Backmapping with Mapping and Isomeric Information.

I present a powerful and flexible backmapping tool named Multiscale Simulation Tool (mstool) that converts a coarse-grained (CG) system into all-atom (AA) resolution and only requires AA to CG mapping and isomeric information (cis/trans/dihedral/chiral). The backmapping procedure includes two simple steps: (a) AA atoms are randomly placed near the corresponding CG beads according to the provided mapping scheme. (b) Energy minimization is performed with two modifications in the AA force field (FF). First, nonbonded interactions are replaced with cosine functions to ensure the numerical stability. Second, additional torsions are imposed to maintain the molecules' isomeric properties. To test the simplicity and robustness of the tool, I backmapped multiple membrane and protein CG structures into AA resolution, including a four-bead CG lipid model (resolution increased by a factor of 34) without using intermediate resolution. The tool is freely available at github.com/ksy141/mstool.

The structure of phosphatidylinositol remodeling MBOAT7 reveals its catalytic mechanism and enables inhibitor identification.

Cells remodel glycerophospholipid acyl chains via the Lands cycle to adjust membrane properties. Membrane-bound O-acyltransferase (MBOAT) 7 acylates lyso-phosphatidylinositol (lyso-PI) with arachidonyl-CoA. MBOAT7 mutations cause brain developmental disorders, and reduced expression is linked to fatty liver disease. In contrast, increased MBOAT7 expression is linked to hepatocellular and renal cancers. The mechanistic basis of MBOAT7 catalysis and substrate selectivity are unknown. Here, we report the structure and a model for the catalytic mechanism of human MBOAT7. Arachidonyl-CoA and lyso-PI access the catalytic center through a twisted tunnel from the cytosol and lumenal sides, respectively. N-terminal residues on the ER lumenal side determine phospholipid headgroup selectivity: swapping them between MBOATs 1, 5, and 7 converts enzyme specificity for different lyso-phospholipids. Finally, the MBOAT7 structure and virtual screening enabled identification of small-molecule inhibitors that may serve as lead compounds for pharmacologic development.

Structure and function of lipid droplet assembly complexes.

Cells store lipids as a reservoir of metabolic energy and membrane component precursors in organelles called lipid droplets (LDs). LD formation occurs in the endoplasmic reticulum (ER) at LD assembly complexes (LDAC), consisting of an oligomeric core of seipin and accessory proteins. LDACs determine the sites of LD formation and are required for this process to occur normally. Seipin oligomers form a cage-like structure in the membrane that may serve to facilitate the phase transition of neutral lipids in the membrane to form an oil droplet within the LDAC. Modeling suggests that, as the LD grows, seipin anchors it to the ER bilayer and conformational shifts of seipin transmembrane segments open the LDAC dome toward the cytoplasm, enabling the emerging LD to egress from the ER.

Effects of Exhaustive Exercise on Inflammatory, Apoptotic, and Antioxidative Signaling Pathways in Human Peripheral Blood Mononuclear Cells.

In the present study, we investigated the effects of exhaustive exercise and recovery on inflammatory, pro-apoptotic, and anti-oxidative responses in human peripheral blood mononuclear cells (PBMCs). Sixteen volunteers participated in a guided physical activity program in which they were subjected to progressive exercise on the treadmill until they were exhausted followed by an 1-hour recovery period. Isolated human PBMCs were collected before exercise, immediately after exercise, and after 1-hour recovery. Exhaustive exercise induced expression of heme oxygenase-1 and glutamate cysteine ligase catalytic subunit and activation of NF-κB and NF-E2 related factor 2 (Nrf2). Apoptosis, as measured by activity and cleavage of caspase-3 and its substrate PARP also significantly increased. However, induction of redox signaling and the pro-apoptotic response fully returned to the baseline level during the 1-hour recovery period. On the other hand, COX-2 expression was continuously elevated after exercise cessation throughout the 1-hour recovery period. Taking all these findings into account, we conclude that exhaustive exercise transiently induces Nrf2-mediated antioxidant gene expression and eliminates damaged cells through apoptosis as part of an adaptive cytoprotective response against oxidative and inflammatory stress.

A Phase II Trial of S-1 and Oxaliplatin in Patients with Metastatic Breast Cancer Previously Treated with Anthracycline and Taxane (KCSG-BR07-03).

