maintenance of mouse incisors

Asymmetric division of stem cells allows for maintenance of the cell population and differentiation for harmonious progress. Developing mouse incisors allows for examination of how the stem cell niche employs specific insights into essential phases. Microtubule-associated serine/threonine kinase family member 4 (Mast4) knockout (KO) mice showed abnormal incisor development with weak hardness as the apical bud was reduced and preameloblasts were shifted to the apical side, resulting in Amelogenesis Imperfecta. In addition, Mast4 KO incisors showed abnormal enamel maturation, and stem cell maintenance was inhibited as amelogenesis accelerated. Distal-Less Homeobox 3 (DLX3), known to be a critical factor Tricho-Dento-Osseous (TDO) syndrome, is considered to be responsible for Amelogenesis Imperfecta in humans. MAST4 directly binds to DLX3 and induces phosphorylation at three residues within the nuclear localization sites (NLS) that promote the nuclear translocation of DLX3. MAST4-mediated phosphorylation of DLX3 ultimately controls the transcription of DLX3 target genes, which are carbonic anhydrase and ion transporter genes involved in the pH regulation process during ameloblast maturation. Taken together, our data reveal a novel role of MAST4 as a critical regulator of ameloblast maturation, which controls DLX3 transcriptional activity.

developmental biology↗

bioRxiv2021

Moderate-intensity exercise might be a competitive protocol to prevent obesity and obesity-related metabolic diseases

To determine effects of exercise intensity on expression levels of cytokines and neurotransmitters beneficial for the prevention and improvement of obesity and obesity-related metabolic diseases. Expression levels of irisin, fibroblast growth factor-21 (FGF21), adiponectin, free fatty acid (FFA), dopamine (DA), and serotonin (5-HT) levels after moderate-intensity (50% of maximal oxygen uptake (VO2max)) and high-intensity treadmill running (80% VO2max) for 30 min in 20 healthy men were compared. Blood samples were collected at three time points: before treadmill running (pre-EX), immediately after treadmill running (post-EX), and at 60 min after treadmill running (60 min post-EX). Expression levels of irisin, FGF21, adiponectin, FFA, DA, and 5-HT were increased after 30 min of treadmill running exercise regardless of exercise intensity. Their levels were increased at 60 min post-EX. They showed no significant difference immediately after exercise regardless of exercise intensity. Only irisin, FGF21, FFA, and DA levels showed significant differences between moderate-intensity (50% VO2max) group and high-intensity group (80% VO2max) at 60 min post-EX. Considering that FFA level at 80% VO2max was significantly lower than that at 50% VO2max during post-EX and side effects of high-intensity exercise, moderate-intensity exercise would be a more competitive method compared to high-intensity for the prevention and improvement of obesity and obesity-related metabolic diseases.

physiology↗

bioRxiv2021

α-Synuclein polymorphism determines oligodendroglial dysfunction

Synucleinopathies, such as Parkinsons disease (PD) and Multiple System Atrophy (MSA) are progressive and unremitting neurological diseases. For both PD and MSA, -synuclein fibril inclusions inside brain cells are neuropathological hallmarks. In addition, amplification of -synuclein fibrils from body fluids is a potential biomarker distinguishing PD from MSA. However, little is known about the structure of -synuclein fibrils amplified from human samples and its connection to -synuclein fibril structure in the human brain. Here we amplified -synuclein fibrils from PD and MSA brain tissue, characterized its seeding potential in oligodendroglia, and determined the 3D structures by cryo-electron microscopy. We show that the -synuclein fibrils from a MSA patient are more potent in recruiting the endogenous -synuclein and evoking a redistribution of TPPP/p25 protein in mouse primary oligoden-droglial cultures compared to those amplified from a PD patient. Cryo-electron microscopy shows that the PD- and MSA-amplified -synuclein fibrils share a similar protofilament fold but differ in their inter-protofilament interface. The structures of the brain-tissue amplified -synuclein fibrils are also similar to other in vitro and ex vivo -synuclein fibrils. Together with published data, our results suggest that Syn fibrils differ between PD and MSA in their quaternary arrangement and could further vary between different forms of PD and MSA.

neuroscience↗

bioRxiv2021

Recombinant haplotypes reflect sexual reproduction in symbiotic arbuscular mycorrhizal fungi

Arbuscular mycorrhizal fungi (AMF) are part of the most widespread fungal-plant symbiosis. They colonize at least 80% of plant species, promote plant growth and plant diversity. These fungi are multinucleated and contain either one or two haploid nuclear genotypes (monokaryon and dikaryon) identified by the alleles at a putative mating-type locus. This taxon has been considered as an ancient asexual scandal because of the lack of observable sexual structures. Despite identification of a putative mating-type locus and functional activation of genes related to mating when two isolates co-exist, it remains unknown if AMF take part in a mainly sexual life cycle. We used publicly available genome sequences to test if dikaryon genomes display signatures of sexual reproduction in the form of reciprocal recombination patterns, or if they display signatures of parasexual reproduction involving gene conversion. We used short-read and long-read sequence data to identify nucleus genotype-specific haplotypes within dikaryons and then compared them to orthologous gene sequences from related monokaryon isolates displaying the same putative MAT-types. We observed that these genotype-specific haplotypes display reciprocal recombination and not gene conversion. These results are consistent with a sexual origin of the dikaryon rather than a parasexual origin and provides an important step to understand the life cycle of these globally important symbiotic fungi.

