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| Open AccessImproving the generalizability of protein-ligand binding predictions with AI-Bind
State-of-the-art machine learning models in drug discovery fail to reliably predict the binding properties of poorly annotated proteins and small molecules. Here, the authors present AI-Bind, a machine learning pipeline to improve generalizability and interpretability of binding predictions.
- Ayan Chatterjee
- , Robin Walters
- & Giulia Menichetti
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Article
| Open AccessSafe drugs with high potential to block malaria transmission revealed by a spleen-mimetic screening
Authors propose their splenic mimetic filtration method, microsphiltration, and utilise this approach in a drug-screen, to identify compounds that induce a stiffening effect on Plasmodium falciparum-infected erythrocytes. They proceed to assess safety and tolerability of one identified compound in a phase I clinical trial.
- Mario Carucci
- , Julien Duez
- & Pierre Buffet
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Article
| Open AccessA Vaccinia-based system for directed evolution of GPCRs in mammalian cells
G protein-coupled receptors are a major class of drug targets. Here, the authors develop a method whereby their biophysical and functional properties can be altered through directed evolution in mammalian cells, leading to variants exhibiting features such as high stability and expression, or increased allosteric coupling.
- Christoph Klenk
- , Maria Scrivens
- & Andreas Plückthun
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Article
| Open AccessTrio-pharmacophore DNA-encoded chemical library for simultaneous selection of fragments and linkers
Dual-pharmacophore DNA-Encoded Libraries (DELs) can generate large libraries, but linker optimisation is challenging. Here, the authors report a trio-pharmacophore DEL (T-DEL) for both de novo fragment identification and linker optimization of known fragment pairs.
- Meiying Cui
- , Dzung Nguyen
- & Yixin Zhang
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Article
| Open Access1,2,4,5-Tetrazine-tethered probes for fluorogenically imaging superoxide in live cells with ultrahigh specificity
Specific detection of cellular superoxide is challenging. Here, the authors designed 1,2,4,5-tetrazine based fluorogenic probes for specific and sensitive imaging of superoxide, and applied them in high throughput screening of modulators of oxidative stress.
- Xuefeng Jiang
- , Min Li
- & Xin Li
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Article
| Open AccessNon-falciparum malaria infection and IgG seroprevalence among children under 15 years in Nigeria, 2018
Nigeria has a high burden of malaria which mostly results from Plasmodium falciparum infection, but other species are also endemic. Here, the authors estimate the prevalence of active infection and previous exposure to P malariae, P ovale, and P vivax in children in Nigeria in 2018 and determine risk factors.
- Camelia Herman
- , Colleen M. Leonard
- & Eric Rogier
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Article
| Open AccessWidespread perturbation of ETS factor binding sites in cancer
Few cancer drivers in non-coding regions have been identified so far. Here, the authors develop a transcription factor-aware burden test to predict non-coding variants and analyze the impact on transcription factor binding - especially ETS factors - as well as their impact on transcriptional activity.
- Sebastian Carrasco Pro
- , Heather Hook
- & Juan Ignacio Fuxman Bass
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Article
| Open AccessEvolution of protease activation and specificity via alpha-2-macroglobulin-mediated covalent capture
Custom proteases find applications as therapeutics, in research and in biotechnological applications. Here, the authors establish a protease selection system based on bacterial alpha-2-macroglobulin protease inhibitors and evolve staphylococcal proteases for increased activity and altered specificity.
- Philipp Knyphausen
- , Mariana Rangel Pereira
- & Florian Hollfelder
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Article
| Open AccessModeling CRISPR-Cas13d on-target and off-target effects using machine learning approaches
Application of CRISPR-Cas13d is limited by the inability to predict on- and off-targets. Here the authors perform CRISPR-Cas13d proliferation screens followed by modeling of Cas13d on- and off-targets; they design a deep learning model, DeepCas13, to predict the on-target activity of a gRNA.
- Xiaolong Cheng
- , Zexu Li
- & Wei Li
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Article
| Open AccessTwo distinct binding modes provide the RNA-binding protein RbFox with extraordinary sequence specificity
Here the authors show that the RRM of RbFox accomplishes extraordinary sequence specificity by employing functionally and structurally distinct binding modes - one for its cognate RNA and one for all non-cognate RNAs.