PURPOSE: This single-arm phase II trial investigate the efficacy and safety of S-1 plus oxaliplatin (SOX) in patients with metastatic breast cancer. Materials and Methods: Patients with metastatic breast cancer previously treated with anthracyclines and taxanes were enrolled. Patients received S-1 (40-60 mg depending on patient's body surface area, twice a day, day 1-14) and oxaliplatin (130 mg/m2, day 1) in 3 weeks cycle until disease progression or unacceptable toxicity. The primary endpoint was objective response rate (ORR) according to Response Evaluation Criteria in Solid Tumor 1.1. Secondary endpoints included time-to-progression (TTP), duration-of-response (DoR), overall survival (OS), and adverse events. RESULTS: A total of 87 patients were enrolled from 11 institutions in Korea. Hormone receptor was positive in 54 (62.1%) patients and six (6.9%) had human epidermal growth factor receptor 2-positive disease. Forty-eight patients (85.1%) had visceral metastasis and 74 (55.2%) had more than three sites of metastases. The ORR of SOX regimen was 38.5% (95% confidence interval [CI], 26.9 to 50.0) with a median TTP of 6.0 months (95% CI, 5.1 to 6.9). Median DoR and OS were 10.3 months (95% CI, 5.5 to 15.1) and 19.4 (95% CI, not estimated) months, respectively. Grade 3 or 4 neutropenia was reported in 28 patients (32.1%) and thrombocytopenia was observed in 23 patients (26.6%). CONCLUSION: This phase II study showed that SOX regimen is a reasonable option in metastatic breast cancer previously treated with anthracyclines and taxanes.

APOE ε4-dependent effects on the early amyloid pathology in induced neurons of patients with Alzheimer's disease.

BACKGROUND: The ε4 allele of apolipoprotein E (APOE ε4) is the strongest known genetic risk factor for late-onset Alzheimer's disease (AD), associated with amyloid pathogenesis. However, it is not clear how APOE ε4 accelerates amyloid-beta (Aß) deposition during the seeding stage of amyloid development in AD patient neurons. METHODS: AD patient induced neurons (iNs) with an APOE ε4 inducible system were prepared from skin fibroblasts of AD patients. Transcriptome analysis was performed using RNA isolated from the AD patient iNs expressing APOE ε4 at amyloid-seeding and amyloid-aggregation stages. Knockdown of IGFBP3 was applied in the iNs to investigate the role of IGFBP3 in the APOE ε4-mediated amyloidosis. RESULTS: We optimized amyloid seeding stage in the iNs of AD patients that transiently expressed APOE ε4. Remarkably, we demonstrated that Aß  pathology was aggravated by the induction of APOE ε4 gene expression at the amyloid early-seeding stage in the iNs of AD patients. Moreover, transcriptome analysis in the early-seeding stage revealed that IGFBP3 was functionally important in the molecular pathology of APOE ε4-associated AD. CONCLUSIONS: Our findings suggest that the presence of APOE ε4 at the early Aß-seeding stage in patient iNs is critical for aggravation of sporadic AD pathology. These results provide insights into the importance of APOE ε4 expression for the progression and pathogenesis of sporadic AD.

Seipin transmembrane segments critically function in triglyceride nucleation and lipid droplet budding from the membrane.

Lipid droplets (LDs) are organelles formed in the endoplasmic reticulum (ER) to store triacylglycerol (TG) and sterol esters. The ER protein seipin is key for LD biogenesis. Seipin forms a cage-like structure, with each seipin monomer containing a conserved hydrophobic helix and two transmembrane (TM) segments. How the different parts of seipin function in TG nucleation and LD budding is poorly understood. Here, we utilized molecular dynamics simulations of human seipin, along with cell-based experiments, to study seipin's functions in protein-lipid interactions, lipid diffusion, and LD maturation. An all-atom simulation indicates that seipin TM segment residues and hydrophobic helices residues located in the phospholipid tail region of the bilayer attract TG. Simulating larger, growing LDs with coarse-grained models, we find that the seipin TM segments form a constricted neck structure to facilitate conversion of a flat oil lens into a budding LD. Using cell experiments and simulations, we also show that conserved, positively charged residues at the end of seipin's TM segments affect LD maturation. We propose a model in which seipin TM segments critically function in TG nucleation and LD growth.

