Neutral lysophosphatidylcholine mediates α-synuclein-induced synaptic vesicle clustering.

α-synuclein (α-Syn) is a presynaptic protein that is involved in Parkinson's and other neurodegenerative diseases and binds to negatively charged phospholipids. Previously, we reported that α-Syn clusters synthetic proteoliposomes that mimic synaptic vesicles. This vesicle-clustering activity depends on a specific interaction of α-Syn with anionic phospholipids. Here, we report that α-Syn surprisingly also interacts with the neutral phospholipid lysophosphatidylcholine (lysoPC). Even in the absence of anionic lipids, lysoPC facilitates α-Syn-induced vesicle clustering but has no effect on Ca2+-triggered fusion in a single vesicle-vesicle fusion assay. The A30P mutant of α-Syn that causes familial Parkinson disease has a reduced affinity to lysoPC and does not induce vesicle clustering. Taken together, the α-Syn-lysoPC interaction may play a role in α-Syn function.

Probing the dynamic crosstalk of lysosomes and mitochondria with structured illumination microscopy.

Structured illumination microscopy (SIM) is a super-resolution technology for imaging living cells and has been used for studying the dynamics of lysosomes and mitochondria. Recently, new probes and analyzing methods have been developed for SIM imaging, enabling the quantitative analysis of these subcellular structures and their interactions. This review provides an overview of the working principle and advances of SIM, as well as the organelle-targeting principles and types of fluorescence probes, including small molecules, metal complexes, nanoparticles, and fluorescent proteins. Additionally, quantitative methods based on organelle morphology and distribution are outlined. Finally, the review provides an outlook on the current challenges and future directions for improving the combination of SIM imaging and image analysis to further advance the study of organelles. We hope that this review will be useful for researchers working in the field of organelle research and help to facilitate the development of SIM imaging and analysis techniques.

lpla (lipoprotein lipase a) is a marker of early adipogenesis rather than late adipogenesis in grass carp (Ctenopharyngodon idellus).

Lipoprotein lipase (LPL) functions as a marker of adipocyte differentiation in mammals, but little is known about its role in fish adipogenesis. The aim of this research is to investigate the function of Lpl in adipocyte differentiation in fish. In this paper, we isolated and characterized lipoprotein lipase a (lpla) and lipoprotein lipase b (lplb) from grass carp (Ctenopharyngodon idellus). The complete coding sequence of lpla and lplb was 1524 bp and 1503 bp in length, coding for 507 amino acids and 500 amino acids, respectively. Both lpla and lplb mRNA were expressed in a great number of tissues. During adipogenesis, the level of lpla mRNA reached its maximum at day 2 and then dropped gradually, while the level of lplb mRNA had no significant changes, indicating that lpla and lplb may have different function in the differentiation of grass carp adipocyte. Furthermore, inhibition of lpla by inhibitor of LPL(GSK264220A) at early time points most clearly reduced adipogenesis, whereas these effects were less pronounced at later stages, suggesting that lpla predominantly affects early adipogenesis rather than late adipogenesis. Based on these findings, it can be inferred that lpla and lplb in grass carp may have distinct roles in the differentiation of grass carp adipocyte, and lpla may play an important role in the early adipogenesis rather than late adipogenesis in grass carp.

Multiple-Color Platinum Complex with Super-Large Stokes Shift for Super-Resolution Imaging of Autolysosome Escape.

It is of great significance to track the platinum drugs in real time with super-resolution to elucidate their mechanism of action, such as their behavior and distribution in live cells. Such information is required for further drug development. However, it is always challenging to design platinum complexes suitable for such research. Herein, we design a luminescent building block (L) for metal complexes and a dinuclear platinum complex (Pt2 L) for super-resolution imaging. Because of its super-large Stokes shift and excellent photophysical properties, Pt2 L is capable of serving as an ideal candidate for super-resolution imaging with extremely low luminescence background and high photobleaching resistance. Moreover, upon light stimulation, a matter flux of Pt2 L escaping from autolysosomes to nucleus was observed, which represents a new transportation path. Utilizing the photoactivated escape properties, we can regulate the nuclear accessibility of Pt2 L form autolysosomes with photo-selectivity, which provides a new way to improve the targeting of platinum drugs.

Quantitation and mapping of the epigenetic marker 5-hydroxymethylcytosine.

