Pharmacology of LRRK2 with type I and II kinase inhibitors revealed by cryo-EM.

LRRK2 is one of the most promising drug targets for Parkinson's disease. Though type I kinase inhibitors of LRRK2 are under clinical trials, alternative strategies like type II inhibitors are being actively pursued due to the potential undesired effects of type I inhibitors. Currently, a robust method for LRRK2-inhibitor structure determination to guide structure-based drug discovery is lacking, and inhibition mechanisms of available compounds are also unclear. Here we present near-atomic-resolution structures of LRRK2 with type I (LRRK2-IN-1 and GNE-7915) and type II (rebastinib, ponatinib, and GZD-824) inhibitors, uncovering the structural basis of LRRK2 inhibition and conformational plasticity of the kinase domain with molecular dynamics (MD) simulations. Type I and II inhibitors bind to LRRK2 in active-like and inactive conformations, so LRRK2-inhibitor complexes further reveal general structural features associated with LRRK2 activation. Our study provides atomic details of LRRK2-inhibitor interactions and a framework for understanding LRRK2 activation and for rational drug design.

Engineering Nanosensitizer to Remodel the TME for Hypoimmunogenic "Cold"-"Hot" Tumor Transformations.

α-PD-L1 therapy has shown encouraging results at harnessing the immune system to combat cancer. However, the treatment effect is relatively low due to the dense extracellular matrix (ECM) and tumor immunosuppressive microenvironment (TIME). Therefore, an ultrasound (US)-responsive nanosensitizer (URNS) is engineered to deliver losartan (LST) and polyethylenimine (PEI) to remolde the TME, driving "cold"-"hot" tumor transformation and enhancing the sensitivity of α-PD-L1 therapy. In the tumor site, noninvasive US can make MTNP generate ROS, which cleave ROS-sensitive bonds to dissociate MTNPtK@LST-PEI, shedding PEI and releasing LST from mesoporous spheres. The results demonstrated that URNS combined with α-PD-L1 therapy effectively inhibited tumor growth with an inhibition rate as high as 90%, which was 1.7-fold higher than that of the α-PD-L1 treatment in vivo. In summary, the URNS improves the sensitivity of α-PD-L1 therapy by remodeling the TME, which provides promising insights for optimizing cancer immunotherapy.

Enantioselective synthesis of [4]helicenes by organocatalyzed intermolecular C-H amination.

Catalytic asymmetric synthesis of helically chiral molecules has remained an outstanding challenge and witnessed fairly limited progress in the past decades. Current methods to construct such compounds almost entirely rely on catalytic enantiocontrolled fused-ring system extension. Herein, we report a direct terminal peri-functionalization strategy, which allows for efficient assembling of 1,12-disubstituted [4]carbohelicenes via an organocatalyzed enantioselective amination reaction of 2-hydroxybenzo[c]phenanthrene derivates with diazodicarboxamides. The key feature of this approach is that the stereochemical information of the catalyst could be transferred into not only the helix sense but also the remote C-N axial chirality of the products, thus enabling the synthesis of [4]- and [5]helicenes with both structural diversity and stereochemical complexity in good efficiency and excellent enantiocontrol. Besides, the large-scale preparations and representative transformations of the helical products further demonstrate the practicality of this protocol. Moreover, DFT calculations reveal that both the hydrogen bonds and the C-H---π interactions between the substrates and catalyst contribute to the ideal stereochemical control.

Rab29-dependent asymmetrical activation of leucine-rich repeat kinase 2.

Gain-of-function mutations in LRRK2, which encodes the leucine-rich repeat kinase 2 (LRRK2), are the most common genetic cause of late-onset Parkinson's disease. LRRK2 is recruited to membrane organelles and activated by Rab29, a Rab guanosine triphosphatase encoded in the PARK16 locus. We present cryo-electron microscopy structures of Rab29-LRRK2 complexes in three oligomeric states, providing key snapshots during LRRK2 recruitment and activation. Rab29 induces an unexpected tetrameric assembly of LRRK2, formed by two kinase-active central protomers and two kinase-inactive peripheral protomers. The central protomers resemble the active-like state trapped by the type I kinase inhibitor DNL201, a compound that underwent a phase 1 clinical trial. Our work reveals the structural mechanism of LRRK2 spatial regulation and provides insights into LRRK2 inhibitor design for Parkinson's disease treatment.