Dormant state of quiescent neural stem cells links Shank3 mutation to autism development.

Autism spectrum disorders (ASDs) are common neurodevelopmental disorders characterized by deficits in social interactions and communication, restricted interests, and repetitive behaviors. Despite extensive study, the molecular targets that control ASD development remain largely unclear. Here, we report that the dormancy of quiescent neural stem cells (qNSCs) is a therapeutic target for controlling the development of ASD phenotypes driven by Shank3 deficiency. Using single-cell RNA sequencing (scRNA-seq) and transposase accessible chromatin profiling (ATAC-seq), we find that abnormal epigenetic features including H3K4me3 accumulation due to up-regulation of Kmt2a levels lead to increased dormancy of qNSCs in the absence of Shank3. This result in decreased active neurogenesis in the Shank3 deficient mouse brain. Remarkably, pharmacological and molecular inhibition of qNSC dormancy restored adult neurogenesis and ameliorated the social deficits observed in Shank3-deficient mice. Moreover, we confirmed restored human qNSC activity rescues abnormal neurogenesis and autism-like phenotypes in SHANK3-targeted human NSCs. Taken together, our results offer a novel strategy to control qNSC activity as a potential therapeutic target for the development of autism.

Bifidobacterium bifidum BGN4 and Bifidobacterium longum BORI promotes neuronal rejuvenation in aged mice.

An increasing number of studies have indicated that alterations in gut microbiota affect brain function, including cognition and memory ability, via the gut-brain axis. In this study, we aimed to determine the protective effect of Bifidobacterium bifidum BGN4 (B. bifidum BGN4) and Bifidobacterium longum BORI (B. longum BORI) on age-related brain damage in mice. We found that administration of B. bifidum BGN4 and B. longum BORI effectively elevates brain-derived neurotrophic factor expression which was mediated by increased histone 3 lysine 9 trimethylation. Furthermore, administration of probiotic supplementation reversed the DNA damage and apoptotic response in aged mice and also improved the age-related cognitive and memory deficits of these mice. Taken together, the present study highlights the anti-aging effects of B. bifidum BGN4 and B. longum BORI in the aged brain and their beneficial effects for age-related brain disorders.

Computational Studies of Lipid Droplets.

Lipid droplets (LDs) are intracellular organelles whose primary function is energy storage. Known to emerge from the endoplasmic reticulum (ER) bilayer, LDs have a unique structure with a core consisting of neutral lipids, triacylglycerol (TG) or sterol esters (SE), surrounded by a phospholipid (PL) monolayer and decorated by proteins that come and go throughout their complex lifecycle. In this Feature Article, we review recent developments in computational studies of LDs, a rapidly growing area of research. We highlight how molecular dynamics (MD) simulations have provided valuable molecular-level insight into LD targeting and LD biogenesis. Additionally, we review the physical properties of TG from different force fields compared with experimental data. Possible future directions and challenges are discussed.

Tracing microplastics from raw water to drinking water treatment plants in Busan, South Korea.

The increasing amount of plastic waste has raised concerns about microplastics (MPs) in aquatic environments. MPs can be fragmented into nanoplastics that can pass through water treatment processes and into tap water; potentially threatening human health because of their high adsorption capacity for hazardous organic materials and their intrinsic toxicity. This case study investigates the identification, fate, and removal efficiency of MPs in Korean drinking water treatment plants. Two sites on the Nakdong River, two lake reservoirs (raw water sources), and four corresponding drinking water treatment plants were targeted to trace the amounts, types, and sizes of MPs throughout the treatment process. Monthly quantitative and qualitative analyses were conducted by chemical image mapping using micro-Fourier-transform infrared spectroscopy. MPs larger than 20 µm were detected, and their sizes and types were quantified using siMPle software. Overall, the number of MPs in the river sites (January to April and October to November) exceeded those in the reservoirs, but only slight differences in the number of MPs between rivers and lake reservoirs were detected from June to October. The annual average number of MPs in River A, B and Lack C and D was not distinctively different (2.65, 2.48, 2.46 and 1.87 particles/L, respectively). The majority of MPs found in raw waters were polyethylene (PE)/polypropylene (PP) (> 60%) and polyethylene terephthalate (PET)/poly(methyl methacrylate) (PMMA) (20%), in addition to polyamide (<10%) in the river and polystyrene (<10%) in the lake reservoirs. Approximately 70-80% of the MPs were removed by pre-ozonation/sedimentation; 81-88% of PE/PP was removed by this process. PET/PMMA was removed by filtration. Correlation of MPs with water quality parameters showed that the Mn concentration was moderately correlated with the MP abundance in rivers and lake reservoirs, excluding the lake with the lowest Mn concentration, while the total organic carbon was negatively correlated with the MP abundance in both rivers (A and B) and lake reservoir C.