We here review primary methods used in quantifying and mapping 5-hydroxymethylcytosine (5hmC), including global quantification, restriction enzyme-based detection, and methods involving DNA-enrichment strategies and the genome-wide sequencing of 5hmC. As discovered in the mammalian genome in 2009, 5hmC, oxidized from 5-methylcytosine (5mC) by ten-eleven translocation (TET) dioxygenases, is increasingly being recognized as a biomarker in biological processes from development to pathogenesis, as its various detection methods have shown. We focus in particular on an ultrasensitive single-molecule imaging technique that can detect and quantify 5hmC from trace samples and thus offer information regarding the distance-based relationship between 5hmC and 5mC when used in combination with fluorescence resonance energy transfer.

Role of glycans in cholesteryl ester transfer protein revealed by molecular dynamics simulation.

Current cholesteryl ester transfer protein (CETP) inhibitors are designed based on the unglycosylated crystal structure, and most of them have failed to cure cardiovascular disease (CVD). It is particularly important for us to investigate the glycosylation structure of CETP (CETP-G) and effect of glycans on the structure and function of CETP. Here, we used a total of 3.0-µs molecular dynamics (MD) trajectories of nascent structure of CETP (CETP-N) and CETP-G to study their structural differentiations, to shed new light on the CETP-mediated lipid exchange. In accordance with our simulations and previous mutation studies, relative to CETP-N, CETP-G adopts a more stretched shape with higher hydrophobic and hydrophilic solvent-accessible surface area (SASA) of N-terminal oscillating with larger amplitude, in which Glycan88 provides partial assistance for CEs through the N-terminal. Glycan341 reduces the flexibility of neck flap, with the interference of CEs through the neck region. Besides, Glycan240 reduces the flexibility of Helix-X to interfere the CEs transfer. Glycan396 decreases the flexibility and increases the hydrophobic SASA of C-terminal. Overall, these glycans affect the dynamics and structure of CETP through forming H-bonds with surrounding residues, and the sampled conformations of glycan is also affected by its surrounding residues. Thus, glycans are an integral part of CETP, further studies on the CETP inhibition and treatment of CVD should fully consider the effect of glycans.

Super-Resolution Tracking of Mitochondrial Dynamics with An Iridium(III) Luminophore.

Combining luminescent transition metal complex with super-resolution microscopy is an excellent strategy for the long-term visualization of the dynamics of subcellular structures in living cells. However, it remains unclear whether iridium(III) complexes are applicable for a particular type of super-resolution technique, structured illumination microscopy (SIM), to image subcellular structures. Herein, an iridium(III) dye, to track mitochondrial dynamics in living cells under SIM is described. The dye demonstrates excellent specificity and photostability and satisfactory cell permeability. While using SIM to image mitochondria, an ≈80 nm resolution is achieved that allows the clear observation of the structure of mitochondrial cristae. The dye is used to monitor and quantify mitochondrial dynamics relative to lysosomes, including fusion involved in mitophagy, and newly discovered mitochondria-lysosome contact (MLC) under different conditions. The MLC remains intact and fusion vanishes when five receptors, p62, NDP52, OPTN, NBR1, and TAX1BP1, are knocked out, suggesting that these two processes are independent.

Structure-property relationships of cell clusters in biotissues: 2D analysis.

Gaining insight into the relationships between the self-organized cell structures and the properties of biotissues is helpful for revealing the function of biomaterials and its designing principle. However, the traditionally used random foam model neglects several important details of the frameworks of cell clusters resulting in incomplete conclusions. Herein, we use a more complete model, the cell adhesion model, to investigate the mechanical and morphological properties of the two-dimensional (2D) dry cell foams. Since these 2D structures are formed by cell adhesion, the system can reach equilibrium through minimizing free energy. Under the equilibrium conditions without volume constraint, shape equations for highly symmetrical structures are derived, and the analytical results of the corresponding mechanical parameters, such as the Young's modulus, bulk modulus and failure strength, are obtained. Moreover, with volume constraint, numerical simulation method is applied to study the complex shapes and obtain several stable multicellular structures. Symmetry breaking caused by the volume change is also observed. Moreover, typical periodic shapes and the corresponding phase transformations are also explored. Our study provides a new potential method to bridge the microstructure and macro-mechanical parameters of biotissues. The results are also useful for understanding the formation mechanism of biotissue structures.