Assessing base-resolution DNA mechanics on the genome scale.

Intrinsic DNA properties including bending play a crucial role in diverse biological systems. A recent advance in a high-throughput technology called loop-seq makes it possible to determine the bendability of hundred thousand 50-bp DNA duplexes in one experiment. However, it's still challenging to assess base-resolution sequence bendability in large genomes such as human, which requires thousands of such experiments. Here, we introduce 'BendNet'-a deep neural network to predict the intrinsic DNA bending at base-resolution by using loop-seq results in yeast as training data. BendNet can predict the DNA bendability of any given sequence from different species with high accuracy. To explore the utility of BendNet, we applied it to the human genome and observed DNA bendability is associated with chromatin features and disease risk regions involving transcription/enhancer regulation, DNA replication, transcription factor binding and extrachromosomal circular DNA generation. These findings expand our understanding on DNA mechanics and its association with transcription regulation in mammals. Lastly, we built a comprehensive resource of genomic DNA bendability profiles for 307 species by applying BendNet, and provided an online tool to assess the bendability of user-specified DNA sequences (http://www.dnabendnet.com/).

TRAmHap: accurate prediction of transcriptional activity from DNA methylation haplotypes in bisulfite-sequencing data.

Deoxyribonucleic acid (DNA) methylation (DNAm) is an important epigenetic mechanism that plays a role in chromatin structure and transcriptional regulation. Elucidating the relationship between DNAm and gene expression is of great importance for understanding its role in transcriptional regulation. The conventional approach is to construct machine-learning-based methods to predict gene expression based on mean methylation signals in promoter regions. However, this type of strategy only explains about 25% of gene expression variation, and hence is inadequate in elucidating the relationship between DNAm and transcriptional activity. In addition, using mean methylation as input features neglects the heterogeneity of cell populations that can be reflected by DNAm haplotypes. We here developed TRAmaHap, a novel deep-learning framework that predicts gene expression by utilizing the characteristics of DNAm haplotypes in proximal promoters and distal enhancers. Using benchmark data of human and mouse normal tissues, TRAmHap shows much higher accuracy than existing machine-learning based methods, by explaining 60~80% of gene expression variation across tissue types and disease conditions. Our model demonstrated that gene expression can be accurately predicted by DNAm patterns in promoters and long-range enhancers as far as 25 kb away from transcription start site, especially in the presence of intra-gene chromatin interactions.

Human IFT-A complex structures provide molecular insights into ciliary transport.

Intraflagellar transport (IFT) complexes, IFT-A and IFT-B, form bidirectional trains that move along the axonemal microtubules and are essential for assembling and maintaining cilia. Mutations in IFT subunits lead to numerous ciliopathies involving multiple tissues. However, how IFT complexes assemble and mediate cargo transport lacks mechanistic understanding due to missing high-resolution structural information of the holo-complexes. Here we report cryo-EM structures of human IFT-A complexes in the presence and absence of TULP3 at overall resolutions of 3.0-3.9 Å. IFT-A adopts a "lariat" shape with interconnected core and peripheral subunits linked by structurally vital zinc-binding domains. TULP3, the cargo adapter, interacts with IFT-A through its N-terminal region, and interface mutations disrupt cargo transport. We also determine the molecular impacts of disease mutations on complex formation and ciliary transport. Our work reveals IFT-A architecture, sheds light on ciliary transport and IFT train formation, and enables the rationalization of disease mutations in ciliopathies.

Structural basis of DNA polymerase θ mediated DNA end joining.