- Xuan Ye
- , Wen Yang
- & Fan Yang
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Article
| Open AccessDiscovery of synthetic lethal interactions from large-scale pan-cancer perturbation screens
Synthetic lethality can be used to identify potential drug targets in cancer based on simultaneous inactivation of two genes through genetic aberrations and gene silencing. Here, the authors develop a statistical framework to identify synthetic lethal pairs from large scale perturbation screens across multiple cancer types.
- Sumana Srivatsa
- , Hesam Montazeri
- & Niko Beerenwinkel
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Article
| Open AccessHigh-throughput screening of caterpillars as a platform to study host–microbe interactions and enteric immunity
Here, combining diagnostic imaging modalities and in vivo assays, Windfelder and colleagues established tobacco hornworm larvae Manduca sexta as an alternative high-throughput platform to study the innate immunity of the gut and host-pathogen interactions. Using the platform, the authors identify mediators of gut inflammation, differentiate pathogens from gut mutualist bacteria, and demonstrate pharmacological interventions.
- Anton G. Windfelder
- , Frank H. H. Müller
- & Ulrich Flögel
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Article
| Open AccessMultiplexed and reproducible high content screening of live and fixed cells using Dye Drop
It is currently difficult to perform accurate single-cell assays in 384-well plates. Here the authors report Dye Drop which uses sequential density displacement and microscopy for multi-step assays on cells, and use this to collect single-cell dose-response data for small molecules in breast cancer cells.
- Caitlin E. Mills
- , Kartik Subramanian
- & Peter K. Sorger
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Article
| Open AccessMaprotiline restores ER homeostasis and rescues neurodegeneration via Histamine Receptor H1 inhibition in retinal ganglion cells
ER stress is associated with various neurodegenerative diseases. This study found that FDA approved drug, maprotiline, inhibits histamine receptor H1-mediated ER stress and provides significant neuroprotection in mouse glaucoma model.
- Wei Chen
- , Pingting Liu
- & Yang Hu
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Article
| Open AccessUse of a glycomics array to establish the anti-carbohydrate antibody repertoire in type 1 diabetes
Type I diabetes is characterized by autoantibodies directed against protein or non-protein self-antigens. Here the authors profile glycan reactive anti-carbohydrate antibodies (ACA) in a longitudinal and cross-sectional childhood diabetes cohort and associate clusters of ACA with disease progression.
- Paul M. H. Tran
- , Fran Dong
- & Sharad Purohit
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Article
| Open AccessHigh-throughput small molecule screen identifies inhibitors of microsporidia invasion and proliferation in C. elegans
Microsporidia are fungal-related intracellular parasites that infect animals and humans. Here, Murareanu et al. develop a high-throughput screening method using the nematode C. elegans as a host, and identify several compounds that inhibit microsporidia infection through different mechanisms.
- Brandon M. Murareanu
- , Noelle V. Antao
- & Aaron W. Reinke
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Article
| Open AccessHigh-throughput mutagenesis identifies mutations and RNA-binding proteins controlling CD19 splicing and CART-19 therapy resistance
Multiple alternative splicing events in CD19 mRNA have been associated with resistance/relapse to CD19 CAR-T therapy in patients with B cell malignancies. Here, by combining patient data and a high-throughput mutagenesis screen, the authors identify single point mutations and RNA-binding proteins that can control CD19 splicing and be associated with CD19 CAR-T therapy resistance.
- Mariela Cortés-López
- , Laura Schulz
- & Julian König
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Article
| Open AccessTraject3d allows label-free identification of distinct co-occurring phenotypes within 3D culture by live imaging
There are currently a lack of tools to detect heterogeneity in 3D cultures. Here the authors report Traject3d as a framework to identify heterogeneous states in 3D culture and to understand how these give rise to distinct phenotypes using label-free multi-day time-lapse imaging.
- Eva C. Freckmann
- , Emma Sandilands
- & David M. Bryant
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Article
| Open AccessRational design of a sensitivity-enhanced tracer for discovering efficient APC–Asef inhibitors
The adenomatous polyposis coli (APC)–Asef protein interaction is essential for colorectal cancer metastasis. Here, the authors present the rational design of a sensitivity-enhanced tracer for fluorescence polarization assays, enabling them to discover more efficient APC–Asef interaction inhibitors.
- Jie Zhong
- , Yuegui Guo
- & Jian Zhang
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Article
| Open AccessTP53-dependent toxicity of CRISPR/Cas9 cuts is differential across genomic loci and can confound genetic screening
Toxicity of CRISPR/Cas9 induced DNA breaks depends on their repair mechanism, and on the chromatin environment at the cut site. Here the authors show that edits in active genes or regulatory elements can incur a higher toxicity via a TP53-dependent mechanism.
