Clinical and genetic analysis of familial neuromyelitis optica spectrum disorder in Chinese: associated with ubiquitin-specific peptidase USP18 gene variants.

BACKGROUND: Familial clustering of neuromyelitis optica spectrum disorder (NMOSD) was present in Chinese. This study was to investigate the clinical characteristics and genetic background of familial NMOSD. METHODS: Through questionnaires in four medical centres in 2016-2020, we identified 10 families with NMOSD aggregation. The statistical differences of clinical characteristics between familial and sporadic NMOSD (22 cases and 459 cases) were summarised. The whole-exome sequencing (WES) for seven families (13 cases and 13 controls) was analysed, compared with our previous WES data for sporadic NMOSD (228 cases and 1 400 controls). The family-based and population-based association and linkage analysis were conducted to identify the pathogenetic genes, the variant impacts were predicted. RESULTS: The familial occurrence was 0.87% in Chinese. Familial patients had higher expanded disability status scale score than sporadic patients (p=0.03). The single-nucleotide polymorphism (SNP) rs2252257 in the promoter and enhancer of ubiquitin-specific peptidase USP18 was linked to familial NMOSD (p=7.8E-05, logarithm of the odds (LOD)=3.1), SNPs rs361553, rs2252257 and rs5746523 were related to sporadic NMOSD (p=1.29E-10, 3.45E-07 and 2.01E-09, respectively). Patients with the SNP rs361553 T/T genotype had higher recurrence rate than C/T or C/C genotype (1.22±0.85 vs 0.69±0.57 and 0.81±0.65, p=0.003 and 0.001, respectively). SNPs rs361553 and rs2252257 altered USP18 expression in brain and nerve tissues. CONCLUSION: Most clinical characteristics of familial NMOSD were indistinguishable from sporadic NMOSD except for the worst episodes severity. USP18 with impaired intronic regulatory function contributed to the pathogenesis of NMOSD.

Osmotic Concentration-Controlled Particle Uptake and Wrapping-Induced Lysis of Cells and Vesicles.

In vivo, high protein and ion concentrations determine the preferred volumes of cells, organelles, and vesicles. Deformations of their lipid-bilayer membranes by nanoparticle wrapping reduce the interior volumes available to solutes and thus induce large osmotic pressure differences. Osmotic concentration can therefore be an important control parameter for wrapping of nanoparticles. We employ a curvature-elasticity model of the membrane and contact interaction with spherical particles to study their wrapping at initially spherical vesicles. Although the continuous particle-binding transition is independent of the presence of solutes, the discontinuous envelopment transition shifts to higher adhesion strengths and the corresponding energy barrier increases with increasing osmotic concentration. High osmotic concentrations stabilize partial-wrapped, membrane-bound states for both, particle attachment to the inside and the outside. In this regime, wrapping of particles controls membrane tension, with power-law dependencies on osmotic concentration and adhesion strength. For high adhesion strengths, particle wrapping can lead to the opening of mechanosensitive channels in cell membranes and to lysis. Membrane tension-induced stabilization of partial-wrapped states as well as wrapping-induced lysis play important roles not only for desired mechano-bacteriocidal effects of engineered nanomaterials but may also determine viral burst sizes of bacteria and control endocytosis for mammalian cells.

Cold-induced FOXO1 nuclear transport aids cold survival and tissue storage.

Cold-induced injuries severely limit opportunities and outcomes of hypothermic therapies and organ preservation, calling for better understanding of cold adaptation. Here, by surveying cold-altered chromatin accessibility and integrated CUT&Tag/RNA-seq analyses in human stem cells, we reveal forkhead box O1 (FOXO1) as a key transcription factor for autonomous cold adaptation. Accordingly, we find a nonconventional, temperature-sensitive FOXO1 transport mechanism involving the nuclear pore complex protein RANBP2, SUMO-modification of transporter proteins Importin-7 and Exportin-1, and a SUMO-interacting motif on FOXO1. Our conclusions are supported by cold survival experiments with human cell models and zebrafish larvae. Promoting FOXO1 nuclear entry by the Exportin-1 inhibitor KPT-330 enhances cold tolerance in pre-diabetic obese mice, and greatly prolongs the shelf-life of human and mouse pancreatic tissues and islets. Transplantation of mouse islets cold-stored for 14 days reestablishes normoglycemia in diabetic mice. Our findings uncover a regulatory network and potential therapeutic targets to boost spontaneous cold adaptation.

