Targeting the splicing factor NONO inhibits GBM progression through GPX1 intron retention

Background: Splicing factors are essential for nascent pre-mRNA processing and critical in cancer progression, suggesting that proteins with splicing functions represent potential molecular targets for cancer therapy. Here, we investigate the role of splicing factors in glioblastoma multiforme (GBM) progression and the possibility of targeting them for the treatment of the disease. Methods: The TCGA and CGGA public databases were used to screen for differentially expressed mRNA splicing factors. Immunohistochemistry and qRT-PCR were used to analyze the expression of non-POU domain-containing octamer-binding protein (NONO), a Drosophila behavior human splicing (DBHS) protein. Knockdown/overexpression of NONO with siRNA and lentiviral expression constructs was used to examine cell growth, apoptosis, and invasion in GBM cells. RNA sequencing was used to identify potential downstream molecular targets of NONO. RIP-PCR and RNA pulldown were used to determine the interaction between NONO and pre-mRNA. JC-1 staining and the seahorse assay were performed to assess redox homeostasis. Results: Expression of NONO was increased in GBM samples and associated with poor survival in patients (P = 0.04). Knockdown of NONO suppressed GBM growth, and overexpression of NONO promoted GBM tumorigenesis in vitro and in vivo. RNA sequencing-based transcriptomic profiling confirmed that knockdown of NONO in U251 and P3 cells resulted in global intron retention of pre-mRNA and led to abnormal splicing of specific pre-mRNAs for GPX1 and CCN1. NONO bound to a consensus motif in the intron of GPX1 pre-mRNA in association with another DBHS protein family member, PSPC1. Knockdown of NONO impaired tumor growth, invasion, and redox homeostasis through aberrant splicing of GPX1. Finally, Auranofin, a small molecule inhibitor of NONO, suppressed GBM tumor growth in an orthotopic xenograft model in mice. Conclusions: We demonstrated that intron retention was a critical alternative RNA splicing event to occur in GBM progression, and that NONO was a key regulator of mRNA splicing in GBM. Targeting NONO represents a novel, potential therapeutic strategy for GBM treatment.publishedVersio

A targeted next-generation sequencing method for identifying clinically relevant mutation profiles in lung adenocarcinoma

Molecular profiling of lung cancer has become essential for prediction of an individual’s response to targeted therapies. Next-generation sequencing (NGS) is a promising technique for routine diagnostics, but has not been sufficiently evaluated in terms of feasibility, reliability, cost and capacity with routine diagnostic formalin-fixed, paraffin-embedded (FFPE) materials. Here, we report the validation and application of a test based on Ion Proton technology for the rapid characterisation of single nucleotide variations (SNVs), short insertions and deletions (InDels), copy number variations (CNVs), and gene rearrangements in 145 genes with FFPE clinical specimens. The validation study, using 61 previously profiled clinical tumour samples, showed a concordance rate of 100% between results obtained by NGS and conventional test platforms. Analysis of tumour cell lines indicated reliable mutation detection in samples with 5% tumour content. Furthermore, application of the panel to 58 clinical cases, identified at least one actionable mutation in 43 cases, 1.4 times the number of actionable alterations detected by current diagnostic tests. We demonstrated that targeted NGS is a cost-effective and rapid platform to detect multiple mutations simultaneously in various genes with high reproducibility and sensitivity

Self correction of refractive error among young people in rural China: results of cross sectional investigation

Objective To compare outcomes between adjustable spectacles and conventional methods for refraction in young people

Perpustakaan Umum Malang Dengan Kombinasi Taman Vertikal Dan Ventilasi Untuk Perancangan Ruang Baca

Kualitas udara dalam ruang merupakan sebuah interaksi yang dapat berubah baiksecara konstan mau pun tidak yang diakibatkan oleh beberapa faktor yangmempengaruhi baik dari lingkungan luar mau pun lingkungan dalam. Salah saturuangan yang berpotensi tinggi untuk mengalami masalah polusi udara dalam ruangadalah ruang perpustakaan. Hal ini disebabkan oleh kondisi lingkungan eksternalseperti debu yang terbawa angin dan kondisi internal yaitu bakteri yang terbawa padabuku-buku lama yang dihirup oleh pelaku aktifitas perpustakaan. Dari faktor eksternal,salah satu penyebabnya ialah debu, tanah, dan polutan yang terbawa di udara masuk kedalam ruang perpustakaan. Pengoperasian sistem ventilasi bangunan berperan pentingdalam membawa udara masuk ke dalam ruangan. Salah satu strategi yang telahdisebutkan ialah penggunaan filter. Filter pada ventilasi berfungsi sebagai penyerappolusi yang terbawa angin luar masuk ke ruang dalam. Terdapat beberapa cara untukfiltrasi pada bangunan salah satunya adalah taman vertikal. Diharapkan penggunaankombinasi taman vertikal dan ventilasi dapat menjadi sumber penghawaan alami yangtetap memperhatikan kualitas udara dalam pada perpustakaan agar masalah buruknyakualitas udara ruang dalam pada perpustakaan dapat direduksi