Key Factors Governing Initial Stages of Lipid Droplet Formation.

Lipid droplets (LDs) are neutral lipid storage organelles surrounded by a phospholipid (PL) monolayer. LD biogenesis from the endoplasmic reticulum is driven by phase separation of neutral lipids, overcoming surface tension and membrane deformation. However, the core biophysics of the initial steps of LD formation remains relatively poorly understood. Here, we use a tunable, phenomenological coarse-grained model to study triacylglycerol (TG) nucleation in a bilayer membrane. We show that PL rigidity has a strong influence on TG lensing and membrane remodeling: when membrane rigidity increases, TG clusters remain more planar with high anisotropy but a minor degree of phase nucleation. This finding is confirmed by advanced sampling simulations that calculate nucleation free energy as a function of the degree of nucleation and anisotropy. We also show that asymmetric tension, controlled by the number of PL molecules on each membrane leaflet, determines the budding direction. A TG lens buds in the direction of the monolayer containing excess PL molecules to allow for better PL coverage of TG, consistent with the reported experiments. Finally, two governing mechanisms of the LD growth, Ostwald ripening and merging, are observed. Taken together, this study characterizes the interplay between two thermodynamic quantities during the initial LD phases, the TG bulk free energy and membrane remodeling energy.

Incidence and Direct Medical Cost of Adjustment Disorder and in Korea Using National Health Insurance Service Claims Data From 2011 to 2017.

OBJECTIVE: We aimed to investigate the annual incidence and direct medical cost of adjustment disorder in general population using the National Health Insurance Service Database (NHID) in Korea. METHODS: To examine the incidence, we selected patients who had at least one medical claim for adjustment disorder and had not been diagnosed in the previous 365 days, from 2010 to 2017. RESULTS: The number of newly diagnosed cases of adjustment disorder from 2011 to 2017 were total 101,922. Annual incidence of adjustment disorder was ranged from 22.0 to 36.8 per 100,000 persons. The incidence of adjustment disorder was found more in female and highest among 70-79 years of age group and medical aid beneficiaries group. Annual prevalence of adjustment disorder was in the range from 95.4 to 116.4 per 100,000 persons. Estimated annual medical cost per person of adjustment disorder was ranged from 162 to 231.4 US dollars. CONCLUSION: From 2011 to 2017, the annual incidence and direct medical cost of adjustment disorder in Korea were increased. Proper information on adjustment disorder will not only allows us to accumulate more knowledge but also lead to more appropriate therapeutic interventions.

Physical Characterization of Triolein and Implications for Its Role in Lipid Droplet Biogenesis.

Lipid droplets (LDs) are neutral lipid-storing organelles surrounded by a phospholipid (PL) monolayer. At present, how LDs are formed in the endoplasmic reticulum (ER) bilayer is poorly understood. In this study, we present a revised all-atom (AA) triolein (TG) model, the main constituent of the LD core, and characterize its properties in a bilayer membrane to demonstrate the implications of its behavior in LD biogenesis. In bilayer simulations, TG resides at the surface, adopting PL-like conformations (denoted in this work as SURF-TG). Free energy sampling simulation results estimate the barrier for TG relocating from the bilayer surface to the bilayer center to be ∼2 kcal/mol in the absence of an oil lens. SURF-TG is able to modulate membrane properties by increasing PL ordering, decreasing bending modulus, and creating local negative curvature. The other neutral lipid, dioleoyl-glycerol (DAG), also reduces the membrane bending modulus and populates negative curvature regions. A phenomenological coarse-grained (CG) model is also developed to observe larger-scale SURF-TG-mediated membrane deformation. CG simulations confirm that TG nucleates between the bilayer leaflets at a critical concentration when SURF-TG is evenly distributed. However, when one monolayer contains more SURF-TG, the membrane bends toward the other leaflet, followed by TG nucleation if a concentration is higher than the critical threshold. The central conclusion of this study is that SURF-TG is a negative curvature inducer, as well as a membrane modulator. To this end, a model is proposed in which the accumulation of SURF-TG in the luminal leaflet bends the ER bilayer toward the cytosolic side, followed by TG nucleation.