Kink turn sRNA folding upon L7Ae binding using molecular dynamics simulations.

The kink-turn sRNA motif in archaea, whose combination with protein L7Ae initializes the assembly of small ribonucleoprotein particles (sRNPs), plays a key role in ribosome maturation and the translation process. Although many studies have been reported on this motif, the mechanism of sRNA folding coupled with protein binding is still poorly understood. Here, room and high temperature molecular dynamics (MD) simulations were performed on the complex of 25-nt kink-turn sRNA and L7Ae. The average RMSD values between the bound and corresponding apo structures and Kolmogorov-Smirnov P test analysis indicate that sRNA may follow an induced fit mechanism upon binding with L7Ae, both locally and globally. These conclusions are further supported by high-temperature unfolding kinetic analysis. Principal component analysis (PCA) found both closing and opening motions of the kink-turn sRNA. This might play a key role in the sRNP assembly and methylation catalysis. These combined computational methods can be used to study the specific recognition of other sRNAs and proteins.

Molecular dynamics simulations of amyloid fibrils: an in silico approach.

Amyloid fibrils play causal roles in the pathogenesis of amyloid-related degenerative diseases such as Alzheimer's disease, type II diabetes mellitus, and the prion-related transmissible spongiform encephalopathies. The mechanism of fibril formation and protein aggregation is still hotly debated and remains an important open question in order to develop therapeutic method of these diseases. However, traditional molecular biological and crystallographic experiments could hardly observe atomic details and aggregation process. Molecular dynamics (MD) simulations could provide explanations for experimental results and detailed pathway of protein aggregation. In this review, we focus on the applications of MD simulations on several amyloidogenic protein systems. Furthermore, MD simulations could help us to understand the mechanism of amyloid aggregation and how to design the inhibitors.

Nanoparticle wrapping at small non-spherical vesicles: curvatures at play.

Nanoparticles in biological systems encounter lipid-bilayer membranes as barriers. They interact with plasma membranes, membranous organelles, such as the endoplasmic reticulum and the Golgi apparatus, the nucleus, and intracellular and extracellular vesicles, such as autophagosomes, lysosomes, and exosomes. Extracellular vesicles have recently attracted particular attention, as they are involved in the transmission of biological signals and as regulators for biological processes. For example, exosomes, small vesicles containing proteins, mRNA, and miRNA, that are released by cells into the extracellular environment, have been suggested to participate in tumor metastasis. Furthermore, vesicles can be applied as targeted-drug-delivery systems. We systematically characterize wrapping of spherical nanoparticles that enter and exit vesicles, depending on particle size, vesicle size, vesicle reduced volume, and membrane spontaneous curvature. We predict the complex wrapping behavior, in particular for large particle-to-vesicle size ratios, where the shape changes of the free membrane contribute significantly to the deformation energy and where nanoparticle wrapping transitions and vesicle shape transitions are coupled. Partial-wrapped membrane-bound particles impose boundary conditions on the membrane that stabilise oblates and stomatocytes for particle entry, and prolates and stomatocytes for particle exit. Our results suggest that nanoparticles may stimulate autophagocytic engulfment, which would facilitate transport of the nanoparticles into lysosomes and would lead to subsequent degradation of nanoparticle-attached proteins.

Specific recognition mechanism between RNA and the KH3 domain of Nova-2 protein.

