Abstract：
:The human Ras superfamily of small GTPases controls essential cellular processes such as gene expression and cell proliferation. As their deregulation is widely associated with human cancer, small GTPases and their regulatory proteins have become increasingly attractive for the development of novel therapeutics. Classical methods to monitor GTPase activation include pulldown assays that limit the analysis of GTP-bound form of proteins from cell lysates. Alternatively, live-cell FRET biosensors may be used to study GTPase activation dynamics in response to stimuli, but these sensors often require further optimization for high-throughput applications. Here, we describe a cell-based approach that is suitable to monitor the modulation of small GTPase activity in a high-content analysis. The assay relies on a genetically encoded tripartite split-GFP (triSFP) system that we integrated in an optimized cellular model to monitor modulation of RhoA and RhoB GTPases. Our results indicate the robust response of the reporter, allowing the interrogation of inhibition and stimulation of Rho activity, and highlight potential applications of this method to discover novel modulators and regulators of small GTPases and related protein-binding domains.
journal_name
J Cell Scijournal_title
Journal of cell scienceauthors
Koraïchi F,Gence R,Bouchenot C,Grosjean S,Lajoie-Mazenc I,Favre G,Cabantous Sdoi
10.1242/jcs.210419subject
Has Abstractpub_date
2018-01-08 00:00:00issue
1eissn
0021-9533issn
1477-9137pii
jcs.210419journal_volume
131pub_type
杂志文章abstract：:In 1993, a Commentary in this journal described how a simple mechanical model of cell structure based on tensegrity architecture can help to explain how cell shape, movement and cytoskeletal mechanics are controlled, as well as how cells sense and respond to mechanical forces (J. Cell Sci. 104, 613-627). The cellular ...
journal_title：Journal of cell science
pub_type： 杂志文章,评审
doi：10.1242/jcs.00359
更新日期：2003-04-01 00:00:00
abstract：:The roles of P21-activated kinase (PAK) in the regulation of axon outgrowth downstream of extracellular matrix (ECM) proteins are poorly understood. Here we show that PAK1-3 and PIX are expressed in the developing spinal cord and differentially localize to point contacts and filopodial tips within motile growth cones....
journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.112607
更新日期：2013-03-01 00:00:00
abstract：:Integrins mediate cell adhesion and signal transduction at focal adhesions. Here we investigate the roles of integrin beta subunits in the regulation of actin cytoskeletal structure and the activities of Rho and Rac. The overexpression of beta3 integrin in Chinese hamster ovary cells enhances Rho activity and stress f...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：
更新日期：2002-05-15 00:00:00
abstract：:Myogenic differentiation in adult muscle is normally suppressed and can be activated by myogenic cues in a subset of activated satellite cells. The switch mechanism that turns myogenesis on and off is not defined. In the present study, we demonstrate that tissue inhibitor of metalloproteinase 3 (TIMP3), the endogenous...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.057620
更新日期：2010-09-01 00:00:00
abstract：:Zoospores of Phytophthora palmivora adhered to a plastic film surface were examined by electron microscopy. Three stages of adhesion were compared: (1) non-adhesive, unencysted zoospores, (2) adhered incipient cysts, and (3) adhered mature cysts. Thin sections of incipient cysts revealed cells attached to the film sur...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：
更新日期：1975-06-01 00:00:00
abstract：:A Sertoli-spermatid coculture model is described in which a large percentage (greater than 76%) of round spermatids remain viable for 48 h and bind to Sertoli cells. The effects of follicle-stimulating hormone (FSH) and testosterone on spermatid binding (expressed as the spermatid density; SD = the number of spermatid...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：
更新日期：1991-11-01 00:00:00
abstract：:The Rho-family proteins make up a major branch of the Ras superfamily of small GTPases. To date, 22 human genes encoding at least 25 proteins have been described. The best known 'classical' members are RhoA, Rac1 and Cdc42. Highly related isoforms of these three proteins have not been studied as intensively, in part b...
