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A sequence-verified collection of human transcription factors is reported. The authors used it in the enhanced yeast-one hybrid (eY1H) assay to map human gene regulatory networks. Also in this issue, Reece-Hoyes et al. describe the eY1H pipeline.
The authors describe the enhanced yeast one-hybrid platform for large-scale screening of protein-DNA interactions and test its performance by mapping Caenorhabditis elegans gene regulatory networks. Also in this issue, Hens et al. describe an alternative platform for this purpose and apply it to screen for transcription factor–DNA interactions in Drosophila melanogaster.
A fluorescent molecular tension sensor for spatially and temporally mapping the mechanical strain exerted by cell-surface receptors in living cells is described.
Described is a high-throughput yeast one-hybrid platform for mapping protein-DNA interactions and a sequence-verified clone collection of Drosophila transcription factors. Also in this issue, Reece-Hoyes et al. report enhanced yeast one-hybrid assays, an alternative system for large-scale protein-DNA interaction screens.
This paper describes a method to enrich for homozygous mutant mouse embryonic stem cells without an inherently selectable phenotype. It is used to construct a bank of homozygous and heterozygous mutant cells and should prove useful for cellular phenotypic screens.
A human embryonic stem cell line expressing a fluorescent reporter of cardiac differentiation is described. The authors use this tool to optimize differentiation methods and to identify cell-surface markers in the cardiac lineage.
Live-cell volumetric super-resolution imaging with 120-nm lateral and 360-nm axial resolution using structured-illumination microscopy at speeds of up to 5 s per cell volume over >50 time points captures fine cellular dynamics using only low illumination intensities.
The combination of light-sheet microscopy and localization-based super-resolution imaging allows deep subdiffraction resolution imaging in thick scattering specimens as demonstrated by three-dimensional super-resolution imaging of proteins in live 150-μm-diameter cell spheroids.
A simple episomal fluorescent reporter for flow cytometric enrichment of cells with zinc-finger nuclease– or TALE nuclease–induced genomic modifications is described.
Owing to a lack of tools and a lack of a consensus sequence for O-glycosylation, studies of the O-glycoproteome have been few and far between, despite the biological importance of O-glycosylation. This method to analyze O-glycan attachment sites to proteins using glycoengineered cell lines with simplified, homogenous O-glycoproteomes should facilitate future O-glycoproteomics studies.
A computational approach for analysis of gene expression in heterogeneous samples of varying composition is presented. The authors used it to study expression in brain samples from humans with Huntington's disease.
Soft hydrogel microwell arrays with controllably variable stiffness are spotted with proteins of interest, individually or in combination. The system permits in vitro study of the biophysical and biochemical aspects of the stem-cell niche.
A triple-stage mass spectrometry (MS3)-based method is used to remove ratio interference, resulting in accurate, large-scale, multiplexed quantitative proteomics measurements using isobaric labeling. Also in this issue, Wenger et al. provide a different solution to the same problem.
A mass spectrometry instrument control method—QuantMode—allows accurate, large-scale, multiplexed quantitative proteomics measurements using isobaric tagging by removing the problem of precursor interference. Also in this issue, Ting et al. provide a different solution to the same problem.
The question of whether transcription factories containing RNA polymerases exist has been controversial, owing to the fact that they have not been isolated previously. Now, a method to carefully isolate these complexes and analyze their proteomes by mass spectrometry is described.
Reported is the robust directed differentiation of mouse epiblast stem cells to oligodendrocyte precursor cells, which then differentiate into myelinating oligodendrocytes in vitro and in vivo. The system should prove useful for basic research and for drug screens.
Implementation of pair correlation analysis for photoactivated localization microscopy allows quantitative analysis of protein clustering in the plasma membrane, revealing the degree to which different perturbations alter protein arrangements.
A comparison of the proteomes and phosphoproteomes of four human embryonic stem cell lines and four induced pluripotent stem cell lines is reported, revealing subtle differences in these cell types at the protein level. Also introduced is the Stem Cell-Omics Repository (SCOR), a database of quantitative information for transcripts, proteins and post-translational modifications.