DNA polymerase θ (Pol θ) plays an essential role in the microhomology-mediated end joining (MMEJ) pathway for repairing DNA double-strand breaks. However, the mechanisms by which Pol θ recognizes microhomologous DNA ends and performs low-fidelity DNA synthesis remain unclear. Here, we present cryo-electron microscope structures of the polymerase domain of Lates calcarifer Pol θ with long and short duplex DNA at up to 2.4 Šresolution. Interestingly, Pol θ binds to long and short DNA substrates similarly, with extensive interactions around the active site. Moreover, Pol θ shares a similar active site as high-fidelity A-family polymerases with its finger domain well-closed but differs in having hydrophilic residues surrounding the nascent base pair. Computational simulations and mutagenesis studies suggest that the unique insertion loops of Pol θ help to stabilize short DNA binding and assemble the active site for MMEJ repair. Taken together, our results illustrate the structural basis of Pol θ-mediated MMEJ.

Enriched-Bromine Surface State for Stable Sky-Blue Spectrum Perovskite QLEDs With an EQE of 14.6.

Halogen vacancies are of great concern in blue-emitting perovskite quantum-dot light-emitting diodes because they affect their efficiency and spectral shift. Here, an enriched-bromine surface state is realized using a facile strategy that employs a PbBr2 stock solution for anion exchange based on Cd-doped perovskite quantum dots. It is found that the doped Cd ions are expected to reduce the formation energy of halogen vacancies filled by the external bromine ions, and the excess free bromine ions in solution are enriched in the surface by anchoring with halogen vacancies as sites, accompanied with the shedding of surface long-chain ligands during the anion exchange process, resulting in a Br-rich and "neat" surface. Moreover, the surface state exhibits good passivation of the surface defects of the controlled perovskite QDs and simultaneously increases the exciton binding energy, leading to excellent optical properties and stability. Finally, the sky-blue emitting perovskite quantum-dot light-emitting diodes (QLEDs) (490 nm) are conducted with a record external quantum efficiency of 14.6% and current efficiency of 19.9 cd A-1 . Meanwhile, the electroluminescence spectra exhibit great stability with negligible shifts under a constant operating voltage from 3 to 7 V. This strategy paves the way for improving the efficiency and stability of perovskite QLEDs.

Risk factors of sitting-induced tachycardia syndrome in children and adolescents.

BACKGROUND: The study was designed to explore the risk factors for sitting-induced tachycardia syndrome (STS) in children and adolescents. METHODS AND RESULTS: In this case-control study, 46 children with STS and 184 healthy children and adolescents were recruited. Demographic characteristics, lifestyle habits, allergy history, and family history were investigated using a questionnaire. The changes in heart rate and blood pressure from supine to sitting were monitored using a sitting test. The possible differences between STS patients and healthy children were analyzed using univariate analysis. Logistic regression analysis was used to explore the independent risk factors for STS. Univariate analysis showed that the daily sleeping time of the STS children were significantly shorter than that of the control group [(8.8 ± 1.2) hours/day vs. (9.3 ± 1.0) hours/day, P = 0.009], and the proportion of positive family history of syncope in the STS patients was higher than the controls (4/42 vs. 3/181, P = 0.044). Multivariate logistic regression studies showed that reduced daily sleeping time was an independent risk factor of STS in children (P = 0.006). Furthermore, when daily sleeping time was prolonged by 1 h, the risk of STS was decreased by 37.3%. CONCLUSION: Reduced daily sleeping was an independent risk factor for STS in children and adolescents.

Purimeth: an integrated web-based tool for estimating and accounting for tumor purity in cancer DNA methylation studies.

Proportion of cancerous cells in a tumor sample, known as "tumor purity", is a major source of confounding factor in cancer data analyses. Lots of computational methods are available for estimating tumor purity from different types of genomics data or based on different platforms, which makes it difficult to compare and integrate the estimated results. To rectify the deviation caused by tumor purity effect, a number of methods for downstream data analysis have been developed, including tumor sample clustering, association study and differential methylation between tumor samples. However, using these computational tools remains a daunting task for many researchers since they require non-trivial computational skills. To this end, we present Purimeth, an integrated web-based tool for estimating and accounting for tumor purity in cancer DNA methylation studies. Purimeth implements three state-of-the-art methods for tumor purity estimation from DNA methylation array data: InfiniumPurify, MEpurity and PAMES. It also provides graphical interface for various analyses including differential methylation (DM), sample clustering, and purification of tumor methylomes, all with the consideration of tumor purities. In addition, Purimeth catalogs estimated tumor purities for TCGA samples from nine methods for users to visualize and explore. In conclusion, Purimeth provides an easy-operated way for researchers to explore tumor purity and implement cancer methylation data analysis. It is developed using Shiny (Version 1.6.0) and freely available at http://purimeth.comp-epi.com/.

