Structural determinants for membrane binding of the EGFR juxtamembrane domain.

Overactivation of the epidermal growth factor receptor (EGFR) is critical for the development of multiple cancers. Previous studies have shown that the cell membrane is a key regulator of EGFR kinase activity through its interaction with the EGFR juxtamembrane domain (JM). However, the lipid recognition specificity of EGFR-JM and its interaction details remain unclear. Using lipid strip and liposome pulldown assays, we showed that EGFR-JM could specifically interact with PI(4,5)P2-or phosphatidylserine-containing membranes. We further characterized the JM-membrane interaction using NMR-titration-based chemical shift perturbation and paramagnetic relaxation enhancement analyses, and found that residues I649 - L659 comprised the membrane-binding site. Furthermore, the membrane-binding region contains the predicted dimerization motif of JM, 655LRRLL659, suggesting that membrane binding may affect JM dimerization and, therefore, regulate kinase activation.

New Insights into the Dependence of CPEB3 Ribozyme Cleavage on Mn2+ and Mg2.

CPEB3 ribozyme is a self-cleaving RNA that occurs naturally in mammals and requires divalent metal ions for efficient activity. Ribozymes exhibit preferences for specific metal ions, but the exact differences in the catalytic mechanisms of various metal ions on the CPEB3 ribozyme remain unclear. Our findings reveal that Mn2+ functions as a more effective cofactor for CPEB3 ribozyme catalysis compared to Mg2+, as confirmed by its stronger binding affinity to CPEB3 by EPR. Cleavage assays of CPEB3 mutants and molecular docking analyses further showed that excessive Mn2+ ions can bind to a second binding site near the catalytic site, hindering CPEB3 catalytic efficiency and contributing to the Mn2+ bell-shaped curve. These results implicate a pivotal role for the local nucleobase-Mn2+ interactions in facilitating RNA folding and modulating the directed attack of nucleophilic reagents. Our study provides new insights and experimental evidence for exploring the divalent cation dependent cleavage mechanism of the CPEB3 ribozyme.

Astragaloside IV inhibits idiopathic pulmonary fibrosis through activation of autophagy by miR-21-mediated PTEN/PI3K/AKT/mTOR pathway.

As the main active ingredient of Astragalus, Astragaloside IV (AS-IV) can ameliorate pulmonary fibrosis. In this experiment, we studied how AS-IV reduces idiopathic pulmonary fibrosis (IPF). Bleomycin (BLM) or TGF-ß1 was treated in mice or alveolar epithelial cells to mimic IPF in vivo as well as in vitro. ASV-IV alleviated levels of inflammatory cytokines and fibrosis markers in IPF model. Through detection of autophagy-related genes, ASV-IV was observed to induce autophagy in IPF. Besides, ASV-IV inhibited miR-21 expression in IPF models, and overexpression of miR-21 could reverse the protective potential of ASV-IV on IPF. PTEN was targeted by miR-21 and was up-regulated by ASV-IV in IPF models. In addition, levels of inflammatory cytokines and fibrosis markers, autophagy, as well as the PI3K/AKT/mTOR pathway regulated by ASV-IV could be neutralized after treatment with autophagy inhibitors, miR-21 mimics, or si-PTEN. Our study demonstrates that ASV-IV inhibits IPF through activation of autophagy by miR-21-mediated PTEN/PI3K/AKT/mTOR pathway, suggesting that ASV-IV could be acted to be a promising therapeutic method for IPF.

Brain magnetic resonance imaging findings in children with neurological complications of coronavirus disease 2019 (Omicron variant): a multicenter retrospective observational study.