Stressed Lipid Droplets: How Neutral Lipids Relieve Surface Tension and Membrane Expansion Drives Protein Association.

Lipid droplets (LDs) are intracellular storage organelles composed of neutral lipids, such as triacylglycerol (TG), surrounded by a phospholipid (PL) monolayer decorated with specific proteins. Herein, we investigate the mechanism of protein association during LD and bilayer membrane expansion. We find that the neutral lipids play a dynamic role in LD expansion by further intercalating with the PL monolayer to create more surface-oriented TG molecules (SURF-TG). This interplay both reduces high surface tension incurred during LD budding or growth and also creates expansion-specific surface features for protein recognition. We then show that the autoinhibitory (AI) helix of CTP:phosphocholine cytidylyltransferase, a protein known to target expanding monolayers and bilayers, preferentially associates with large packing defects in a sequence-specific manner. Despite the presence of three phenylalanines, the initial binding with bilayers is predominantly mediated by the sole tryptophan due to its preference for membrane interfaces. Subsequent association is dependent on the availability of large, neighboring defects that can accommodate the phenylalanines, which are more probable in the stressed systems. Tryptophan, once fully associated, preferentially interacts with the glycerol moiety of SURF-TG in LDs. The calculation of AI binding free energy, hydrogen bonding and depth analysis, and in silico mutation experiments support the findings. Hence, SURF-TG can both reduce surface tension and mediate protein association, facilitating class II protein recruitment during LD expansion.

Serum Dickkopf-1 in Combined with CA 19-9 as a Biomarker of Intrahepatic Cholangiocarcinoma.

Dickkopf-related protein 1 (DKK-1) has a diagnostic and prognostic value in various malignant tumors. We investigated the diagnostic and prognostic performance of DKK-1 in combination with carbohydrate antigen 19-9 (CA 19-9) in cholangiocarcinoma (CCC) patients. Serum DKK-1 levels were measured using enzyme-linked immunosorbent assay. The receiver operating characteristic (ROC) curve, area under ROC (AUROC) analyses, Kaplan-Meier method, and Cox proportional hazard model were used to evaluate the diagnostic and prognostic performance of DKK-1 in combination with CA 19-9. We checked DKK-1 levels in 356 CCC patients and found that DKK-1 was significantly elevated only in 79 intrahepatic CCC (ICC) patients compared to controls (340.5 vs. 249.8 pg/mL, p = 0.002). The optimal cutoff level of DKK-1 used to identify ICC patients was 258.0 pg/mL (AUROC = 0.637, sensitivity = 59.5%, specificity = 56.9%, positive predictive value (PPV) = 40.5%, negative predictive value (NPV) = 74.0%, positive likelihood ratio (LR) = 1.38, and negative LR = 0.71). Using this cutoff, 47 (59.5%) patients were correctly diagnosed with ICC. DKK-1 in combination with CA 19-9 showed a better diagnostic performance (AUROC = 0.793, sensitivity = 74.7%, specificity = 56.3%, PPV = 45.7, NPV = 81.8, positive LR = 1.71, and negative LR = 0.45) than CA 19-9 alone. The low DKK-1 and CA 19-9 expression group had a significantly longer overall survival (OS) than the high expression group (p = 0.006). The higher level of DKK-1 and CA 19-9 was independently associated with shorter OS (hazard ratio = 3.077, 95% confidence interval 1.389-6.819, p = 0.006). The diagnostic and prognostic performance of DKK-1 in combination with CA 19-9 might be better than those of CA 19-9 alone in ICC patients.

The Surface and Hydration Properties of Lipid Droplets.