A matter of timing.

A change in the electric charge of autophagosome membranes controls the recruitment of SNARE proteins to ensure that membrane fusion occurs at the right time during autophagy.

Prevalence of overweight/obesity, and associated factors among adolescents aged 12 ∼ 15 in Shandong Province, China: A cross-sectional study.

Overweight/obesity among adolescents in Shandong Province, China, has been rising, posing significant public health challenge. Comprehensive investigation is needed to develop effective interventions. Following the Strengthening the Reporting of Observational Studies in Epidemiology guidelines, a stratified random cluster sampling approach was used from September to October 2023 across 17 cities in Shandong Province. The study included 165 middle schools, surveying 99,638 students aged 12 ∼ 15. After applying exclusion criteria, 97,356 students (97.71% effective rate) completed anthropometric measurements and questionnaires. Overweight/obesity were assessed based on national and international standards. Univariable chi-square test and multivariable logistic regression were used to analyze factors influencing overweight/obesity. In 2023, the overweight/obesity rate among 12 ∼ 15-year-olds in Shandong was 19.75%. Significant factors included sex, age, residence, family income, parental weight status and activity, mother's gestational diabetes history, birth weight, physical activity, sleep, screen time, homework, and diet. Girls, older adolescents, and those with physically active parents or who themselves engaged in over 1.5 h of daily physical activity had lower odds of being overweight/obese. Adequate sleep and frequent consumption of vegetable and fruit were also protective. Higher odds were associated with urban residence, high family income, overweight/obese parents, maternal gestational diabetes, high birth weight, excessive screen time, extensive homework, and frequent fast food consumption. Overweight/obesity in Shandong adolescents is influenced by multiple determinants. Holistic interventions addressing genetic, behavioral, and environmental factors are essential for promoting healthier lifestyles and reducing the prevalence in this demographic.

Prevalence and associated factors of myopia among adolescents aged 12-15 in Shandong Province, China: a cross-sectional study.

Myopia is a growing concern worldwide, especially among adolescents. This study aimed to investigate the prevalence and associated factors of myopia in adolescents aged 12-15 in Shandong Province, China. This cross-sectional study adhered to the Strengthening the Reporting of Observational Studies in Epidemiology guidelines and involved stratified random cluster sampling of 128,678 students from 186 middle schools across 17 cities in Shandong Province. Data collection was conducted from March to April 2024. We excluded students with organic eye diseases, abnormal binocular vision functions, or a history of eye injuries or surgeries. Myopia was assessed using the standard logarithmic visual acuity chart and autorefractor without inducing ciliary muscle paralysis. A comprehensive questionnaire survey was conducted to gather demographic characteristics and daily life behaviors. With the chi-squared test for univariable analysis and multivariable logistic regression for identifying significant factors. This study included 126,375 participants, with a gender distribution of 51.02% male and 48.98% female. The overall prevalence of myopia was 71.34%. Higher prevalence was observed in girls (72.26%) compared to boys (70.45%), and the prevalence increased with age, peaking at 73.12% in 15-year-olds. Urban residents had a higher prevalence (71.86%) than rural (70.39%). Factors such as increased frequency of eye usage while lying down or leaning forward, frequent use of eyes while walking or riding in a car, prolonged screen time, and extended homework duration were associated with higher odds of developing myopia. Conversely, higher frequency of outdoor exercise, maintaining proper posture during reading and writing, greater distance from eyes to screen, and longer sleep duration were associated with lower odds. Additionally, female gender, older age, urban residence, and parental history of myopia increased the risk. The high prevalence of myopia among adolescents in Shandong Province was influenced by a combination of demographic, behavioral, and environmental factors. The study highlighted the importance of lifestyle modifications, such as increasing outdoor activities and maintaining proper visual habits, limiting the duration of screen exposure and homework sessions, and extending sleep duration, to mitigate the risk of developing myopia. These findings underscored the need for targeted public health interventions and educational campaigns to address this significant public health issue.

Comparative study on the transcriptional activities of grass carp (Ctenopharyngodon idellus) peroxisome proliferator-activated receptors induced by different fatty acids.

