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. Author manuscript; available in PMC: 2016 May 1.
Published in final edited form as: Cell Signal. 2015 Feb 12;27(5):908–922. doi: 10.1016/j.cellsig.2015.02.003

Figure 5. UCR1C does not alter global PKA activity.

Figure 5

(A) Diagram of A kinase activity reporter (AKAR2.2), adapted from [64]. In AKAR2.2, PKA phosphorylation of the P sites results in binding to FHA, which induces the proximity of CFP and citrine to increase the FRET ratio. AKAR2.2 was used to report the overall PKA activity inside cells. HEK293T cells were co-transfected with AKAR2.2 and RFP-tagged UCR1C or RFP control plasmid. Twenty-four hours later, cells were plated on slides and imaged after another 24 hours. (B) Representative pseudo-color images of AKAR2.2 FRET ratio change in response to adenylyl cyclase activator Forskolin (20 μM) stimulation and subsequent PKA inhibition using H89 (20 μM). (C) Time-course of average AKAR FRET ratio. Cells with similar RFP expression intensity were selected for analysis. Data are expressed as mean ± s.e.m.. Black squares represent cells expressing RFP vector (n=12 cells), open diamonds represent cells expressing RFP-UCR1C (n=12 cells).