The GAUGAA Motif Is Responsible for the Binding between circSMARCA5 and SRSF1 and Related Downstream Effects on Glioblastoma Multiforme Cell Migration and Angiogenic Potential

Circular RNAs (circRNAs) are a large class of RNAs with regulatory functions within cells. We recently showed that circSMARCA5 is a tumor suppressor in glioblastoma multiforme (GBM) and acts as a decoy for Serine and Arginine Rich Splicing Factor 1 (SRSF1) through six predicted binding sites (BSs). Here we characterized RNA motifs functionally involved in the interaction between circSMARCA5 and SRSF1. Three different circSMARCA5 molecules (Mut1, Mut2, Mut3), each mutated in two predicted SRSF1 BSs at once, were obtained through PCR-based replacement of wild-type (WT) BS sequences and cloned in three independent pcDNA3 vectors. Mut1 significantly decreased its capability to interact with SRSF1 as compared to WT, based on the RNA immunoprecipitation assay. In silico analysis through the “Find Individual Motif Occurrences” (FIMO) algorithm showed GAUGAA as an experimentally validated SRSF1 binding motif significantly overrepresented within both predicted SRSF1 BSs mutated in Mut1 (q-value = 0.0011). U87MG and CAS-1, transfected with Mut1, significantly increased their migration with respect to controls transfected with WT, as revealed by the cell exclusion zone assay. Immortalized human brain microvascular endothelial cells (IM-HBMEC) exposed to conditioned medium (CM) harvested from U87MG and CAS-1 transfected with Mut1 significantly sprouted more than those treated with CM harvested from U87MG and CAS-1 transfected with WT, as shown by the tube formation assay. qRT-PCR showed that the intracellular pro- to anti-angiogenic Vascular Endothelial Growth Factor A (VEGFA) mRNA isoform ratio and the amount of total VEGFA mRNA secreted in CM significantly increased in Mut1-transfected CAS-1 as compared to controls transfected with WT. Our data suggest that GAUGAA is the RNA motif responsible for the interaction between circSMARCA5 and SRSF1 as well as for the circSMARCA5-mediated control of GBM cell migration and angiogenic potential.     

Complement-Opsonized Nano-Carriers Are Bound by Dendritic Cells (DC) via Complement Receptor (CR)3, and by B Cell Subpopulations via CR-1/2, and Affect the Activation of DC and B-1 Cells

The development of nanocarriers (NC) for biomedical applications has gained large interest due to their potential to co-deliver drugs in a cell-type-targeting manner. However, depending on their surface characteristics, NC accumulate serum factors, termed protein corona, which may affect their cellular binding. We have previously shown that NC coated with carbohydrates to enable biocompatibility triggered the lectin-dependent complement pathway, resulting in enhanced binding to B cells via complement receptor (CR)1/2. Here we show that such NC also engaged all types of splenic leukocytes known to express CR3 at a high rate when NC were pre-incubated with native mouse serum resulting in complement opsonization. By focusing on dendritic cells (DC) as an important antigen-presenting cell type, we show that CR3 was essential for binding/uptake of complement-opsonized NC, whereas CR4, which in mouse is specifically expressed by DC, played no role. Further, a minor B cell subpopulation (B-1), which is important for first-line pathogen responses, and co-expressed CR1/2 and CR3, in general, engaged NC to a much higher extent than normal B cells. Here, we identified CR-1/2 as necessary for binding of complement-opsonized NC, whereas CR3 was dispensable. Interestingly, the binding of complement-opsonized NC to both DC and B-1 cells affected the expression of activation markers. Our findings may have important implications for the design of nano-vaccines against infectious diseases, which codeliver pathogen-specific protein antigen and adjuvant, aimed to induce a broad adaptive cellular and humoral immune response by inducing cytotoxic T lymphocytes that kill infected cells and pathogen-neutralizing antibodies, respectively. Decoration of nano-vaccines either with carbohydrates to trigger complement activation in vivo or with active complement may result in concomitant targeting of DC and B cells and thereby may strongly enhance the extent of dual cellular/humoral immune responses.     

