Biological techniques

  • Article
    | Open Access

    Conventional upconversion nanoparticles (UCNPs) cannot activate multiple neuron populations independently using optogenetics. Here the authors report trichromatic UCNPs with excitation-specific luminescence to allow activation of three distinct neuronal populations in the brain of awake mice.

    • Xuan Liu
    • , Heming Chen
    •  & Fan Zhang
  • Article
    | Open Access

    Current high-throughput single-cell transcriptomic methods are incompatible with paraformaldehyde, a common cell fixation technique. Here the authors present FD-seq, a method for droplet-based RNA sequencing of paraformaldehyde-fixed, stained and sorted single cells.

    • Hoang Van Phan
    • , Michiel van Gent
    •  & Savaş Tay
  • Article
    | Open Access

    Practical implementation of genetic circuits is difficult due to low predictability and time-intensive troubleshooting. Here the authors present Cyberloop, which interfaces a computer with single cells to enable cell-in-the-loop testing and optimization of circuit designs before they are built.

    • Sant Kumar
    • , Marc Rullan
    •  & Mustafa Khammash
  • Article
    | Open Access

    Although the first dissection of the human ovary dates back to the 17th century, its characterization is still limited. Here, the authors have unraveled a unique biophysical and topological phenotype of reproductive-age tissue, bridging biophysics and female fertility and providing a blueprint for the artificial ovary.

    • Emna Ouni
    • , Alexis Peaucelle
    •  & Christiani A. Amorim
  • Article
    | Open Access

    Determining the quality of localisation microscopy images is currently challenging. Here the authors report use of the Haar wavelet kernel analysis (HAWK) Method for the Assessment of Nanoscopy, termed HAWKMAN, to assess the reliability of localisation information.

    • Richard J. Marsh
    • , Ishan Costello
    •  & Susan Cox
  • Article
    | Open Access

    Humans are exposed to millions of chemicals but mass spectrometry (MS)-based targeted biomonitoring assays are usually limited to a few hundred known hazards. Here, the authors develop a workflow for MS-based untargeted exposome profiling of known and unidentified environmental chemicals.

    • Xin Hu
    • , Douglas I. Walker
    •  & Dean P. Jones
  • Article
    | Open Access

    Anaplastic large cell lymphoma (ALCL) is an aggressive T-cell lymphoma often with poor prognosis. To identify genes defining ALCL cell state and dependencies, the authors here characterize ALCL-specific super-enhancers and describe the BATF3/IL-2R−module as a therapeutic opportunity for ALCL.

    • Huan-Chang Liang
    • , Mariantonia Costanza
    •  & Olaf Merkel
  • Article
    | Open Access

    Actin-based structures in cells and tissues are built and maintained through a poorly understood balance between assembly and disassembly. Here, our findings provide insights into how factors known to promote these opposing effects dynamically integrate to shape cells and tissue systems.

    • Elena E. Grintsevich
    • , Giasuddin Ahmed
    •  & Jonathan R. Terman
  • Article
    | Open Access

    The ever-growing need for highly functional, compact, and integrated microfluidic devices often incurs lengthy and expensive manufacturing processes. Here, authors introduce a generalized 3D printing process that enables fast parallel fabrication of miniaturized, high resolution 3D components.

    • Jose L. Sanchez Noriega
    • , Nicholas A. Chartrand
    •  & Gregory P. Nordin
  • Article
    | Open Access

    Rapid identification of host genes essential for virus replication may expedite the generation of therapeutic interventions. Here the authors generate mutant clonal intestinal organoids for 19 host genes previously implicated in coronavirus biology and identify the cell surface protease TMPRSS2 as a potential therapeutic target.

    • Joep Beumer
    • , Maarten H. Geurts
    •  & Hans Clevers
  • Article
    | Open Access

    Here, the authors present a hierarchical disorder model for the analysis of disorder in both crystal and cryo-EM structures. They apply their approach to several structures of three proteins, including SARS-CoV-2 proteins, and discuss mechanistic and dynamical implications.

    • Nicholas M. Pearce
    •  & Piet Gros
  • Article
    | Open Access

    Faster, higher throughput antibody engineering methods are needed. Here the authors present CeVICA, a cell-free nanobody engineering platform using ribosome display and computational clustering analysis for in vitro selection; they use this to develop nanobodies against the RBD of SARS-CoV-2 spike protein.

    • Xun Chen
    • , Matteo Gentili
    •  & Aviv Regev
  • Article
    | Open Access

    Transcription in archaea is known to be regulated through the recruitment of RNA polymerase to promoters. Here, the authors show that the archaeon Saccharolobus solfataricus regulates transcription globally through a rate-limiting promoter-proximal elongation step.

    • Fabian Blombach
    • , Thomas Fouqueau
    •  & Finn Werner
  • Article
    | Open Access

    GABAA receptors (GABAARs) cause inhibition in the brain by functioning as heteropentamers formed from multiple subunit types. Here, the authors demonstrate that receptors incorporating β3 subunits can spontaneously gate, which is modulated by protein kinases and neurosteroids to affect tonic inhibition.

