Write a summary on this article and make sure to explain each one of the figures

Write a summary on this article. Given an example. It’s really important that the summary looks like example.Make sure to explain each one of the figures

Abstract
G protein-coupled receptors (GPCRs) are essential mediators of cellular signaling and important targets of drug action. Of the approximately 350 non-olfactory human GPCRs, more than 100 are
still considered “orphans” as their endogenous ligand(s) remain unknown. Here, we describe a unique open-source resource that provides the capacity to interrogate the druggable human GPCR-
ome via a G protein-independent β-arrestin recruitment assay. We validate this unique platform at more than 120 non-orphan human GPCR targets, demonstrate its utility for discovering new
ligands for orphan human GPCRs, and describe a method (PRESTO-TANGO; Parallel Receptor-ome Expression and Screening via Transcriptional Output – TANGO) for the simultaneous and parallel interrogation of the entire human GPCR-ome.G protein-coupled receptors (GPCRs) are proteins with seven transmembrane helices capable of transducing a wide variety of extracellular stimuli into intracellular signals, mediated by G proteins of four groups, Gs, Gi, G12 or 13 or Gq, as well as arrestins and other effectors1. The human genome encodes more than 350 different non-olfactory GPCRs, as well as a similar number of olfactory GPCRs2–4. In addition to their roles as signal
transducers, GPCRs are the targets for more than one-third of currently prescribed medications5, 6. Approximately one-third of the non-olfactory GPCRs in the human genome
are “orphan” GPCRs, i.e., their endogenous or natural ligands are unknown2–4, while many more are inadequately interrogated with respect to their ligands. Thus, much of the
druggable GPCR-ome – like other drug target families such as the kinome7, represents ‘dark
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subject always to the full Conditions of use:http://www.nature.com/authors/editorial_policies/license.html#termsCorresponding author: Bryan L. Roth, 4072 Genetic Medicine, UNC-CH School of Medicine, Chapel Hill, NC 27599-7365; Phone:
919-966-7535; Fax: 919-843-5788; bryan_roth@med.unc.edu.WKK, MFS and X-P H: Contributed equally
AUTHOR CONTRIBUTIONS: BLR and WKK conceived the general approach; WKK designed the constructs; WKK, FS, KL and
XPH executed and analyzed validation, profiling and confirmatory assays; JDM and PMG validated assays; NS assisted with high
content microscopy; FS designed, executed and analyzed the simultaneous profiling strategy; BLR, WKK, FS, KL and XPH wrote the paper; BLR was responsible for the overall strategy.
HHS Public AccessAuthor manuscriptNat Struct Mol Biol. Author manuscript; available in PMC 2015 November 01.
Published in final edited form as:
Nat Struct Mol Biol. 2015 May ; 22(5): 362–369. doi:10.1038/nsmb.3014.
Author Manuscript Author Manuscript Author Manuscript Author Manuscript matter’ of the human genome. As many of these sparsely annotated GPCRs will likely represent fruitful future therapeutic targets, identifying drug-like chemical leads for the
entire family of druggable GPCRs represents a major goal for chemical biology.
Unfortunately, interrogating the druggable GPCR-ome en masse in a parallel and simultaneous fashion is currently technologically and economically unfeasible.The difficulty in screening the entire druggable GPCR-ome in parallel is due mainly to the
inherent diversity of signal transduction cascades, rendering attempts at parallel profiling challenging. Thus, for instance, functional assays for the identification of agonists at orphan
and other sparsely annotated GPCRs have typically used readouts that depend on the native or forced1 coupling of GPCRs with G proteins, e.g., Gs, Gi, Gq or G12 or 138–14.
Unfortunately, these approaches are not well suited for the parallel and simultaneous genome-wide interrogation of the druggable GPCR-ome1. Alternatively, measurement of G-protein independent β-arrestin recruitment provides a feasible and universal assay platform, since nearly all tested GPCRs can induce arrestin translocation15, 16 (Supplementary Table
1). A wide variety of approaches has been described to quantify GPCR-β-arrestin interactions, including high content screening (HCS)17, bioluminescence resonance energy transfer (BRET)18, enzyme complementation19, and transcriptional activation following arrestin translocation (TANGO)20, although none are routinely performed in a genome-wide, parallel manner. As we show here, the TANGO approach has a number of advantages for high throughput assays, including its independence from G protein coupling, generally high signal-to-background ratios, and its amplification of relatively small initial inputs into large readout signals. Independence from G protein coupling facilitates interrogation of orphan GPCRs, whose coupling partners are unknown. Some of the advantages of the TANGO assay might also be shared by other readout systems, including, for example,
assays for changes in impedance or dynamic mass redistribution (reviewed in 21) and indeed, arrestin recruitment may be part of the dynamic mass redistribution response measured in receptor-expressing cells responding to agonists, as suggested, but not directly shown, by the results of Hennen et al22. Our goal was to develop the TANGO assay into a platform that could encompass the entire druggable GPCR-ome. Although our assay does not differ significantly in terms of the general concept from that of Barnea et al.20, several notable changes including the design of the plasmid constructs and assay execution have
distinct advantages, as we describe below. We also demonstrate a method (PRESTO-TANGO; Parallel Receptor-ome Expression and Screening via Transcriptional Output-TANGO) that facilitates the rapid, efficacious, parallel, and simultaneous profiling of
biologically active compounds across essentially the entire human druggable GPCR-ome.
Additionally, we document how our approach leads to the facile identification of new
synthetic and naturally-occurring agonists for orphan GPCRs. Finally, as our platform is open-source, our methods and reagents are made freely available to the scientific community.
Kroeze et al. Page 2
Nat Struct Mol Biol. Author manuscript; available in PMC 2015 November 01.
Author Manuscript Author Manuscript Author Manuscript Author Manuscript

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Baird et al. – 2014 – HSF-1-mediated cytoskeletal integrity determines thermotolerance and life span
Discovered that thermotolerance is regulated by HSF-1 but not through upregulation of heat shock proteins as previously thought.
They authors used molecular biology techniques such as PCR, plasmid construction methods, etc. to show that truncated versions of the HSF-1 gene have decreased transcriptional activity in C. elegans.
Fig 1. Only the full-length HSF-1 gene leads to heat shock protein expression, however both full length and truncated genes increase thermotolerance and life span.
Fig 2. HSF-1 regulates PAT-10 expression. Knock out of PAT-10 causes a decrease in thermotolerance and overexpression increases thermotolerance. The PAT-10 effect is independent of heat shock proteins.
Fig 3. Use fluorescence microscopy and GFP to assess the actin cytoskeleton. Loss of PAT-10 leads to a messed-up actin cytoskeleton when animals were heat stressed. Overexpression of PAT-10 protected the actin cytoskeleton from heat stress. Overexpression also delayed the decrease in F actin that is normally seen with aging.
Fig 4. Using pharmacology they showed that disrupting the actin cytoskeleton in human cells also impaired their thermotolerance.
The transcription factor HSF-1 affects thermotolerance by controlling the expression of PAT-10, which is required to keep the actin cytoskeleton from collapsing. HSF-1 regulation of PAT-10 is independent of its heat shock protein responsibilities. Over-expressing PAT-10 increased thermotolerance and life span. Pathways that activate PAT-10 should be explored further because the pathway would make a good target for the treatment of age-related diseases.