GFP-cDNA Live & G-Protein Coupled Receptors
Dynamics of fluorescent cytoplasmic proteins upon activation of NK2 receptor

These web pages present the videos recorded during a live-imaging screen in HEK293 cells. This was aimed at identifying novel cytoplasmic proteins implicated in signaling pathways of G-protein coupled receptors, based on a translocation criterion (1).

The plasmids transfected in cells during the screen were from the GFP-tagged human full-length ORF plasmid collection and encoded proteins that localize in the cytoplasm (2-3). This plasmid library is composed of two vectors for each cDNA, one with the cDNA encoding CFP in frame 5’ of the ORF, the other with the cDNA encoding YFP in frame 3’ of the ORF.

An extensive study of REDD1, one of the hit proteins can be found in (4).

  1. Lecat S, Matthes HW, de Mey J, Pepperkok R, Simpson JC and Galzi JL. (2015) « A fluorescent live imaging screening assay based on translocation criteria identifies novel cytoplasmic proteins implicated in GPCR signalling pathways». Molecular and Cellular Proteomics. May;14(5):1385-99 First Published on March 10, 2015, doi:10.1074/mcp.M114.046698 http://www.mcponline.org/content/early/2015/03/10/mcp.M114.046698.full.pdf.
  2. Simpson JC, Wellenreuther R, Poustka A, Pepperkok R and Wiemann S (2000) “Systematic subcellular localization of novel proteins identified by large-scale cDNA sequencing”. EMBO Rep. 1(3):287-92. EMBL GFP-cDNA localization project, http://gfp-cdna.embl.de.
  3. Stadler C, Rexhepaj E, Singan VR, Murphy RF, Pepperkok R, Uhlén M, Simpson JC, and Lundberg E. (2013) « Immunofluorescence and fluorescent-protein tagging show high correlation for protein localization in mammalian cells.” Nat Methods. 4:315-23. doi: 10.1038/nmeth.2377.
  4. Michel G, Matthes HW, Hachet-Haas M, El Baghdadi K, de Mey J, Pepperkok R, Simpson JC, Galzi JL and Lecat S. (2014) “Plasma membrane translocation of REDD1 governed by GPCRs contributes to mTORC1 activation” Journal of Cell Science, 127:773-787. doi:10.1242/jcs.136432.

To get access to the list of the screened proteins, press « Show all ».

To refine the Search, use keywords such as : Name, full-Name, cDNA ID, Swissprot ID, biological process.

Show all
e.g WDR91, WD repeat-containing protein 91, AL136940, Q9H062, transcription
2 result(s)
Id Name Full Name Biological Process Clones cDNA ID Swissprot ID Endogenous localisation in Stadler et al. 2013 Fusion Orientation Localisation in Simpson et al. 2000 Localisation during screening Comments during screening
289 WDR91 WD repeat-containing protein 91 unknown hute1_24b4 AL136940 Q9H062 IF cyt/nuc not nucleoli/ Golgi CFP-ORF cyt cyt
290 WDR91 WD repeat-containing protein 91 unknown hute1_24b4 AL136940 Q9H062 IF cyt/nuc not nucleoli/ Golgi ORF-YFP cyt/ves cyt + PM sometimes +vesicles
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