- Miguel M. Álvarez
- , Josep Biayna
- & Fran Supek
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Article
| Open AccessCombi-seq for multiplexed transcriptome-based profiling of drug combinations using deterministic barcoding in single-cell droplets
Current screens to assess tumour drug resistance require a large amount of material, normally not available from patients. Here the authors report CombiSeq, a scalable microfluidic workflow to screen hundreds of drug combinations in picoliter-size droplets using transcriptome changes as a readout.
- L. Mathur
- , B. Szalai
- & C. A. Merten
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Article
| Open AccessLow-cost anti-mycobacterial drug discovery using engineered E. coli
Whole-cell screening for Mycobacterium tuberculosis inhibitors is complicated by the pathogen’s slow growth and biocontainment requirements. Here the authors develop engineered E. coli as a synthetic biology tool to express and screen metabolic targets from Mycobacterium tuberculosis.
- Nadine Bongaerts
- , Zainab Edoo
- & Edwin H. Wintermute
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Article
| Open AccessSynthesis and direct assay of large macrocycle diversities by combinatorial late-stage modification at picomole scale
Macrocycles have potential as therapeutics, but their libraries are currently not large enough for high-throughput screening. Here, the authors show a combinatorial approach to generate a library of almost 20’000 macrocycles by conjugating carboxylic-acid fragments to macrocyclic scaffolds, identifying nanomolar inhibitors against thrombin and binders of MDM2.
- Sevan Habeshian
- , Manuel Leonardo Merz
- & Christian Heinis
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Article
| Open AccessSimple synthesis of massively parallel RNA microarrays via enzymatic conversion from DNA microarrays
RNA microarrays have many potential applications, but are difficult to produce. Here, the AUs present a method for converting commercial, customizable DNA microarrays into RNA microarrays using an accessible three-step process involving primer photocrosslinking, extension, and template degradation.
- Erika Schaudy
- , Kathrin Hölz
- & Mark M. Somoza
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Article
| Open AccessPervasive translation of circular RNAs driven by short IRES-like elements
Unbiased screen of random sequences identified many short IRES-like elements to drive circular RNA translation and hundreds of rolling circle translation events, suggesting a pervasive cap-independent translation in human transcriptome.
- Xiaojuan Fan
- , Yun Yang
- & Zefeng Wang
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Article
| Open AccessTargeting fungal membrane homeostasis with imidazopyrazoindoles impairs azole resistance and biofilm formation
There is an urgent need for novel strategies to combat life-threatening fungal infections. Here, Revie et al. identify a compound that induces alterations in fungal membranes, synergizes with azole antifungals against the pathogen Candida albicans, and inhibits virulence traits and biofilm formation in animal models of infection.
- Nicole M. Revie
- , Kali R. Iyer
- & Leah E. Cowen
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Article
| Open AccessA high-throughput multiparameter screen for accelerated development and optimization of soluble genetically encoded fluorescent biosensors
Fluorescent biosensors are important tools for studying cellular metabolism, but development and optimization are challenging. Koveal et al. present a high-throughput multiparameter screen for sensor performance, and used it to generate LiLac, a high-performance, quantitative lactate sensor.
- Dorothy Koveal
- , Paul C. Rosen
- & Gary Yellen
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Article
| Open AccessMachine learning-coupled combinatorial mutagenesis enables resource-efficient engineering of CRISPR-Cas9 genome editor activities
Screening combinatorial mutants is too massive for wet-lab experiment alone. Here the authors present a machine learning-coupled combinatorial mutagenesis approach to vastly reduce experimental burden for engineering Cas9 genome editing enzymes.
- Dawn G. L. Thean
- , Hoi Yee Chu
- & Alan S. L. Wong
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Article
| Open AccessIntegrated profiling of human pancreatic cancer organoids reveals chromatin accessibility features associated with drug sensitivity
The chromatin accessibility landscape and gene regulatory network of pancreatic cancer has not been fully characterised. Here, the authors perform multi-omics analysis of 84 pancreatic cancer organoid lines and reveal gene regulatory networks and distinct molecular subtypes.