    • Craig A. Sexton
    • , Reka Penzinger
    •  & Trevor G. Smart
  • Article
    | Open Access

    Detection of the tumour boundary in prostate cancer is required for surgery. Here the authors present a fluorescent molecular rotor probe to target a prostate cancer marker, prostate-specific membrane antigen (PSMA), which they use in a xenograft mouse model to show it can be used for in vivo imaging.

    • Jingming Zhang
    • , Anastasia Rakhimbekova
    •  & Xing Yang
  • Article
    | Open Access

    Molecular tethers physically bridge transport vesicles to their target membranes as a prerequisite step for fusion. Here the authors control vesicle tethering using optogenetic approaches to study the interplay between vesicle tethering and fusion.

    • Seong J. An
    • , Felix Rivera-Molina
    •  & Derek Toomre
  • Article
    | Open Access

    Here, Broszeit et al. show that circulating A/H3N2 viruses have evolved binding specificity to α2,6-sialosides on extended LacNAc moieties and therefore cannot agglutinate erythrocytes. Applying glycan remodeling allows to install functional receptors on erythrocytes and promotes identification of newly circulating variants to facilitate vaccine design.

    • Frederik Broszeit
    • , Rosanne J. van Beek
    •  & Geert-Jan Boons
  • Article
    | Open Access

    Circularised nanodiscs (cNDs) are able to stabilise large lipid bilayer patches and are used for structural and functional studies. Current techniques to build cNDs have numerous steps and low yields; here the authors report a single step construction method using the SpyCatcher-SpyTag system.

    • Shanwen Zhang
    • , Qian Ren
    •  & Huan Bao
  • Article
    | Open Access

    Humans are exposed to many xenobiotic chemicals, but identification of low abundance xenobiotic exposures is limited by a lack of authentic standards for xenobiotic metabolites. Here the authors develop methods for enzymatic generation of diverse xenobiotic metabolites for use with high-resolution mass spectrometry for biology-based chemical identification.

    • Ken H. Liu
    • , Choon M. Lee
    •  & Dean P. Jones
  • Article
    | Open Access

    Photoacoustic imaging is limited by a lack of contrast agents which can enable combined molecular and physiological imaging at depth. Here the authors address these limitations by developing and validating a contrast agent based on targeted liposomes loaded with J-aggregated indocyanine green dye.

    • Cayla A. Wood
    • , Sangheon Han
    •  & Richard R. Bouchard
  • Article
    | Open Access

    Photosynthetic Complex I (PS-CI) is proposed to couple ferredoxin oxidation and plastoquinone reduction to proton pumping across thylakoid membranes. Here the authors determine the reduction potentials of the iron-sulphur clusters of PS-CI and thus the bioenergetics of the electron transfer relay.

    • Katherine H. Richardson
    • , John J. Wright
    •  & Maxie M. Roessler
  • Article
    | Open Access

    The Spike protein of SARS-CoV-2 has a C-terminal cytoplasmic tail. Here the authors show that this tail binds trafficking machinery via sequences that appear optimised to ensure that Spike accumulates at the site of viral budding in the Golgi but that some can also traffic to the cell surface to induce syncytia formation.

    • Jérôme Cattin-Ortolá
    • , Lawrence G. Welch
    •  & Sean Munro
  • Article
    | Open Access

    Light-driven micromotors can convert energy to motion in sub-millimeter dimensions. Here, the authors extend this concept and introduce reconfigurable micromachines with multiple components, driven by optoelectronic tweezers, and demonstrate new functionalities.

    • Shuailong Zhang
    • , Mohamed Elsayed
    •  & Aaron R. Wheeler
  • Article
    | Open Access

    The concentration of a biomarker in solution can be determined by counting single molecules. Here the authors report a digital immunoassay scheme with solid-state nanopore readout to quantify a target protein and use this to measure thyroid-stimulating hormone from human serum.

    • Liqun He
    • , Daniel R. Tessier
    •  & Vincent Tabard-Cossa
  • Article
    | Open Access

    Intricate color patterns are a defining aspect of morphological diversity in the Felidae. Here the authors apply morphological and single-cell gene expression analysis to fetal skin of domestic cats to identify when, where, and how, during fetal development, felid color patterns are established.

    • Christopher B. Kaelin
    • , Kelly A. McGowan
    •  & Gregory S. Barsh
  • Article
    | Open Access

    Elucidating specific effects of protein kinase Akt isoforms remains challenging. Here, the authors establish an Akt isoform-dependent cellular model system and use it, together with X-ray crystallography and structure-based ligand design, to develop isoform-selective covalent-allosteric Akt inhibitors

    • Lena Quambusch
    • , Laura Depta
    •  & Daniel Rauh
  • Article
    | Open Access

    Silicon probes for electrical recording from neurons usually have fewer wires than recording channels available to carry signals off the probe, which restricts the number of channels that can be recorded simultaneously. The authors propose to pool electrodes, using a single wire to serve many channels through a set of controllable switches.