BACKGROUND: An increasing rate of encephalopathy associated with coronavirus disease 2019 (COVID-19) has been observed among children. However, the literature on neuroimaging data in children with COVID-19 is limited. OBJECTIVE: To analyze brain magnetic resonance imaging (MRI) of pediatric COVID-19 patients with neurological complications. MATERIALS AND METHODS: This multicenter retrospective observational study analyzed clinical (n=102, 100%) and neuroimaging (n=93, 91.2%) data of 102 children with COVID-19 infections and comorbid acute neurological symptoms. These children were hospitalized at five pediatric intensive care units (PICUs) in China between December 1, 2022, and January 31, 2023. RESULTS: All patients were positive for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) as detected via reverse transcriptase polymerase chain reaction. About 75.7% of the children were infected with the Omicron variant BF.7 strain. Brain MRI was performed 1-12 days following the onset of neurological symptoms, which revealed acute neuroimaging findings in 74.2% (69/93) of cases, including evidence of acute necrotizing encephalopathy (33/69, 47.8%), encephalitis (31/69, 44.9%), reversible splenial lesion syndrome (3/69, 4.3%), reversible posterior leukoencephalopathy (1/69, 1.4%), and hippocampal atrophy (1/69, 1.4%). CONCLUSIONS: Overall, these data highlighted five neuroimaging patterns associated with the outbreak of the SARS-CoV-2 Omicron variant, with acute necrotizing encephalopathy being the most common of these neuroimaging findings. Rarely, the brain MRI of these pediatric COVID-19 patients also demonstrate hippocampal atrophy.

A Case Report of Zosteriform Cutaneous Metastases from Breast Carcinoma.

We described a 58-year-old female diagnosed with zosteriform cutaneous metastases from breast carcinoma. She was initially diagnosed with herpes zoster. Correct diagnosis was obtained after pathological biopsy. Various forms of cutaneous metastases have various forms, which require careful discrimination by dermatologists to reduce the rate of misdiagnosis.

Molecular targets and mechanisms of Jiawei Jiaotai Pill on diabetic cardiomyopathy based on network pharmacology.

BACKGROUND: Jiawei Jiaotai Pill is commonly used in clinical practice to reduce apoptosis, increase insulin secretion, and improve blood glucose tolerance. However, its mechanism of action in the treatment of diabetic cardiomyopathy (DCM) remains unclear, hindering research efforts aimed at developing drugs specifically for the treatment of DCM. AIM: To explore the pharmacodynamic basis and molecular mechanism of Jiawei Jiaotai Pill in DCM treatment. METHODS: We explored various databases and software, including the Traditional Chinese Medicine Systems Pharmacology Database, Uniport, PubChem, GenCards, String, and Cytoscape, to identify the active components and targets of Jiawei Jiaotai Pill, and the disease targets in DCM. Protein-protein interaction network, gene ontology, and Kyoto Encyclopedia of Genes and Genomes analyses were used to determine the mechanism of action of Jiawei Jiaotai Pill in treating DCM. Molecular docking of key active components and core targets was verified using AutoDock software. RESULTS: Total 42 active ingredients and 142 potential targets of Jiawei Jiaotai Pill were identified. There were 100 common targets between the DCM and Jiawei Jiaotai Pills. Through this screening process, TNF, IL6, TP53, EGFR, INS, and other important targets were identified. These targets are mainly involved in the positive regulation of the mitogen-activated protein kinase (MAPK) MAPK cascade, response to xenobiotic stimuli, response to hypoxia, positive regulation of gene expression, positive regulation of cell proliferation, negative regulation of the apoptotic process, and other biological processes. It was mainly enriched in the AGE-RAGE signaling pathway in diabetic complications, DCM, PI3K-Akt, interleukin-17, and MAPK signaling pathways. Molecular docking results showed that Jiawei Jiaotai Pill's active ingredients had good docking activity with DCM's core target. CONCLUSION: The active components of Jiawei Jiaotai Pill may play a role in the treatment of DCM by reducing oxidative stress, cardiomyocyte apoptosis and fibrosis, and maintaining metabolic homeostasis.

The role of mitochondrial dynamics in disease.