Dimensional Gradient Structure of CoSe2@CNTs-MXene Anode Assisted by Ether for High-Capacity, Stable Sodium Storage.

Recently, abundant resources, low-cost sodium-ion batteries are deemed to the new-generation battery in the field of large-scale energy storage. Nevertheless, poor active reaction dynamics, dissolution of intermediates and electrolyte matching problems are significant challenges that need to be solved. Herein, dimensional gradient structure of sheet-tube-dots is constructed with CoSe2@CNTs-MXene. Gradient structure is conducive to fast migration of electrons and ions with the association of ether electrolyte. For half-cell, CoSe2@CNTs-MXene exhibits high initial coulomb efficiency (81.7%) and excellent cycling performance (400 mAh g-1 cycling for 200 times in 2 A g-1). Phase transformation pathway from crystalline CoSe2-Na2Se with Co and then amorphous CoSe2 in the discharge/charge process is also explored by in situ X-ray diffraction. Density functional theory study discloses the CoSe2@CNTs-MXene in ether electrolyte system which contributes to stable sodium storage performance owing to the strong adsorption force from hierarchical structure and weak interaction between electrolyte and electrode interface. For full cell, CoSe2@CNTs-MXene//Na3V2 (PO4)3/C full battery can also afford a competitively reversible capacity of 280 mAh g-1 over 50 cycles. Concisely, profiting from dimensional gradient structure and matched electrolyte of CoSe2@CNTs-MXene hold great application potential for stable sodium storage.

Structural analysis of the full-length human LRRK2.

Mutations in leucine-rich repeat kinase 2 (LRRK2) are commonly implicated in the pathogenesis of both familial and sporadic Parkinson's disease (PD). LRRK2 regulates critical cellular processes at membranous organelles and forms microtubule-based pathogenic filaments, yet the molecular basis underlying these biological roles of LRRK2 remains largely enigmatic. Here, we determined high-resolution structures of full-length human LRRK2, revealing its architecture and key interdomain scaffolding elements for rationalizing disease-causing mutations. The kinase domain of LRRK2 is captured in an inactive state, a conformation also adopted by the most common PD-associated mutation, LRRK2G2019S. This conformation serves as a framework for structure-guided design of conformational specific inhibitors. We further determined the structure of COR-mediated LRRK2 dimers and found that single-point mutations at the dimer interface abolished pathogenic filamentation in cells. Overall, our study provides mechanistic insights into physiological and pathological roles of LRRK2 and establishes a structural template for future therapeutic intervention in PD.

Suppressing ion migration of CsPbBrxI3-xnanocrystals by Nickel doping and the application in high-efficiency WLEDs.

All inorganic perovskite nanocrystals CsPbX3(X = Cl, Br, I) are the great potential candidates for the application of high-performance light emitting diodes (LED) due to their high Photoluminescence Quantum Yield (PLQY), high defect tolerance, narrow full-width half-maximum and tunable wavelength of 410-700 nm. However, the application of red-emitting (630-650 nm) CsPbBrxI3-xnanocrystals are perplexed by phase segregation due to the composition of mixed halides and the difference in halide ion mobility. Herein, we provide an effective strategy to suppressing the migration of Br/I ions through Ni2+doping via a facile Hot-Injection method and the PLQY was improved as well. DFT calculations show that the introduction of Ni2+causes a slight contraction of the host crystal structure, which improves the bond energy between Pb and halides and reduces the level of surface defects. Therefore, the phase stability is improved by Ni2+doping because the phase segregation caused by ion migration in the mixed phase is effectively inhibited. Meanwhile, the non-radiative recombination in the exciton transition process is reduced and the PLQY is improved. What's more, benefiting from the suppressed ion migration and enhanced PLQY, we combine the Ni2+-doped CsPbBrxI3-xnanocrystals with different Br/I ratios and YAG: Ce3+phosphors as color conversion layers to fabricate high efficiency WLED. When the ratio of Br/I is 9:11, WLED has a color coordinate of (0.3621, 0.3458), the color temperature of 4336 K and presents a high luminous efficiency of 113.20 lm W-1, color rendering index of 94.9 under the driving current of 20 mA and exhibits excellent stability, which shows great potential in the application of LED.