Peroxisome proliferator-activated receptors (PPARs) are ligand-activated transcription factors that are part of the nuclear hormone receptor family, playing a crucial role in gene expression regulation. They serve as a connection between lipid metabolism disorders and innate immunity by being activated by fatty acids and their derivatives, facilitating signal transduction between the cell surface and nucleus. However, the specific transcriptional effects of different fatty acids (FAs) in fish are not yet fully understood. In our research, we identified and characterized PPARs in grass carp (Ctenopharyngodon idellus). The complete coding sequences of pparαa, pparαb, pparγ, pparδa, and pparδb were 1443 bp, 1404 bp, 1569 bp, 1551 bp, and 1560 bp in length, respectively. Pparα showed the highest expression in the liver, pparγ was mainly expressed in abdominal adipose tissue, and pparδ exhibited increased expression in the heart compared to other tissues. Gene localization analysis revealed that only pparδa was present in both the nucleus and cytoplasm, while the other four genes were exclusively located in the nucleus. Furthermore, our study explored the influence of various fatty acids (docosahexaenoic acid, palmitic acid, lauric acid and oleic acid at concentrations of 0, 50, 100, and 200 µM) on the transcriptional activities of different PPARs, demonstrating the diverse effects of fatty acid ligands on PPAR transcriptional activity. These results have significant implications for understanding the regulation of PPARs transcriptional activity.

Comparative analysis of Scarb1 and Cd36 in grass carp (Ctenopharyngodon idellus): Implications for DHA uptake.

The polyunsaturated fatty acid docosahexaenoic acid (DHA) significantly influences fish growth and lipid metabolism. Nevertheless, the specific mechanism by which DHA is transported and exerts its effects remains unclear. Scavenger receptor class B type I (SCARB1) is essential for maintaining cellular cholesterol levels and regulating the immune system in mammals, as well as facilitating the uptake of fatty acids (FAs). Another class B scavenger receptor, cluster-determinant 36 (CD36), is involved in promoting the uptake and transport of long-chain fatty acids. However, the molecular characteristics of the grass carp scarb1 gene have not yet been reported, and the potential role of Scarb1 and Cd36 in mediating DHA transport and metabolism remains uncertain. This study aimed to investigate the effects of Scarb1 and Cd36 on DHA transport. Initially, grass carp scarb1-1 and scarb1-2 were cloned. Predictions were made regarding their structural characteristics, including number and presence of transmembrane domains and glycosylation sites. Furthermore, gene structure analysis revealed that scarb1-1 has two additional exons in the 3'-region compared to scarb1-2. The multiple sequence alignment indicated that Scarb1 exhibits conserved motifs and amino acid residues across vertebrates. mRNA expression of scarb1-1 was the highest in the intestine, while scarb1-2 was highest expressed in adipose tissue, with both having lower expression levels in muscle tissue. Scarb1-1 was primarily localized on the cell membrane, whereas Scarb1-2 was found in both the cell membrane and cytoplasm. After overexpression of grass carp Scarb1-1, Scarb1-2, and Cd36 in HEK 293 T cells, DHA incubation showed that only Cd36 significantly increased cellular DHA relative content, suggesting a potential role of Cd36 in DHA transport. These findings will serve as a basis for further research on fatty acid transport in fish.

VAMP2 chaperones α-synuclein in synaptic vesicle co-condensates.

α-Synuclein (α-Syn) aggregation is closely associated with Parkinson's disease neuropathology. Physiologically, α-Syn promotes synaptic vesicle (SV) clustering and soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complex assembly. However, the underlying structural and molecular mechanisms are uncertain and it is not known whether this function affects the pathological aggregation of α-Syn. Here we show that the juxtamembrane region of vesicle-associated membrane protein 2 (VAMP2)-a component of the SNARE complex that resides on SVs-directly interacts with the carboxy-terminal region of α-Syn through charged residues to regulate α-Syn's function in clustering SVs and promoting SNARE complex assembly by inducing a multi-component condensed phase of SVs, α-Syn and other components. Moreover, VAMP2 binding protects α-Syn against forming aggregation-prone oligomers and fibrils in these condensates. Our results suggest a molecular mechanism that maintains α-Syn's function and prevents its pathological amyloid aggregation, the failure of which may lead to Parkinson's disease.

Double-Transmembrane Domain of SNAREs Decelerates the Fusion by Increasing the Protein-Lipid Mismatch.