Mitochondria are multifaceted and dynamic organelles regulating various important cellular processes from signal transduction to determining cell fate. As dynamic properties of mitochondria, fusion and fission accompanied with mitophagy, undergo constant changes in number and morphology to sustain mitochondrial homeostasis in response to cell context changes. Thus, the dysregulation of mitochondrial dynamics and mitophagy is unsurprisingly related with various diseases, but the unclear underlying mechanism hinders their clinical application. In this review, we summarize the recent developments in the molecular mechanism of mitochondrial dynamics and mitophagy, particularly the different roles of key components in mitochondrial dynamics in different context. We also summarize the roles of mitochondrial dynamics and target treatment in diseases related to the cardiovascular system, nervous system, respiratory system, and tumor cell metabolism demanding high-energy. In these diseases, it is common that excessive mitochondrial fission is dominant and accompanied by impaired fusion and mitophagy. But there have been many conflicting findings about them recently, which are specifically highlighted in this view. We look forward that these findings will help broaden our understanding of the roles of the mitochondrial dynamics in diseases and will be beneficial to the discovery of novel selective therapeutic targets.

Facile and stereospecific synthesis of diverse ß-N-glycosyl sulfonamide scaffolds via palladium catalysis.

Glycosylation is an important strategy to improve the druggability of lead compounds. Here, we present a palladium-catalysed stereospecific N-glycosylation of sulfonamides. This approach stands out with wide substrate scope, high functional group tolerance, and easy scalability, furnishing a broad spectrum of densely functionalized ß-N-glycosyl sulfonamides with good efficiency and exceptional regio-/stereoselectivity. Diverse drug-like glycosulfonamido scaffolds have been constructed via a late-stage diversification strategy and various facile synthetic transformations of the products. Collectively, the established protocol provides a valuable tool for efficiently preparing glycosyl sulfonamides to facilitate drug discovery.

Performance-Based Distributed Control of Multiagent Systems: A Dual Phase Approach.

In this article, we investigate the distributed tracking control problem for networked uncertain nonlinear strict-feedback systems with unknown time-varying gains under a directed interaction topology. A dual phase performance-guaranteed approach is established. In the first phase, a fully distributed robust filter is constructed for each agent to estimate the desired trajectory with prescribed performance such that the control directions of all agents are allowed to be nonidentical. In the second phase, by establishing a novel lemma regarding Nussbaum function, a new adaptive control protocol is developed for each agent based on backstepping technique, which not only steers the output to track the corresponding estimated signal asymptotically with arbitrarily prescribed transient response but also extends the application scope of the proposed control scheme largely since the unknown control gains are allowed to be time-varying and even state-dependent. In such a way, the underlying problem is tackled with the output tracking error converging into an arbitrarily preassigned residual set exhibiting an arbitrarily predefined convergence rate. Besides, all the internal signals are ensured to be semi-globally ultimately uniformly bounded (SGUUB). Finally, two examples are provided to illustrate the effectiveness of the co-designed scheme.

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/).

Deep Learner System Based on Focal Color Retinal Fundus Images to Assist in Diagnosis.

Retinal diseases are a serious and widespread ophthalmic disease that seriously affects patients' vision and quality of life. With the aging of the population and the change in lifestyle, the incidence rate of retinal diseases has increased year by year. However, traditional diagnostic methods often require experienced doctors to analyze and judge fundus images, which carries the risk of subjectivity and misdiagnosis. This paper will analyze an intelligent medical system based on focal retinal image-aided diagnosis and use a convolutional neural network (CNN) to recognize, classify, and detect hard exudates (HEs) in fundus images (FIs). The research results indicate that under the same other conditions, the accuracy, recall, and precision of the system in diagnosing five types of patients with pathological changes under color retinal FIs range from 86.4% to 98.6%. Under conventional retinopathy FIs, the accuracy, recall, and accuracy of the system in diagnosing five types of patients ranged from 70.1% to 85%. The results show that the application of focus color retinal FIs in the intelligent medical system has high accuracy and reliability for the early detection and diagnosis of diabetic retinopathy and has important clinical applications.

Dexmedetomidine improves the outcomes for pediatric severe sepsis with mechanical ventilation.