Structure, substrate specificity, and catalytic mechanism of human D-2-HGDH and insights into pathogenicity of disease-associated mutations.

D-2-hydroxyglutarate dehydrogenase (D-2-HGDH) catalyzes the oxidation of D-2-hydroxyglutarate (D-2-HG) into 2-oxoglutarate, and genetic D-2-HGDH deficiency leads to abnormal accumulation of D-2-HG which causes type I D-2-hydroxyglutaric aciduria and is associated with diffuse large B-cell lymphoma. This work reports the crystal structures of human D-2-HGDH in apo form and in complexes with D-2-HG, D-malate, D-lactate, L-2-HG, and 2-oxoglutarate, respectively. D-2-HGDH comprises a FAD-binding domain, a substrate-binding domain, and a small C-terminal domain. The active site is located at the interface of the FAD-binding domain and the substrate-binding domain. The functional roles of the key residues involved in the substrate binding and catalytic reaction and the mutations identified in D-2-HGDH-deficient diseases are analyzed by biochemical studies. The structural and biochemical data together reveal the molecular mechanism of the substrate specificity and catalytic reaction of D-2-HGDH and provide insights into the pathogenicity of the disease-associated mutations.

A Ni-doping-induced phase transition and electron evolution in cobalt hexacyanoferrate as a stable cathode for sodium-ion batteries.

Prussian blue analogues are potential competitive energy storage materials due to their diverse metal combinations and wide three-dimensional ion channels. Here, we prepared a new highly crystalline monoclinic nickel-doped cobalt hexacyanoferrate via a feasible and simple one-step co-precipitation method. In the process of sodium-ion de-intercalation, three stable charge and discharge platforms, which are consistent with the cyclic voltammetry performance, are seen for the first time, showing the function of nickel ions in Prussian blue. Furthermore, the charge transfer and structural evolution caused by the transmission of sodium ions were well revealed via ex situ XRD, ex situ XPS, and in situ EIS studies. Simulation calculations are performed relating to the energy band structure and the highest-occupied bonding orbitals of the system in different charge states, revealing the charge and discharge mechanism of the nickel-doped material and the reason for the emergence of the new platform at low voltages. In addition, NaNi0.17Co0.83Fe(CN)6 also delivers a striking capacity of 146 mA h g-1 and superior cyclability, with 93% capacity retention over 100 cycles; it can be considered as a promising alternative cathode material for use in sodium-ion batteries.

Synthesis and Biological Characterization of Cyclic Disulfide-Containing Peptide Analogs of the Multifunctional Opioid/Neuropeptide FF Receptor Agonists That Produce Long-Lasting and Nontolerant Antinociception.

In a previously described chimeric peptide, we reported that the multifunctional opioid/neuropeptide FF (NPFF) receptor agonist 0 (BN-9) produced antinociception for 1.5 h after supraspinal administration. Herein, four cyclic disulfide analogs containing l- and/or d-type cysteine at positions 2 and 5 were synthesized. The cyclized analogs and their linear counterparts behaved as multifunctional agonists at both opioid and NPFF receptors in vitro and produced potent analgesia without tolerance development. In comparison to 0, cyclized peptide 6 exhibited sevenfold more potent µ-opioid receptor agonistic activity in vitro. Interestingly, the cyclized analog 6 possessed an improved stability in the brain and an increased blood-brain barrier permeability compared to the parent peptide 0 and produced more potent analgesia after supraspinal or subcutaneous administration with improved duration of action of 4 h. In addition, antinociceptive tolerance of analog 6 was greatly reduced after subcutaneous injection compared to fentanyl, as was the rewarding effect, withdrawal reaction, and gastrointestinal inhibition.

