title: Akt signaling dynamics in individual cells.
abstract: The protein kinase Akt (for which there are three isoforms) is a key intracellular mediator of many biological processes, yet knowledge of Akt signaling dynamics is limited. Here, we have constructed a fluorescent reporter molecule in a lentiviral delivery system to assess Akt kinase activity at the single cell level. The reporter, a fusion between a modified FoxO1 transcription factor and clover, a green fluorescent protein, rapidly translocates from the nucleus to the cytoplasm in response to Akt stimulation. Because of its long half-life and the intensity of clover fluorescence, the sensor provides a robust readout that can be tracked for days under a range of biological conditions. Using this reporter, we find that stimulation of Akt activity by IGF-I is encoded into stable and reproducible analog responses at the population level, but that single cell signaling outcomes are variable. This reporter, which provides a simple and dynamic measure of Akt activity, should be compatible with many cell types and experimental platforms, and thus opens the door to new insights into how Akt regulates its biological responses.
authors.affiliation: Department of Biochemistry and Molecular Biology, Oregon Health & Science University, Portland, OR 97239, USA.
authors.forename: Sean M
authors.initials: SM
authors.lastname: Gross
authors.affiliation: Department of Biochemistry and Molecular Biology, Oregon Health & Science University, Portland, OR 97239, USA Department of Biomedical Sciences, Paul L. Foster School of Medicine, Texas Tech Health University Health Sciences Center, El Paso, TX 79905, USA peter.rotwein@ttuhsc.edu.
authors.forename: Peter
authors.initials: P
authors.lastname: Rotwein
mesh_terms: D000818:Animals; D002460:Cell Line; D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008954:Models, Biological; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D000071161:Forkhead Box Protein O1; D051858:Forkhead Transcription Factors; C467200:Foxo1 protein, mouse; D051057:Proto-Oncogene Proteins c-akt
keywords: Akt; IGF-I; Live-cell imaging; Signaling dynamics; Signaling pathways