BACKGROUND: The sedative dexmedetomidine has been shown to reduce mortality in adult patients with severe sepsis, but it is not known whether children benefit. This study explored the effects of dexmedetomidine on the outcomes of children with severe sepsis with mechanical ventilation. METHODS: In this retrospective cohort study, children with severe sepsis requiring mechanical ventilation from 2016 to 2020 were categorized as dexmedetomidine and non-dexmedetomidine group. The propensity score matching was performed to match cases in both groups. The primary outcome was 28-day mortality, and the secondary outcomes were acute kidney injury, ventilator-free days, lengths of PICU and hospital stays. The Kaplan-Meier method and was the log-rank test used to estimate the 28-day mortality rate and assess between-group differences. RESULTS: In total, 250 patients were eligible patients: 138 in the dexmedetomidine group and 112 in the non-dexmedetomidine group. After 1:1 propensity score matching, 61 children in each group. dexmedetomidine group showed more lower 28-day mortality (9.84% vs. 26.23%, P = 0.008). During the 7-day observation period after PICU admission, the dexmedetomidine group showed significantly lower neurological and renal sub-scores at day 7 and serum creatinine level at day 3 and day 7. There were no statistical differences in the incidence of acute kidney injury, ventilator-free days, lengths of PICU and hospital stays between the two groups. CONCLUSIONS: dexmedetomidine treatment in children with severe sepsis is associated with better outcomes and should therefore be considered for the sedation strategy.

[Progress in neural network mechanism of tinnitus using functional magnetic resonance imaging].

Tinnitus refers to the perception of abnormal sound in the absence of external sound stimulation. It can have an impact on a person's mood, memory, attention, and mental state, although the mechanism of tinnitus is still unclear. In recent years, the research on the central neural mechanism of tinnitus has attracted the attention of scholars.Functional magnetic resonance imaging （fMRI）,as an effective imaging technology, has been actively employed in this field. This paper provides a systematic summary of studies on the central neural mechanism of tinnitus by fMRI in recent years,revealed the changes of functional connections among tinnitus-related neural networks,such as auditory network,limbic system,default mode network and salience network. The central neural mechanism of tinnitus involves multiple networks that interact with each other. By understanding this mechanism, we hope to develop more targeted prevention and treatment strategies to help patients alleviate long-term tinnitus.

The distinct initiation sites and processing activities of TTLL4 and TTLL7 in glutamylation of brain tubulin.

Mammalian brain tubulins undergo a reversible posttranslational modification-polyglutamylation-which attaches a secondary polyglutamate chain to the primary sequence of proteins. Loss of its erasers can disrupt polyglutamylation homeostasis and cause neurodegeneration. Tubulin tyrosine ligase like 4 (TTLL4) and TTLL7 were known to modify tubulins, both with preference for the ß-isoform, but differently contribute to neurodegeneration. However, differences in their biochemical properties and functions remain largely unknown. Here, using an antibody-based method, we characterized the properties of a purified recombinant TTLL4 and confirmed its sole role as an initiator, unlike TTLL7, which both initiates and elongates the side chains. Unexpectedly, TTLL4 produced stronger glutamylation immunosignals for α-isoform than ß-isoform in brain tubulins. Contrarily, the recombinant TTLL7 raised comparable glutamylation immunoreactivity for two isoforms. Given the site selectivity of the glutamylation antibody, we analyzed modification sites of two enzymes. Tandem mass spectrometry analysis revealed their incompatible site selectivity on synthetic peptides mimicking carboxyl termini of α1- and ß2-tubulins and a recombinant tubulin. Particularly, in the recombinant α1A-tubulin, a novel region was found glutamylated by TTLL4 and TTLL7, that again at distinct sites. These results pinpoint different site specificities between two enzymes. Moreover, TTLL7 exhibits less efficiency to elongate microtubules premodified by TTLL4, suggesting possible regulation of TTLL7 elongation activity by TTLL4-initiated sites. Finally, we showed that kinesin behaves differentially on microtubules modified by two enzymes. This study underpins the different reactivity, site selectivity, and function of TTLL4 and TTLL7 on brain tubulins and sheds light on their distinct role in vivo.

Antibacterial 3D-Printed Silver Nanoparticle/Poly Lactic-Co-Glycolic Acid (PLGA) Scaffolds for Bone Tissue Engineering.