Sitting-induced hemodynamic changes and association with sitting intolerance in children and adolescents: a cross-sectional study.

Hemodynamic alteration with postural change from supine to sitting has been unclear in the young. In the cross-sectional study, 686 participants (371 boys and 315 girls, aged 6-18 years) were recruited from 4 schools in Kaifeng city, the central area of China. The active sitting test was performed to obtain heart rate (HR) and blood pressure (BP) changes from supine to sitting in children and adolescents. Hemodynamic change-associated sitting intolerance was analyzed. In the study participants, the 95th percentile (P95) values of changes in HR and BP within 3 min from supine to sitting were 25 beats/min and 18/19 mm Hg, respectively. Sixty-six participants had sitting intolerance symptoms. Compared with participants without sitting intolerance symptoms, those with symptoms more frequently had HR increase ≥ P95 or BP increase ≥ P95 within 3 min from supine to sitting (P < 0.001). Risk factors for sitting intolerance were age (odds ratio 1.218, 95% confidence interval 1.072-1.384, P = 0.002) and changes in HR or BP ≥ P95 within 3 min after sitting (odds ratio 2.902, 95% confidence interval 1.572-5.357, P = 0.001). We firstly showed hemodynamic changing profiles from supine to sitting and their association with sitting intolerance in children and adolescents. Sitting tachycardia is likely suggested with a change in HR ≥ 25 beats/min and sitting hypertension with a change in BP ≥ 20/20 mm Hg when changing from supine to sitting within 3 min. The age and changes in HR or BP were independent risk factors for sitting intolerance.

VF-13, a chimeric peptide of VD-hemopressin(α) and neuropeptide VF, produces potent antinociception with reduced cannabinoid-related side effects.

Pharmacological evidence indicated a functional interaction between neuropeptide FF (NPFF) and cannabinoid systems, and the cannabinoids combined with the NPFF receptor agonist neuropeptide VF (NPVF) produced antinociception without tolerance. In the present study, VF-13, a chimeric peptide containing the pharmacophores of the endogenous cannabinoid peptide VD-hemopressin(α) (VD-Hpα) and NPVF, was synthesized and pharmacologically evaluated. In vitro, VF-13 significantly upregulated the phosphorylated level of extracellular signal-regulated kinase 1/2 (ERK1/2) in CHO cells stably expressing CB1 receptors and inhibited forskolin-induced cAMP accumulation in HEK293 cells stably expressing NPFF1 or NPFF2 receptors. Moreover, VF-13 induced neurite outgrowth in Neuro 2A cells via CB1 and NPFF receptors. These results suggest that VF-13 exhibits multifunctional agonism at CB1, NPFF1 and NPFF2 receptors in vitro. Interestingly, intracerebroventricular VF-13 produced dose-dependent antinociception in mouse models of tail-flick and carrageenan-induced inflammatory pain via the TRPV1 receptor. In contrast, the reference compound (m)VD-Hpα-NH2 induced CB1 receptor-mediated supraspinal antinociception. Additionally, subcutaneous injection of (m)VD-Hpα-NH2 and VF-13 produced significant antinociception in carrageenan-induced inflammatory pain model. In the tetrad assay, our data demonstrated that VF-13 elicited hypothermia, but not catalepsy and hypoactivity after intracerebroventricular injection. Notably, VF-13 produced non-tolerance forming antinociception over 6 days treatment in both acute and inflammatory pain models. Furthermore, VF-13 had no apparent effects on gastrointestinal transit, pentobarbitone-induced sedation, food intake, and motor coordination at the supraspinal level. In summary, VF-13, a novel chimeric peptide of VD-Hpα and NPVF, produced non-tolerance forming antinociception in preclinical pain models with reduced cannabinoid-related side effects.