    • Kyu Hyun Lee
    • , Yu-Li Ni
    •  & Markus Meister
  • Article
    | Open Access

    The authors present epiScanpy: a computational framework for the analysis of single-cell epigenomic data, both ATAC-seq and DNA methylation data, with examples for clustering, cell type identification, trajectory learning and atlas integration - and show its performance in distinguishing cell types.

    • Anna Danese
    • , Maria L. Richter
    •  & Maria Colomé-Tatché
  • Article
    | Open Access

    The signal-to-noise ratio is a key consideration when selecting a magnetic resonance imaging protocol. Thermal noise is major issue, especially in high resolution functional images. Here the authors introduce a method to suppress thermal noise in functional images without losses in spatial precision, increasing the signal-to-noise ratio.

    • Luca Vizioli
    • , Steen Moeller
    •  & Kamil Uğurbil
  • Article
    | Open Access

    There is a pressing need to develop representative organ-like platforms recapitulating complex in vivo phenotypes to study human development and disease in vitro. Here the authors present a method to generate human heart organoids by self-assembly using pluripotent stem cells, compare these to age-matched fetal cardiac tissues and recreate a model of pregestational diabetes.

    • Yonatan R. Lewis-Israeli
    • , Aaron H. Wasserman
    •  & Aitor Aguirre
  • Article
    | Open Access

    Previous approaches to derive embryoids either lack physiological morphology and signaling interactions, or are unconducive to model post-gastrulation development. Here the authors use a high-throughput approach to induce mouse embryonic stem cells into epiblast-like aggregates, which are then co-cultured with mouse trophoblast stem cell aggregates, to yield embryoids with axial morphogenesis and anterior development.

    • Mehmet U. Girgin
    • , Nicolas Broguiere
    •  & Matthias P. Lutolf
  • Article
    | Open Access

    The PUF RNAbinding domain has an eight-repeat structure with the ability to bind an eight-base RNA sequence. Here the authors generate a comprehensive library of variants of a PUF domain to find an RNA binding code and design PUF domains that recognize an arbitrary RNA sequence.

    • Wei Zhou
    • , Daniel Melamed
    •  & Stanley Fields
  • Article
    | Open Access

    Micro-crystal electron diffraction (MicroED) has shown great potential for the structure determination of crystals that are too small for X-ray diffraction but MicroED sample preparation remains challenging. Here, the authors present Preassis, a pressure-assisted method for the preparation of MicroED specimens and demonstrate that Preassis can be applied to a wide range of protein crystal suspensions with low and high viscosities, as well as those with low crystal concentrations.

    • Jingjing Zhao
    • , Hongyi Xu
    •  & Xiaodong Zou
  • Article
    | Open Access

    The prognosis of castration-resistant prostate cancers remains dismal, but accurate preclinical models can lead to effective therapies. Here the Melbourne Urological Research Alliance establish prostate cancer patient-derived xenografts, use the tumors for organoids and single-cell RNA-seq, and show the efficacy of PARP inhibitor combination treatments.

    • Gail P. Risbridger
    • , Ashlee K. Clark
    •  & Renea A. Taylor
  • Article
    | Open Access

    Proximity biotinylation is a powerful tool to profile interactomes, but it requires genetic engineering of the target protein. Here, the authors develop a proximity biotinylation enzyme that can be directed to the target using antibodies, enabling interactome profiling of endogenous proteins or PTMs.

    • Irene Santos-Barriopedro
    • , Guido van Mierlo
    •  & Michiel Vermeulen
  • Article
    | Open Access

    Mesomelic dysplasia, a severe shortening and bending of the limb, has been linked to rearrangements in the HoxD cluster in humans and mice. Here the authors engineer a 1 Mb inversion including the HoxD gene cluster and use this model to provide a mechanistic framework to understand and unify the molecular origins of human mesomelic dysplasia associated with 2q31.

    • Christopher Chase Bolt
    • , Lucille Lopez-Delisle
    •  & Denis Duboule
  • Article
    | Open Access

    Tendon self-renewal occurs rarely and reconstructive surgery comes with significant limitations. Here the authors present an induced pluripotent stem cell-based method to generate tenocytes, analyze their developmental trajectory using scRNA-seq, and demonstrate their contribution to motor function recovery after Achilles tendon injury via engraftment and paracrine effects.

    • Taiki Nakajima
    • , Akihiro Nakahata
    •  & Makoto Ikeya
  • Article
    | Open Access

    Existing methods for multimeric protein complex quantification in single cells suffer from limited selectivity and sensitivity. Here the authors report Single-cell protein Interaction Fractionation Through Electrophoresis and immunoassay Readout (SIFTER) and use this to probe the effects of cellular stress.

    • Julea Vlassakis
    • , Louise L. Hansen
    •  & Amy E. Herr