Local Protein Translation and RNA Processing of Synaptic Proteins in Autism Spectrum Disorder

Autism spectrum disorder (ASD) is a heritable neurodevelopmental condition associated with impairments in social interaction, communication and repetitive behaviors. While the underlying disease mechanisms remain to be fully elucidated, dysfunction of neuronal plasticity and local translation control have emerged as key points of interest. Translation of mRNAs for critical synaptic proteins are negatively regulated by Fragile X mental retardation protein (FMRP), which is lost in the most common single-gene disorder associated with ASD. Numerous studies have shown that mRNA transport, RNA metabolism, and translation of synaptic proteins are important for neuronal health, synaptic plasticity, and learning and memory. Accordingly, dysfunction of these mechanisms may contribute to the abnormal brain function observed in individuals with autism spectrum disorder (ASD). In this review, we summarize recent studies about local translation and mRNA processing of synaptic proteins and discuss how perturbations of these processes may be related to the pathophysiology of ASD.     

Accurate Receptor-Ligand Binding Free Energies from Fast QM Conformational Chemical Space Sampling

Small molecule receptor-binding is dominated by weak, non-covalent interactions such as van-der-Waals hydrogen bonding or electrostatics. Calculating these non-covalent ligand-receptor interactions is a challenge to computational means in terms of accuracy and efficacy since the ligand may bind in a number of thermally accessible conformations. The conformational rotamer ensemble sampling tool (CREST) uses an iterative scheme to efficiently sample the conformational space and calculates energies using the semi-empirical ‘Geometry, Frequency, Noncovalent, eXtended Tight Binding’ (GFN2-xTB) method. This combined approach is applied to blind predictions of the modes and free energies of binding for a set of 10 drug molecule ligands to the cucurbit[n]urils CB[8] receptor from the recent ‘Statistical Assessment of the Modeling of Proteins and Ligands’ (SAMPL) challenge including morphine, hydromorphine, cocaine, fentanyl, and ketamine. For each system, the conformational space was sufficiently sampled for the free ligand and the ligand-receptor complexes using the quantum chemical Hamiltonian. A multitude of structures makes up the final conformer-rotamer ensemble, for which then free energies of binding are calculated. For those large and complex molecules, the results are in good agreement with experimental values with a mean error of 3 kcal/mol. The GFN2-xTB energies of binding are validated by advanced density functional theory calculations and found to be in good agreement. The efficacy of the automated QM sampling workflow allows the extension towards other complex molecular interaction scenarios.     

基于3D scSE-UNet的肝脏CT图像半监督学习分割方法

针对分割神经网络需要大量的高质量标签但较难获取的问题,提出基于3D scSE-UNet的半监督学习分割方法. 该方法使用自训练的半监督学习框架,将包含改进的并行空间/特征通道压缩和激励模块（scSE-block+）的3D scSE-UNet作为分割网络. scSE-block+可以从图像空间和特征通道2个方面自动学习图像的有效特征,抑制无用冗余特征,更好地保留图像边缘信息. 在自训练过程中加入全连接条件随机场,对分割网络产生的伪标签进行边缘细化,提升伪标签的精确度. 在LiTS17 Challenge和SLIVER07数据集上验证所提出方法的有效性. 当有标签图像占训练集总图像的30%时,所提方法的Dice相似系数(dice score)为0.941. 结果表明,所提出的半监督学习分割方法可以在仅使用少量标注数据的情况下,取得与全监督分割方法相当的分割效果,有效减轻肝脏CT图像分割对专家标注数据的依赖.     