Infectious bone defects present a major challenge in the clinical setting currently. In order to address this issue, it is imperative to explore the development of bone tissue engineering scaffolds that are equipped with both antibacterial and bone regenerative capabilities. In this study, we fabricated antibacterial scaffolds using a silver nanoparticle/poly lactic-co-glycolic acid (AgNP/PLGA) material via a direct ink writing (DIW) 3D printing technique. The scaffolds' microstructure, mechanical properties, and biological attributes were rigorously assessed to determine their fitness for repairing bone defects. The surface pores of the AgNPs/PLGA scaffolds were uniform, and the AgNPs were evenly distributed within the scaffolds, as confirmed via scanning electron microscopy (SEM). Tensile testing confirmed that the addition of AgNPs enhanced the mechanical strength of the scaffolds. The release curves of the silver ions confirmed that the AgNPs/PLGA scaffolds released them continuously after an initial burst. The growth of hydroxyapatite (HAP) was characterized via SEM and X-ray diffraction (XRD). The results showed that HAP was deposited on the scaffolds, and also confirmed that the scaffolds had mixed with the AgNPs. All scaffolds containing AgNPs exhibited antibacterial properties against Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli). A cytotoxicity assay using mouse embryo osteoblast precursor cells (MC3T3-E1) showed that the scaffolds had excellent biocompatibility and could be used for repairing bone tissue. The study shows that the AgNPs/PLGA scaffolds have exceptional mechanical properties and biocompatibility, effectively inhibiting the growth of S. aureus and E. coli. These results demonstrate the potential application of 3D-printed AgNPs/PLGA scaffolds in bone tissue engineering.

Stereospecific Palladium-Catalyzed Direct Glycosylation of Oximes: Access to N-O-Linked Glycosides.

A highly efficient, palladium-catalyzed glycosylation between 3,4-O-carbonate glycals and acid-labile oximes is disclosed. This approach features broad substrate scope, high functional group tolerance, and easy scalability, delivering glycosyl oximes in excellent yields with exclusive ß-selectivity and retention of Z/E geometries. The power of this method is demonstrated by a set of site-selective transformations of glycosylation products and late-stage glycodiversification of bioactive molecules. Overall, our strategy provides an efficient toolkit for facile access to valuable N-O-linked glycosides.

Disruption of the nuclear localization signal in RBM20 is causative in dilated cardiomyopathy.

Human patients carrying genetic mutations in RNA binding motif 20 (RBM20) develop a clinically aggressive dilated cardiomyopathy (DCM). Genetic mutation knockin (KI) animal models imply that altered function of the arginine-serine-rich (RS) domain is crucial for severe DCM. To test this hypothesis, we generated an RS domain deletion mouse model (Rbm20ΔRS). We showed that Rbm20ΔRS mice manifested DCM with mis-splicing of RBM20 target transcripts. We found that RBM20 was mis-localized to the sarcoplasm in Rbm20ΔRS mouse hearts and formed RBM20 granules similar to those detected in mutation KI animals. In contrast, mice lacking the RNA recognition motif showed similar mis-splicing of major RBM20 target genes but did not develop DCM or exhibit RBM20 granule formation. Using in vitro studies with immunocytochemical staining, we demonstrated that only DCM-associated mutations in the RS domain facilitated RBM20 nucleocytoplasmic transport and promoted granule assembly. Further, we defined the core nuclear localization signal (NLS) within the RS domain of RBM20. Mutation analysis of phosphorylation sites in the RS domain suggested that this modification may be dispensable for RBM20 nucleocytoplasmic transport. Collectively, our findings revealed that disruption of RS domain-mediated nuclear localization is crucial for severe DCM caused by NLS mutations.

CCP5 and CCP6 retain CP110 and negatively regulate ciliogenesis.