evolutionary biology↗

bioRxiv2020

Directed Evolution of Adenine Base Editors with Increased Activity and Therapeutic Application

The foundational adenine base editors (e.g. ABE7.10) enable programmable C*G to T*A point mutations but editing efficiencies can be low at challenging loci in primary human cells. Here we further evolve ABE7.10 using a library of adenosine deaminase variants to create ABE8s. At NGG PAM sites, ABE8s result in [~]1.5x higher editing at protospacer positions A5-A7 and [~]3.2x higher editing at positions A3-A4 and A8-A10 compared with ABE7.10. Non-NGG PAM variants have a [~]4.2-fold overall higher on-target editing efficiency than ABE7.10. In human CD34+ cells, ABE8 can recreate a natural allele at the promoter of the {gamma}-globin genes HBG1 and HBG2, with up to 60% efficiency, causing persistence of fetal hemoglobin. In primary human T cells, ABE8s achieve 98-99% target modification which is maintained when multiplexed across three loci. Delivered as mRNA, ABE8s induce no significant levels of sgRNA-independent off-target adenine deamination in genomic DNA and very low levels of adenine deamination in cellular mRNA.

genomics↗

bioRxiv2020

Next-generation cytosine base editors with minimized unguided DNA and RNA off-target events and high on-target activity

Abstract/introductory paragraphCytosine base editors (CBEs) are molecular machines which enable efficient and programmable reversion of T*A to C*G point mutations in the human genome without induction of DNA double strand breaks1, 2. Recently, the foundational cytosine base editor (CBE) BE3, containing rAPOBEC1, was reported to induce unguided, genomic DNA3, 4 and cellular RNA5 cytosine deamination when expressed in living cells. To mitigate spurious off-target events, we developed a sensitive, high-throughput cellular assay to select next-generation CBEs that display reduced spurious deamination profiles relative to rAPOBEC1-based CBEs, whilst maintaining equivalent or superior on-target editing frequencies. We screened 153 CBEs containing cytidine deaminase enzymes with diverse sequences and identified four novel CBEs with the most promising on/off target ratios. These spurious-deamination-minimized CBEs (BE4 with either RrA3F, AmAPOBEC1, SsAPOBEC3B, or PpAPOBEC1) were further optimized for superior on- and off-target DNA editing profiles through structure-guided mutagenesis of the deaminase domain. These next-generation CBEs display comparable overall DNA on-target editing frequencies, whilst eliciting a 10- to 49-fold reduction in C-to-U edits in the transcriptome of treated cells, and up to a 33-fold overall reduction in unguided off-target DNA deamination relative to BE4 containing rAPOBEC1. Taken together, these next-generation CBEs represent a new collection of base editing tools for applications in which minimization of spurious deamination is desirable and high on-target activity is required.

genomics↗

bioRxiv2019

Feasibility of real-time in vivo 89Zr-DFO-labeled CAR T-cell trafficking using PET imaging

IntroductionChimeric antigen receptor (CAR) T-cells have been developed recently, producing impressive outcomes in patients with hematologic malignancies. However, there is no standardized method for cell trafficking and in vivo CAR T-cell monitoring. We assessed the feasibility of real-time in vivo 89Zr-p-Isothiocyanatobenzyl-desferrioxamine (Df-Bz-NCS, DFO) labeled CAR T-cell trafficking using positron emission tomography (PET).\n\nResultsThe 89Zr-DFO radiolabeling efficiency of Jurkat/CAR and human peripheral blood mononuclear cells (hPBMC)/CAR T-cells was 70-79%, and cell radiolabeling activity was 98.1-103.6 kBq/106 cells. Cell viability after radiolabeling was >95%. Compared with unlabeled cells, cell proliferation was not significantly different during the early period after injection; however, the proliferative capacity decreased over time (p = 0.02, day 7 after labeling). IL-2 or IFN-{gamma} secretion was not significantly different between unlabeled and labeled CAR T-cells. PET/magnetic resonance images in the xenograft model showed that most of the 89Zr-DFO-labeled Jurkat/CAR T-cells were distributed in the lung (24.4% {+/-} 3.4%ID) and liver (22.9% {+/-} 5.6%ID) by 1 hour after injection. The cells gradually migrated from lung to the liver and spleen by day 1, and remained stably until day 7 (on day 7: lung 3.9% {+/-} 0.3%ID, liver 36.4% {+/-} 2.7%ID, spleen 1.4% {+/-} 0.3%ID). No significant accumulation of labeled cells was identified in tumors. A similar pattern was observed in ex vivo biodistributions on day 7 (lung 3.0% {+/-} 1.0%ID, liver 19.8% {+/-} 2.2%ID, spleen 2.3% {+/-} 1.7%ID). 89Zr-DFO-labeled hPBMC/CAR T-cells showed the similar distribution on serial PET images as Jurkat/CAR T-cells. The distribution of CAR T-cells was cross-confirmed by flow cytometry, Alu polymerase chain reaction, and immunohistochemistry.\n\nConclusionUsing PET imaging of 89Zr-DFO-labeled CAR T-cells, real time in vivo cell trafficking is feasible. It can be used to investigate cellular kinetics, initial in vivo biodistribution, and the safety profile in future CAR T-cell development.

cancer biology↗