Lipid droplets (LDs) are energy storage organelles composed of neutral lipids, such as triacylglycerol (TG) and sterol esters, surrounded by a phospholipid (PL) monolayer. Their central role in metabolism, complex life cycle, and unique lipid monolayer surface have garnered great attention over the last decade. In this article, results from the largest and longest all-atom simulations to date suggest that 5-8% of the LD surface is occupied by TG molecules, a number that exceeds the maximal solubility reported for TGs in PL bilayers (2.8%). Two distinct classes of TG molecules that interact with the LD monolayer are found. Those at the monolayer surface (SURF-TG) are ordered like PLs with the glycerol moiety exposed to water, creating a significant amount of chemically unique packing defects, and the acyl chains extended toward the LD center. In contrast, the TGs that intercalate just into the PL tail region (CORE-TG) are disordered and increase the amount of PL packing defects and the PL tail order. The degree of interdigitation caused by CORE-TG is stable and determines the width of the TG-PL overlap, whereas that caused by SURF-TG fluctuates and is highly correlated with the area per PL or the expansion of the monolayer. Thus, when the supply of PLs is limited, SURF-TG may reduce surface tension by behaving as a secondary membrane component. The hydration properties of the simulated LD systems demonstrate ∼10 times more water in the LD core than previously reported. Collectively, the reported surface and hydration properties are expected to play a direct role in the mechanisms by which proteins target and interact with LDs.

Epitranscriptomic N6-Methyladenosine Modification Is Required for Direct Lineage Reprogramming into Neurons.

N6-methyladenosine (m6A), a conserved epitranscriptomic modification of eukaryotic mRNA (mRNA), plays a critical role in a variety of biological processes. Here, we report that m6A modification plays a key role in governing direct lineage reprogramming into induced neuronal cells (iNs). We found that m6A modification is required for the remodeling of specific mRNAs required for the neuronal direct conversion. Inhibition of m6A methylation by Mettl3 knockdown decreased the efficiency of direct lineage reprogramming, whereas increased m6A methylation by Mettl3 overexpression increased the efficiency of iN generation. Moreover, we found that transcription factor Btg2 is a functional target of m6A modification for efficient iN generation. Taken together, our results suggest the importance of establishing epitranscriptomic remodeling for the cell fate conversion into iNs.

Methionine Enkephalin Suppresses Osteocyte Apoptosis Induced by Compressive Force through Regulation of Nuclear Translocation of NFATc1.

Mechanical stress stimulates bone remodeling, which occurs through bone formation and resorption, resulting in bone adaptation in response to the mechanical stress. Osteocytes perceive mechanical stress loaded to bones and promote bone remodeling through various cellular processes. Osteocyte apoptosis is considered a cellular process to induce bone resorption during mechanical stress-induced bone remodeling, but the underlying molecular mechanisms are not fully understood. Recent studies have demonstrated that neuropeptides play crucial roles in bone metabolism. The neuropeptide, methionine enkephalin (MENK) regulates apoptosis positively and negatively depending on cell type, but the role of MENK in osteocyte apoptosis, followed by bone resorption, in response to mechanical stress is still unknown. Here, we examined the roles and mechanisms of MENK in osteocyte apoptosis induced by compressive force. We loaded compressive force to mouse parietal bones, resulting in a reduction of MENK expression in osteocytes. A neutralizing connective tissue growth factor (CTGF) antibody inhibited the compressive force-induced reduction of MENK. An increase in osteocyte apoptosis in the compressive force-loaded parietal bones was inhibited by MENK administration. Nuclear translocation of NFATc1 in osteocytes in the parietal bones was enhanced by compressive force. INCA-6, which inhibits NFAT translocation into nuclei, suppressed the increase in osteocyte apoptosis in the compressive force-loaded parietal bones. NFATc1-overexpressing MLO-Y4 cells showed increased expression of apoptosis-related genes. MENK administration reduced the nuclear translocation of NFATc1 in osteocytes in the compressive force-loaded parietal bones. Moreover, MENK suppressed Ca2+ influx and calcineurin and calmodulin expression, which are known to induce the nuclear translocation of NFAT in MLO-Y4 cells. In summary, this study shows that osteocytes expressed MENK, whereas the MENK expression was suppressed by compressive force via CTGF signaling. MENK downregulated nuclear translocation of NFATc1 probably by suppressing Ca2+ signaling in osteocytes and consequently inhibiting compressive force-induced osteocyte apoptosis, followed by bone resorption. © 2020 The Authors. JBMR Plus published by Wiley Periodicals, Inc. on behalf of American Society for Bone and Mineral Research.