MS Binding Assays for Glycine Transporter 2 (GlyT2) Employing Org25543 as Reporter Ligand

This study describes the first binding assay for glycine transporter 2 (GlyT2) following the concept of MS Binding Assays. The selective GlyT2 inhibitor Org25543 was employed as a reporter ligand and it was quantified with a highly sensitive and rapid LC-ESI-MS/MS method. Binding of Org25543 at GlyT2 was characterized in kinetic and saturation experiments with an off-rate of 7.07×10⁻³ s⁻¹, an on-rate of 1.01×10⁶ M⁻¹ s⁻¹, and an equilibrium dissociation constant of 7.45 nM. Furthermore, the inhibitory constants of 19 GlyT ligands were determined in competition experiments. The validity of the GlyT2 affinities determined with the binding assay was examined by a comparison with published inhibitory potencies from various functional assays. With the capability for affinity determination towards GlyT2 the developed MS Binding Assays provide the first tool for affinity profiling of potential ligands and it represents a valuable new alternative to functional assays addressing GlyT2.     

边缘AI的霜雪识别系统设计与实现

为提高霜雪识别系统的识别率,开展了基于边缘AI的霜雪识别系统设计。综合海思嵌入式处理器Hi3559A高算力、低功耗特点,硬件部分采用模块复用的方法降低功耗和资源消耗,软件部分,从模型训练、模型量化、模型部署入手对MobileNetV2图像分类网络和ISP自适应处理算法进行改进,最终识别率99.7%,预处理时间0.3 s,图像分类时间0.8 s,模组平均功率2 W,鲁棒性好、性能稳定。由此可见Hi3559A智能相机模组可有效纠正雪深探测仪的疑误数据。     

Supramolecular Nanofibers from Collagen-Mimetic Peptides Bearing Various Aromatic Groups at N-Termini via Hierarchical Self-Assembly

Self-assembly of artificial peptides has been widely studied for constructing nanostructured materials, with numerous potential applications in the nanobiotechnology field. Herein, we report the synthesis and hierarchical self-assembly of collagen-mimetic peptides (CMPs) bearing various aromatic groups at the N-termini, including 2-naphthyl, 1-naphtyl, anthracenyl, and pyrenyl groups, into nanofibers. The CMPs (R-(GPO)_n: n > 4) formed a triple helix structure in water at 4 °C, as confirmed via CD analyses, and their conformations were more stable with increasing hydrophobicity of the terminal aromatic group and peptide chain length. The resulting pre-organized triple helical CMPs showed diverse self-assembly into highly ordered nanofibers, reflecting their slight differences in hydrophobic/hydrophilic balance and configuration of aromatic templates. TEM analysis demonstrated that 2Np-CMP_n (n = 6 and 7) and Py-CMP₆ provided well-developed natural collagen-like nanofibers and An-CMP_n (n = 5–7) self-assembled into rod-like micelle fibers. On the other hand, 2Np-CMP₅ and 1Np-CMP₆ were unable to form nanofibers under the same conditions. Furthermore, the Py-CMP₆ nanofiber was found to encapsulate a guest hydrophobic molecule, Nile red, and exhibited unique emission behavior based on the specific nanostructure. In addition to the ability of CMPs to bind small molecules, their controlled self-assembly enables their versatile utilization in drug delivery and wavelength-conversion nanomaterials.     

Quest for the Crypto-phosphoproteome

Protein phosphorylation is one of the most studied post-translational modifications (PTMs). Despite the remarkable advances in phosphoproteomics, a chemically less-stable subset of the phosphosites, which we call the crypto-phosphoproteome, has remained underexplored due to technological challenges. In this Viewpoint, we briefly summarize the current understanding of these elusive protein phosphorylations and identify the missing pieces for future studies.     

划痕法测Ni—P合金镀层结合力的研究

本文用划痕法测定了Ni-P合金镀层的结合力;观察了划痕侧面镀层的失效形式;分析了三种评定结合力的参数。结果指出,临界载荷L_C、平均应力σ_A用于评定Ni-P镀层的结合力是有条件的,而B-W公式存在的问题更多。