- Xiaohan Shi
- , Yunguang Li
- & Dong Gao
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Article
| Open AccessA loss-of-adhesion CRISPR-Cas9 screening platform to identify cell adhesion-regulatory proteins and signaling pathways
Targeting integrin-mediated retention of malignant B cells in their protective microenvironment is an efficacious treatment for lymphoma and leukemia. Here, the authors present an unbiased loss-of-adhesion CRISPR screening method, identifying therapeutic targets for these B-cell malignancies.
- Martin F. M. de Rooij
- , Yvonne J. Thus
- & Marcel Spaargaren
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Article
| Open AccessIntegrative multi-omics and drug response profiling of childhood acute lymphoblastic leukemia cell lines
Childhood acute lymphoblastic leukemia is characterised by a range of genetic aberrations. Here, the authors use multi-omics profiling of ALL cell lines to connect molecular phenotypes and drug responses to provide an interactive resource of drug sensitivity.
- Isabelle Rose Leo
- , Luay Aswad
- & Rozbeh Jafari
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Article
| Open AccessIntegrating deep learning and unbiased automated high-content screening to identify complex disease signatures in human fibroblasts
By coupling robotic cell culture systems with artificial intelligence–powered image analysis, Schiff et al. identify previously unseen characteristics of Parkinson’s disease in patient skin cells that distinguish them from healthy controls.
- Lauren Schiff
- , Bianca Migliori
- & Bjarki Johannesson
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Article
| Open AccessTranscriptional repression of estrogen receptor alpha by YAP reveals the Hippo pathway as therapeutic target for ER+ breast cancer
Hippo signalling is reported to be required for proper ESR1 expression. Here the authors reveal that the transcriptional repression of ESR1 is via LATS-YAP-TEAD-VGLL3 axis and the epigenetic regulation of ESR1 super enhancer in ER + breast cancer.
- Shenghong Ma
- , Tracy Tang
- & Kun-Liang Guan
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Article
| Open AccessKansl1 haploinsufficiency impairs autophagosome-lysosome fusion and links autophagic dysfunction with Koolen-de Vries syndrome in mice
Here the authors show that the Koolen-de Vries syndrome associated gene KANSL1 modulates autophagosome-lysosome fusion via transcriptional regulation of autophagosomal gene Syntaxin17, and that 13-cis retinoic acid can reverses mitophagic defects and neurobehavioural abnormalities of mice lacking Kansl1.
- Ting Li
- , Dingyi Lu
- & Xin Pan
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Article
| Open AccessDihydroceramide- and ceramide-profiling provides insights into human cardiometabolic disease etiology
Among lipid classes, ceramides are linked to impaired cardiometabolic health. Here the authors report the association of specific ceramides and dihydroceramides on the risk of developing type 2 diabetes and or cardiovascular disease in a prospective population cohort.
- C. Wittenbecher
- , R. Cuadrat
- & M. B. Schulze
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Article
| Open AccessDroplet-based screening of phosphate transfer catalysis reveals how epistasis shapes MAP kinase interactions with substrates
Here, the authors use a droplet-based screen for phosphate transfer catalysis, testing variants of the human protein kinase MKK1 for its ability to activate its downstream target ERK2. Data reveal a flexible motif in the MKK1 docking domain that promotes efficient activation of ERK2, and suggest epistasis between the residues within that sequence.
- Remkes A. Scheele
- , Laurens H. Lindenburg
- & Florian Hollfelder
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Article
| Open AccessMicrobiome differential abundance methods produce different results across 38 datasets
Many microbiome differential abundance methods are available, but it lacks systematic comparison among them. Here, the authors compare the performance of 14 differential abundance testing methods on 38 16S rRNA gene datasets with two sample groups, and show ALDEx2 and ANCOM-II produce the most consistent results.
- Jacob T. Nearing
- , Gavin M. Douglas
- & Morgan G. I. Langille
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Article
| Open AccessIntegrating gene expression and clinical data to identify drug repurposing candidates for hyperlipidemia and hypertension
Prioritizing drug repurposing candidates for downstream studies remains challenging. Here, the authors present a high-throughput approach to identify and validate drug repurposing candidates, integrating human gene expression, drug perturbation, and clinical data from publicly available resources.
- Patrick Wu
- , QiPing Feng
- & Wei-Qi Wei
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Article
| Open AccessCapillary flow experiments for thermodynamic and kinetic characterization of protein liquid-liquid phase separation
Methods to quantitatively study liquid-liquid phase separation (LLPS) of proteins are lacking. Here the authors report Capillary flow experiments (Capflex) for the quantification of key LLPS parameters; they study Ddx4, the RP3 peptide and the aberrant liquid-to-solid phase transition of α-synuclein.