SNARE is the essential mediator of membrane fusion that highly relies on the molecular structure of SNAREs. For instance, the protein syntaxin-1 involved in neuronal SNAREs, has a single transmembrane domain (sTMD) leading to fast fusion, while the syntaxin 17 has a V-shape double TMDs (dTMDs), taking part in the autophagosome maturation. However, it is not clear how the TMD structure influences the fusion process. Here, we demonstrate that the dTMDs significantly reduce fusion rate compared with the sTMD by using an in vitro reconstitution system. Through theoretical analysis, we reveal that the V-shape dTMDs can significantly increase protein-lipid mismatch, thereby raising the energy barrier of the fusion, and that increasing the number of SNAREs can reduce the energy barrier or protein-lipid mismatch. This study provides a physicochemical mechanistic understanding of SNARE-regulated membrane fusion.

2D MoS2 and BN Nanosheets Damage Mitochondria through Membrane Penetration.

With the progression of nanotechnology, a growing number of nanomaterials have been created and incorporated into organisms and ecosystems, which raises significant concern about potential hazards of these materials on human health, wildlife, and the environment. Two-dimensional (2D) nanomaterials are one type of nanomaterials with thicknesses ranging from that of a single atom or of several atoms and have been proposed for a variety of biomedical applications such as drug delivery and gene therapy, but the toxicity thereof on subcellular organelles remains to be studied. In this work, we studied the impact of two typical 2D nanomaterials, MoS2 and BN nanosheets, on mitochondria, which are a type of membranous subcellular organelle that provides energy to cells. While 2D nanomaterials at a low dose exhibited a negligible cell mortality rate, significant mitochondrial fragmentation and partially reduced mitochondrial functions occurred; cells initiate mitophagy in response to mitochondrial damages, which cleans damaged mitochondria to avoid damage accumulation. Moreover, the molecular dynamics simulation results revealed that both MoS2 and BN nanosheets can spontaneously penetrate the mitochondrial lipid membrane through the hydrophobic interaction. The membrane penetration induced heterogeneous lipid packing resulting in damages. Our results demonstrate that even at a low dose 2D nanomaterials can physically damage mitochondria by penetrating the membrane, which draws attention to carefully evaluating the cytotoxicity of 2D nanomaterials for the potential biomedical application.

Advanced imaging techniques for tracking drug dynamics at the subcellular level.

Optical microscopes are an important imaging tool that have effectively advanced the development of modern biomedicine. In recent years, super-resolution microscopy (SRM) has become one of the most popular techniques in the life sciences, especially in the field of living cell imaging. SRM has been used to solve many problems in basic biological research and has great potential in clinical application. In particular, the use of SRM to study drug delivery and kinetics at the subcellular level enables researchers to better study drugs' mechanisms of action and to assess the efficacy of their targets in vivo. The purpose of this paper is to review the recent advances in SRM and to highlight some of its applications in assessing subcellular drug dynamics.

An ER-targeted "reserve-release" fluorogen for topological quantification of reticulophagy.

The endoplasmic reticulum's (ER) dynamic nature, essential for maintaining cellular homeostasis, can be influenced by stress-induced damage, which can be assessed by examining the morphology of ER dynamics and, more locally, ER properties such as hydrophobicity, viscosity, and polarity. Although numerous ER-specific chemical probes have been developed to monitor the ER's physical and chemical parameters, the quantitative detection and super-resolution imaging of its local hydrophobicity have yet to be explored. Here, we describe a photostable ER-targeted probe with high signal-to-noise ratio for super-resolution imaging that can specifically respond to changes in ER hydrophobicity under stress based on a "reserve-release" mechanism. The probe shows an excellent ability to target ER over commercial ER dyes and can be used to track local changes of hydrophobicity by fluorescence intensity and morphology during the selective autophagy of ER (i.e., reticulophagy). By correlating the level and location of ER damage with the distribution of fluorescence intensity, we were able to assess reticulophagy at the subcellular level. Beyond that, we developed a topological analytical tool adaptable to any ER probe for detecting structural changes in ER and thus quantitatively identifying reticulophagy. The algorithm-assisted tool can also be adapted to a wide range of molecular probes and organelles. Altogether, the new probe and analytical strategy described here show promise for the quantitative detection and analysis of subtle ER damage and stress.