journal_title：Journal of cell science
pub_type： 杂志文章,评审
doi：10.1242/jcs.01118
更新日期：2004-03-15 00:00:00
abstract：:In Drosophila, telomere-capping proteins have the remarkable capacity to recognize chromosome ends in a sequence-independent manner. This epigenetic protection is essential to prevent catastrophic ligations of chromosome extremities. Interestingly, capping proteins occupy a large telomere chromatin domain of several k...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.167825
更新日期：2015-05-15 00:00:00
abstract：:Cultured human erythrocytes infected with Plasmodium falciparum were studied by freeze-fracture electron microscopy. Special emphasis was placed upon the formation of the membrane surface excrescences ('knobs') found on red cells containing mature parasites. Knobs were visualized as conoid projections of the protoplas...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：
更新日期：1986-03-01 00:00:00
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journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.091363
更新日期：2011-12-01 00:00:00
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journal_title：Journal of cell science
pub_type： 杂志文章
doi：
更新日期：1980-04-01 00:00:00
abstract：:When a Euglena gracilis in a Ca2+-containing medium is impaled with a microelectrode, the flagellum is instantly ejected. In a Ca2+-free medium to which 1 mM EGTA has been added, the flagellum remains attached to the organism, but it loses activity upon impalement. Externally added ATP at a concentration of 10 mM will...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：
更新日期：1977-02-01 00:00:00
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journal_title：Journal of cell science
pub_type： 杂志文章
doi：
更新日期：1978-10-01 00:00:00
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journal_title：Journal of cell science
pub_type： 杂志文章
doi：
更新日期：1976-11-01 00:00:00
abstract：:Antithrombin inhibits chemokine-induced migration of neutrophils by activating heparan sulfate proteoglycan-dependent signaling. Whether antithrombin affects migration of other types of leukocytes is not known. We investigated the effects of antithrombin on spontaneous and chemokine-triggered migration of lymphocytes ...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：
更新日期：2002-01-01 00:00:00
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journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.01524
更新日期：2004-12-01 00:00:00
abstract：:The notion that cell shape and spreading can regulate cell proliferation has evolved over several years, but only recently has this been linked to forces from within and upon the cell. This emerging area of mechanical signaling is proving to be wide-spread and important for all cell types. The microenvironment that su...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.067009
更新日期：2011-04-15 00:00:00
abstract：:Eukaryotic cells have developed complex machineries to distribute proteins and lipids from the Golgi complex. Contrary to what has originally been postulated, delivery of proteins to the cell surface is not a simple bulk flow process but involves sorting into distinct pathways from the trans-Golgi network. Here we des...
journal_title：Journal of cell science
pub_type： 杂志文章,评审
doi：
更新日期：1997-12-01 00:00:00
abstract：:It has long been established that neuronal growth cone navigation depends on changes in microtubule (MT) and F-actin architecture downstream of guidance cues. However, the mechanisms by which MTs and F-actin are dually coordinated remain a fundamentally unresolved question. Here, we report that the well-characterized ...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.224311
更新日期：2019-04-30 00:00:00
abstract：:Cyclic stretch is an artificial model of mechanical force loading, which induces the reorientation of vascular endothelial cells and their stress fibers in a direction perpendicular to the stretch axis. Rho family GTPases are crucial for cyclic-stretch-induced endothelial cell reorientation; however, the mechanism und...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.157503
更新日期：2015-05-01 00:00:00
abstract：:Here we report Drosophila Waharan (Wah), a 170-kD predominantly nuclear protein with two potential human homologues, as a newly identified regulator of endosomal trafficking. Wah is required for neuromuscular-junction development and muscle integrity. In muscles, knockdown of Wah caused novel accumulations of tightly ...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.060582
更新日期：2010-07-15 00:00:00
abstract：:A new aspect of research into the pathogenesis of cystic fibrosis (CF) is a genetics-based search for ;modifier genes' that may affect the severity of CF lung disease. Using an alternative, cell biological approach, we show that ITPK1 should be considered a modifier gene. ITPK1 synthesizes an intracellular signal, ino...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.02836
更新日期：2006-04-01 00:00:00
abstract：:The cell surface delivery of extracellular matrix (ECM) and integrins is fundamental for cell migration in wound healing and during cancer cell metastasis. This process is not only driven by several soluble NSF attachment protein (SNAP) receptor (SNARE) proteins, which are key players in vesicle transport at the cell ...
journal_title：Journal of cell science
pub_type： 杂志文章,评审
doi：10.1242/jcs.164459
更新日期：2015-03-15 00:00:00
abstract：:Many of the cilia- and flagella-specific integral membrane proteins identified to date function to sense the extracellular milieu, and there is considerable interest in defining pathways for targeting such proteins to these sensory organelles. The flagellar glucose transporter of Leishmania mexicana, LmxGT1, is target...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.103028
更新日期：2012-07-15 00:00:00
abstract：:The adaptor protein SETA/CIN85/Ruk is involved in regulating diverse signal transduction pathways, including the internalization of tyrosine kinase receptors via the Cbl ubiquitin ligases, and attenuating PI3K activity by interaction with its regulatory subunit. Here we present evidence for a new aspect of SETA functi...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.00522
更新日期：2003-07-15 00:00:00
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journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.03418
更新日期：2007-04-01 00:00:00
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journal_title：Journal of cell science
pub_type： 杂志文章
doi：
更新日期：1998-05-01 00:00:00
abstract：:Fe-S clusters act as co-factors of proteins with diverse functions, for example, in DNA repair. Downregulation of the cytosolic iron-sulfur protein assembly (CIA) machinery promotes genomic instability through the inactivation of multiple DNA repair pathways. Furthermore, CIA deficiencies are associated with so far un...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：10.1242/jcs.211433
更新日期：2018-06-27 00:00:00
abstract：:When expressed in pituitary AtT-20 cells, parotid proline-rich proteins enter the regulated pathway. Because the short N-terminal domain of a basic proline-rich protein is necessary for efficient export from the ER, it has not been possible to evaluate the role of this polypeptide segment as a sorting signal for regul...
journal_title：Journal of cell science
pub_type： 杂志文章
doi：
更新日期：1996-06-01 00:00:00
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journal_title：Journal of cell science
pub_type： 杂志文章
doi：
更新日期：1989-12-01 00:00:00