- Emil G. P. Stender
- , Soumik Ray
- & Alexander K. Buell
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Article
| Open AccessDiscovery of indole-modified aptamers for highly specific recognition of protein glycoforms
Glycosylation is an abundant form of post-translational modification. Here the authors present a generalizable workflow for the selection of indole-modified aptamers that can recognize protein glycoforms with high specificity.
- Alex M. Yoshikawa
- , Alexandra Rangel
- & H. Tom Soh
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Article
| Open AccessBioinformatic and cell-based tools for pooled CRISPR knockout screening in mosquitos
Forward genetic approaches such as CRISPR screens are powerful ways to identify essential genes and those that influence host-pathogen interactions. Here the authors design a bioinformatics portal for sgRNA design and a recombination-mediated cassette system for delivery into mosquito cell lines.
- Raghuvir Viswanatha
- , Enzo Mameli
- & Norbert Perrimon
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Article
| Open AccessMapping enzyme catalysis with metabolic biosensing
The testing of engineered enzymes represents a bottleneck. Here the authors report a screening method combining microfluidics and mass spectrometry, to map the catalysis of a mutated enzyme, characterise the range of products generated and recover the sequences of variants with desired activities.
- Linfeng Xu
- , Kai-Chun Chang
- & Adam R. Abate
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Article
| Open AccessDiscovery of putative tumor suppressors from CRISPR screens reveals rewired lipid metabolism in acute myeloid leukemia cells
CRISPR-based knockout screens in cancer cells have suggested the existence of proliferation suppressor genes (PSG). Here, the authors develop an approach to systematically identify them, and reveal a PSG module involved in fatty acid synthesis and tumour suppression in acute myeloid leukemia cell lines.
- W. Frank Lenoir
- , Micaela Morgado
- & Traver Hart
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Article
| Open AccessPhysics-informed deep learning characterizes morphodynamics of Asian soybean rust disease
Deep learning (DL) can be used to automatically extract complex features from dynamic systems. Here, the authors combine high-content imaging, DL and mechanistic models to extract and explain drug-induced morphological changes in the growth of the fungus responsible for Asian soybean rust.
- Henry Cavanagh
- , Andreas Mosbach
- & Robert G. Endres
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Article
| Open AccessMassively parallel interrogation of protein fragment secretability using SECRiFY reveals features influencing secretory system transit
The exact protein features that control passage through the eukaryotic secretory system remain largely unknown. Here the authors report SECRiFY which they use to evaluate the secretory potential of polypeptides on a proteome-wide scale in yeast, revealing a role for flexibility and intrinsic disorder.
- Morgane Boone
- , Pathmanaban Ramasamy
- & Nico Callewaert
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Article
| Open AccessSingle-cell normalization and association testing unifying CRISPR screen and gene co-expression analyses with Normalisr
Normalisr removes technical bias in single-cell RNA-seq and detects gene differential and coexpression accurately and efficiently. It also infers gene regulatory and co-expression networks from conventional and CRISPR screen single-cell RNA-seq datasets.
- Lingfei Wang
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Article
| Open AccessImprovement of a synthetic live bacterial therapeutic for phenylketonuria with biosensor-enabled enzyme engineering
PKU patients have elevated phenylalanine levels which can result in neurological impairment. Here the authors utilize biosensor-based ultra-high-throughput screening to optimize PAL activity in a synthetic biotic platform for improved in vivo performance.
- Kristin J. Adolfsen
- , Isolde Callihan
- & Vincent M. Isabella
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Article
| Open AccessEngineering digitizer circuits for chemical and genetic screens in human cells
Cell-based transcriptional reporters are an invaluable part of highthroughput screening, but many such reporters have weak or transient responses. Here, the authors describe a digitizer circuit for amplifying reporter activity, increasing sensitivity, and retaining memory of pathway activation.
- Nicole M. Wong
- , Elizabeth Frias
- & Wilson W. Wong
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Article
| Open AccessGAK and PRKCD are positive regulators of PRKN-independent mitophagy
The mechanisms involved in programmed or damage-induced removal of mitochondria by mitophagy remain elusive. Here the authors use an siRNA library to screen lipid-binding proteins, and identify the kinases GAK and PRKCD as positive regulators of PRKN-independent mitophagy.
- Michael J. Munson
- , Benan J. Mathai
- & Anne Simonsen