BACKGROUND: The axonemal microtubules of primary cilium undergo a conserved protein posttranslational modification (PTM) - polyglutamylation. This reversible procedure is processed by tubulin tyrosine ligase-like polyglutamylases to form secondary polyglutamate side chains, which are metabolized by the 6-member cytosolic carboxypeptidase (CCP) family. Although polyglutamylation modifying enzymes have been linked to ciliary architecture and motility, it was unknown whether they also play a role in ciliogenesis. RESULTS: In this study, we found that CCP5 expression is transiently downregulated upon the initiation of ciliogenesis, but recovered after cilia are formed. Overexpression of CCP5 inhibited ciliogenesis, suggesting that a transient downregulation of CCP5 expression is required for ciliation initiation. Interestingly, the inhibitory effect of CCP5 on ciliogenesis does not rely on its enzyme activity. Among other 3 CCP members tested, only CCP6 can similarly suppress ciliogenesis. Using CoIP-MS analysis, we identified a protein that potentially interacts with CCP - CP110, a known negative regulator of ciliogenesis, whose degradation at the distal end of mother centriole permits cilia assembly. We found that both CCP5 and CCP6 can modulate CP110 level. Particularly, CCP5 interacts with CP110 through its N-terminus. Loss of CCP5 or CCP6 led to the disappearance of CP110 at the mother centriole and abnormally increased ciliation in cycling RPE-1 cells. Co-depletion of CCP5 and CCP6 synergized this abnormal ciliation, suggesting their partially overlapped function in suppressing cilia formation in cycling cells. In contrast, co-depletion of the two enzymes did not further increase the length of cilia, although CCP5 and CCP6 differentially regulate polyglutamate side-chain length of ciliary axoneme and both contribute to limiting cilia length, suggesting that they may share a common pathway in cilia length control. Through inducing the overexpression of CCP5 or CCP6 at different stages of ciliogenesis, we further demonstrated that CCP5 or CCP6 inhibited cilia formation before ciliogenesis, while shortened the length of cilia after cilia formation. CONCLUSION: These findings reveal the dual role of CCP5 and CCP6. In addition to regulating cilia length, they also retain CP110 level to suppress cilia formation in cycling cells, pointing to a novel regulatory mechanism for ciliogenesis mediated by demodifying enzymes of a conserved ciliary PTM, polyglutamylation.

JLP/Foxk1/N-cadherin axis fosters a partial epithelial-mesenchymal transition state in epithelial tubular cells.

Renal tubular epithelial cells (TECs) undergoing partial epithelial-mesenchymal transition (pEMT) during renal fibrosis has been recognized as a featuring and detrimental event. However, the mechanism for redirecting the cell fate of pEMT remains unclear. Here we mapped the temporal expression trajectories of a series of EMT-related molecules in renal fibrosis. It revealed a unique expression profile of N-cadherin of initial rising and late dropdown, which is distinct from that of other mesenchymal markers. The transcription factor Foxk1, which serves as a negative regulator of the N-cadherin gene, was induced by TGF-ß1 but was tightly regulated in the presence of JNK-associated leucine zipper protein (JLP). The loss of JLP resulted in Foxk1 induction, leading to N-cadherin downregulation and compromised cell viability. We propose a novel axis consisting of JLP/Foxk1/N-cadherin in shaping the EMT program and suggest JLP as the checkpoint of the EMT continuum during renal fibrosis progression.

Complete mitochondrial genome of Penicillidia jenynsii (Diptera: Hippoboscoidea: Nycteribiidae) and phylogenetic relationship.

In recent years, bat-associated pathogens, such as 2019 novel coronavirus, have been ravaging the world, and ectoparasites of bats have received increasing attention. Penicillidia jenynsii is a member of the family Nycteribiidae which is a group of specialized ectoparasites of bats. In this study, the complete mitochondrial genome of P. jenynsii was sequenced for the first time and a comprehensive phylogenetic analysis of the superfamily Hippoboscoidea was conducted. The complete mitochondrial genome of P. jenynsii is 16 165 base pairs (bp) in size, including 13 protein-coding genes (PCGs), 22 transfer RNA genes, 2 ribosomal RNA genes and 1 control region. The phylogenetic analysis based on 13 PCGs of the superfamily Hippoboscoidea known from the NCBI supported the monophyly of the family Nycteribiidae, and the family Nycteribiidae was a sister group with the family Streblidae. This study not only provided molecular data for the identification of P. jenynsii, but also provided a reference for the phylogenetic analysis of the superfamily Hippoboscoidea.