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| Open AccessDissection of goadsporin biosynthesis by in vitro reconstitution leading to designer analogues expressed in vivo
Goadsporin is an antibacterial peptide highly customized by multiple post-translational modifying enzymes. Here, the authors dissect its biosynthetic pathway byin vitro reconstitution and use their insights to produce goadsporin analogues in vitro and in vivo.
- Taro Ozaki
- , Kona Yamashita
- & Hiroyasu Onaka
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| Open AccessDeep phenotyping unveils hidden traits and genetic relations in subtle mutants
Experimenter scoring of cellular imaging data can be biased. This study describes an automated and unbiased multidimensional phenotyping method that relies on machine learning and complex feature computation of imaging data, and identifies weak alleles affecting synapse morphology in live C. elegans.
- Adriana San-Miguel
- , Peri T. Kurshan
- & Hang Lu
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| Open AccessRapid construction of a whole-genome transposon insertion collection for Shewanella oneidensis by Knockout Sudoku
Knockout collections provide a valuable tool to explore gene function, yet are expensive and technically challenging to produce at a genome-wide scale. Here Baym et al. devise a cost-effective transposon-based method to quickly develop a knockout collection for the electroactive microbe Shewanella oneidensis.
- Michael Baym
- , Lev Shaket
- & Buz Barstow
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| Open AccessHuman genome-wide RNAi screen reveals host factors required for enterovirus 71 replication
Enterovirus 71 (EV71) infection causes a spectrum of symptoms including neurological disease. To improve our understanding of EV71-host interactions, Wu et al. here perform a genome-wide RNAi screen, which implicates cell cycle regulation and ER-associated degradation as important factors in EV71 replication.
- Kan Xing Wu
- , Patchara Phuektes
- & Justin Jang Hann Chu
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| Open AccessLarge-scale microfluidics providing high-resolution and high-throughput screening of Caenorhabditis elegans poly-glutamine aggregation model
Large-scale screens on whole animals could facilitate drug discovery, but are technically challenging. Here, Mondal et al. develop a microfluidic chip combined with an automated imaging platform that enables high-throughput, high-resolution screening of Caenorhabditis elegansdisease models.
- Sudip Mondal
- , Evan Hegarty
- & Adela Ben-Yakar
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| Open AccessIrreversible inhibitors of the 3C protease of Coxsackie virus through templated assembly of protein-binding fragments
Molecular fragments are useful tools in drug-discovery but they might be hard to identify due to their weak affinity to the targets. Here, the authors use a protein-templated assembly to design high affinity inhibitors of Coxsackie virus 3C protease, a pharmacological target against enteroviral infections.
- Daniel Becker
- , Zuzanna Kaczmarska
- & Jörg Rademann
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| Open AccessRapid construction of metabolite biosensors using domain-insertion profiling
In the construction of single fluorescent protein biosensors, selection of the insertion point of a fluorescent protein into a ligand-binding domain is a rate-limiting step. Here, the authors develop an unbiased, high-throughput approach, called domain insertion profiling with DNA sequencing (DIP-seq), to generate a novel trehalose biosensor.
- Dana C. Nadler
- , Stacy-Anne Morgan
- & David F. Savage
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| Open AccessChEC-seq kinetics discriminates transcription factor binding sites by DNA sequence and shape in vivo
In chromatin endogenous cleavage (ChEC), micrococcal nuclease (MNase) is fused to a protein of interest and its cleavage is thus targeted to specific genomic loci in vivo. Here, the authors show that time-resolved ChEC-seq (high-throughput sequencing after ChEC) can detect DNA shape patterns regardless of motif strength.
- Gabriel E. Zentner
- , Sivakanthan Kasinathan
- & Steven Henikoff
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High-throughput and quantitative assessment of enhancer activity in mammals by CapStarr-seq
Characterizing mammalian gene expression regulation by enhancer elements is complicated by the size and complexity of the genome. Here Vanhille et al.demonstrate CapStarr-Seq, a novel high-throughput method for assessing potential enhancers and deciphering the mechanisms regulating transcription
- Laurent Vanhille
- , Aurélien Griffon
- & Salvatore Spicuglia
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Quantitative high throughput screening using a primary human three-dimensional organotypic culture predicts in vivo efficacy
Tumour microenvironment affects the outcome of pharmacological anticancer treatments. Here, Kenny et al. show that organotypic cultures of ovarian cancer cells can recapitulate metastasis. They identify several new compounds that block cancer invasion and metastasis and improve survival in mouse models.
- Hilary A. Kenny
- , Madhu Lal-Nag
- & Ernst Lengyel
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High-throughput detection of miRNAs and gene-specific mRNA at the single-cell level by flow cytometry
Flow cytometry allows high-throughput analysis of multiple proteins in individual cells, but relies on availability of antibodies. Here Porichis et al.report a sensitive method for multi-parameter flow cytometric and imaging detection of proteins together with mRNA or miRNA at the single-cell level.
- Filippos Porichis
- , Meghan G. Hart
- & Daniel E. Kaufmann
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Suppression of the FOXM1 transcriptional programme via novel small molecule inhibition
Overexpression of the FOXM1 transcription factor occurs in several cancer and correlates with poor prognoses. Here, the authors identify a novel small molecule capable of displacing FOXM1 from its DNA consensus motif in vitro, displace it from target promoters and downregulate the expression of its target genes cancer cells.
- Michael V. Gormally
- , Thomas S. Dexheimer
- & Shankar Balasubramanian
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Functional high-throughput screening identifies the miR-15 microRNA family as cellular restriction factors for Salmonella infection
The functional role of microRNAs in the interplay between bacterial pathogens and host cells is not well defined. Here, using an image-based high-throughput screening approach, the authors identify a family of microRNAs that regulates Salmonellainfection and characterize their mechanism of action.
- Claire Maudet
- , Miguel Mano
- & Ana Eulalio
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High-throughput hyperdimensional vertebrate phenotyping
Large-scale screening of animal phenotypes requires automated detection and analysis of complex morphological information. Here, Yanik and colleagues present an imaging system based on optical projection tomography that generates micrometre-resolution 3D images of zebrafish larvae with within tens of seconds per animal.
- Carlos Pardo-Martin
- , Amin Allalou
- & Mehmet Fatih Yanik
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| Open AccessGenome-wide functional screening of miR-23b as a pleiotropic modulator suppressing cancer metastasis
microRNAs are known to be deregulated in cancer. Using a screen for microRNAs that alter cell migration, Zhanget al. show that mir-23b blocks cell migration in vitro and in vivoand is reduced in expression in human colon cancer, suggesting a therapeutic potential for this microRNA.
- Hanshuo Zhang
- , Yang Hao
- & Jianzhong Jeff Xi