Bacterial microcompartment-mediated ethanolamine metabolism in E. coli urinary tract infection

Urinary tract infections (UTIs) are common, in general caused by intestinal Uropathogenic E.coli (UPEC) ascending via the urethra. Microcompartment-mediated catabolism of ethanolamine, a host cell breakdown product, fuels competitive overgrowth of intestinal E. coli, both pathogenic enterohaemorrhagic E. coli and commensal strains. During UTI urease negative E. coli thrive, despite the comparative nutrient limitation in urine. The role of ethanolamine as a potential nutrient source during UTI is understudied. We evaluated the role of metabolism of ethanolamine as a potential nitrogen and carbon source for UPEC in the urinary tract. We analysed infected urine samples by culture, HPLC, qRT-PCR and genomic sequencing. Ethanolamine concentration in urine was comparable to the most abundant reported urinary amino acid D-serine. Transcription of the eut operon was detected in the majority of urine samples screened containing E. coli. All sequenced UPECs had conserved eut operons while metabolic genotypes previously associated with UTI (dsdCXA, metE) were mainly limited to phylogroup B2. In vitro ethanolamine was found to be utilised as a sole source of nitrogen by UPECs. Metabolism of ethanolamine in artificial urine medium (AUM) induced metabolosome formation and provided a growth advantage at the physiological levels found in urine. Interestingly, eutE (acetaldehyde dehydrogenase) was required for UPECs to utilise ethanolamine to gain a growth advantage in AUM, suggesting ethanolamine is also utilised as a carbon source. This data suggests urinary ethanolamine is a significant additional carbon and nitrogen source for infecting E. coli

Frequent alterations in cytoskeleton remodelling genes in primary and metastatic lung adenocarcinomas

The landscape of genetic alterations in lung adenocarcinoma derived from Asian patients is largely uncharacterized. Here we present an integrated genomic and transcriptomic analysis of 335 primary lung adenocarcinomas and 35 corresponding lymph node metastases from Chinese patients. Altogether 13 significantly mutated genes are identified, including the most commonly mutated gene TP53 and novel mutation targets such as RHPN2, GLI3 and MRC2. TP53 mutations are furthermore significantly enriched in tumours from patients harbouring metastases. Genes regulating cytoskeleton remodelling processes are also frequently altered, especially in metastatic samples, of which the high expression level of IQGAP3 is identified as a marker for poor prognosis. Our study represents the first large-scale sequencing effort on lung adenocarcinoma in Asian patients and provides a comprehensive mutational landscape for both primary and metastatic tumours. This may thus form a basis for personalized medical care and shed light on the molecular pathogenesis of metastatic lung adenocarcinoma

Modification of bacterial microcompartments with target biomolecules via post-translational SpyTagging

Bacterial microcompartments (BMCs) are proteinaceous organelle-like structures formed within bacteria, often encapsulating enzymes and cellular processes, in particular, allowing toxic intermediates to be shielded from the general cellular environment. Outside of their biological role they are of interest, through surface modification, as potential drug carriers and polyvalent antigen display scaffolds. Here we use a post-translational modification approach, using copper free click chemistry, to attach a SpyTag to a target protein molecule for attachment to a specific SpyCatcher modified BMC shell protein. We demonstrate that a post-translationally SpyTagged material can react with a SpyCatcher modified BMC and show its presence on the surface of BMCs, enabling future investigation of these structures as polyvalent antigen display scaffolds for vaccine development. This post-translational ‘click’ methodology overcomes the necessity to genetically encode the SpyTag, avoids any potential reduction in expression yield and expands the scope of SpyTag/SpyCatcher vaccine scaffolds to form peptide epitope vaccines and small molecule delivery agents

A Systematic Analysis on DNA Methylation and the Expression of Both mRNA and microRNA in Bladder Cancer

Background: DNA methylation aberration and microRNA (miRNA) deregulation have been observed in many types of cancers. A systematic study of methylome and transcriptome in bladder urothelial carcinoma has never been reported. Methodology/Principal Findings: The DNA methylation was profiled by modified methylation-specific digital karyotyping (MMSDK) and the expression of mRNAs and miRNAs was analyzed by digital gene expression (DGE) sequencing in tumors and matched normal adjacent tissues obtained from 9 bladder urothelial carcinoma patients. We found that a set of significantly enriched pathways disrupted in bladder urothelial carcinoma primarily related to "neurogenesis" and "cell differentiation" by integrated analysis of -omics data. Furthermore, we identified an intriguing collection of cancer-related genes that were deregulated at the levels of DNA methylation and mRNA expression, and we validated several of these genes (HIC1, SLIT2, RASAL1, and KRT17) by Bisulfite Sequencing PCR and Reverse Transcription qPCR in a panel of 33 bladder cancer samples. Conclusions/Significance: We characterized the profiles between methylome and transcriptome in bladder urothelial carcinoma, identified a set of significantly enriched key pathways, and screened four aberrantly methylated and expressed genes. Conclusively, our findings shed light on a new avenue for basic bladder cancer research