research highlights CheMIC aL BIoL oGY IMa GING Cross-linking with suFex super-long single-molecule chemistry tracking Wang, N. et al. J. Am. Chem. Soc. 140, 4995–4999 Tsunoyama, T. A. et al. Nat. Chem. Biol. 14, (2018). 497–506 (2018). Biocompatible, bio-orthogonal chemical Single-molecule tracking can provide reactions such as the Staudinger ligation important insight into protein dynamics, have proven to be useful methods for regulation and function. However, biochemical and imaging studies in living photophysical properties of fluorophores, cells, as well as for drug discovery. Wang such as bleaching and blinking, can make it et al. now add sulfur–fluoride exchange challenging to track labeled molecules over (SuFEx) chemistry to this growing toolbox. extended periods. Although methods for An unnatural amino acid, fluorosulfate--l reducing photobleaching and photoblinking tyrosine (FSY), can be incorporated into are known, they can be incompatible with proteins in bacterial and mammalian cells live imaging because of their toxicity. via expression of a novel, specifically evolved Tsunoyama et al. addressed this challenge tRNA synthetase–tRNA pair. Incorporated by treating cells with low concentrations of FSY selectively reacts with lysine, histidine dissolved oxygen along with a reducing-plus- and tyrosine residues in close proximity, oxidizing system to suppress both bleaching generating a covalent cross-link either within and blinking. They demonstrate that the or between proteins. Importantly, FSY has approach works for numerous commonly low cytotoxicity, which the authors attribute used organic dyes and has only minor effects to the low reactivity of aryl fluorosulfates on cells. Using their approach, they extended inside cells. They used FSY and SuFEx the duration of particle tracking from chemistry to capture and analyze the protein ~10 seconds to ~7 minutes at video rate, while complex between Escherichia coli Afb and maintaining high localization precision. They Z protein, as well as the complex between used their approach to study the behavior of PAPS reductase and thioredoxin. AD integrins at focal adhesions. RS https://doi.org/10.1038/s41592-018-0024-0 https://doi.org/10.1038/s41592-018-0026-y Help your results GeNoMICs GeNoMICs stand out. t issue-specific oncogene rBP census Huang, R. et al. Proc. Natl. Acad. Sci. USA 115, screens Try our expert editing E3879–E3887 (2018). Sack, L. M. et al. Cell 173, 499–514 (2018). service today. Post-transcriptional gene regulation is in Some genes can lead to cancer when large part driven by the proteins associated mutated, whereas others drive cancer with RNAs, and thus it is of great interest as a result of copy-number increases. to establish a comprehensive catalog of Sack et al. generated a library of almost RNA-binding proteins (RBPs). Huang et al. 30,000 barcoded human open reading developed CARIC (click chemistry–assisted frames to screen for genes that either RNA interactome capture) for unbiased stimulate or suppress proliferation when RBP profiling. The method begins with overexpressed. Their lentiviral screening metabolic labeling of RNA to incorporate vectors allowed for inducible expression and an alkyne-containing uridine analog (EU) tagging, as well as quantitative detection. The Language Editing • Scientiﬁ c Editing and a photoactivatable thio-uridine (4SU). researchers carried out screens in mammary, Upon UV light exposure, proteins bound fibroblast and pancreatic cell lines, and Formerly known as NPG Language Editing to RNAs are cross-linked to 4SU, and found that around 10% of genes regulate and MSC Scientiﬁ c Editing azide–biotin is clicked onto EU to provide a proliferation, often in a highly tissue-specific handle by which the RNA can be enriched. manner. Candidate genes from the screen After digestion of the RNA and mass- were enriched for known oncogenes and spectrometry-based protein identification, tumor-suppressor genes, and tissue-specific the researchers obtained a list of nearly drivers were typically associated with the authorservices.springernature.com 600 RBPs from HeLa cells that included tumor tissue of origin. Many of the candidate 130 proteins not known to associate genes are not commonly mutated in cancers with RNA. CARIC can be applied to but are associated with tissue-specific focal any cell type and will enrich RBPs acros s somatic copy-number alterations. TN all RNAs, independent of their https://doi.org/10.1038/s41592-018-0027-x polyadenylation status. NR Allison Doerr, Tal Nawy, Nicole Rusk, https://doi.org/10.1038/s41592-018-0025-z Rita Strack and Nina Vogt Nature Methods | VOL 15 | JUNE 2018 | 403–409 | www.nature.com/naturemethods © 2018 Nature America Inc., part of Springer Nature. All rights reserved.
Nature Methods – Springer Journals
Published: May 31, 2018
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