The KH3 domain of Nova-2 protein can precisely recognize the sequence-specific target RNA of human glycine receptor α2. However, the recognition mechanism between the protein and its target RNA is still hotly debated. In this study, molecular dynamic simulations in explicit solvent were utilized to understand the recognition mechanism. The structural analysis and the Kolmogorov-Smirnov P-test statistics reveal that the KH3 domain might obey a conformational selection mechanism upon RNA binding. However, the induced fit mechanism could not be completely ruled out. Unfolding kinetics indicates that the folding of RNA and KH3 happens first and then the binding between RNA and KH3 follows. Principle component analysis shows that the invariant Gly-Lys-Gly-Gly loop moves toward to the RNA molecule but the C-terminal domain moves away from the RNA molecule upon binding. These specific dominant motions were hypothesized to stabilize the complex structure. The hydrophobic and hydrogen bonding interactions were found to be the driving forces for the specific recognition, in contrast to the dominant electrostatic interactions for nonspecific recognition.

Global conformational selection and local induced fit for the recognition between intrinsic disordered p53 and CBP.

The transactivation domain (TAD) of tumor suppressor p53 can bind with the nuclear coactivator binding domain (NCBD) of cyclic-AMP response element binding protein (CBP) and activate transcription. NMR experiments demonstrate that both apo-NCBD and TAD are intrinsic disordered and bound NCBD/TAD undergoes a transition to well folded. The recognition mechanism between intrinsic disordered proteins is still hotly debated. Molecular dynamics (MD) simulations in explicit solvent are used to study the recognition mechanism between intrinsic disordered TAD and NCBD. The average RMSD values between bound and corresponding apo states and Kolmogorov-Smirnov P test analysis indicate that TAD and NCBD may follow an induced fit mechanism. Quantitative analysis indicates there is also a global conformational selection. In summary, the recognition of TAD and NCBD might obey a local induced fit and global conformational selection. These conclusions are further supported by high-temperature unbinding kinetics and room temperature landscape analysis. These methods can be used to study the recognition mechanism of other intrinsic disordered proteins.

Conformational selection and induced fit in specific antibody and antigen recognition: SPE7 as a case study.

Antibody-antigen specific recognition is essential in autoimmunity. In this study, antibody SPE7 binding to protein antigens and to hapten molecules were carefully analyzed in order to gain insight into their binding mechanisms. X-ray crystal structures show that SPE7 can adopt at least four different conformations, as in the two observed free isomers (Ab(1) and Ab(2)) and the two observed bound conformers (Ab(3) and Ab(4)). Multidimensional scaling analysis reveals that antibody SPE7 may obey a global conformational selection mechanism upon its binding to an antigen. The conformations of key residue at the binding site (Trp93L) further reveals that bound isomer Ab(3) may come from free isomer Ab(2), and bound isomer Ab(4) from free isomer Ab(1). The average root-mean-square deviation (RMSD) values between the bound isomers and the corresponding free isomers and Kolmogorov-Smimov P test analysis indicate that the antibody may also follow a local induced fit mechanism at the binding interface. Quantitative analysis indicates that the magnitude of the local induced fit interaction at the binding site is more pronounced than that of the global conformational selection interaction. These conclusions are further supported by high-temperature unbinding kinetics analysis. The computational methods proposed here can also be used to study the specific recognitions between other antibody and antigen systems.

Insight into the stability of cross-ß amyloid fibril from VEALYL short peptide with molecular dynamics simulation.

Amyloid fibrils are found in many fatal neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, type II diabetes, and prion disease. The VEALYL short peptide from insulin has been confirmed to aggregate amyloid-like fibrils. However, the aggregation mechanism of amyloid fibril is poorly understood. Here, we utilized molecular dynamics simulation to analyse the stability of VEALYL hexamer. The statistical results indicate that hydrophobic residues play key roles in stabilizing VEALYL hexamer. Single point and two linkage mutants confirmed that Val1, Leu4, and Tyr5 of VEALYL are key residues. The consistency of the results for the VEALYL oligomer suggests that the intermediate states might be trimer (3-0) and pentamer(3-2). These results can help us to obtain an insight into the aggregation mechanism of amyloid fibril. These methods can be used to study the stability of amyloid fibril from other short peptides.