Many physiological and pathophysiological processes are regulated by cAMP. Different therapies directly or indirectly influence the cellular concentration of this second messenger. A wide variety of receptors either activates or inhibits adenylate cyclases in order to induce proper physiological responses. A key event in this signalling system is the direct and dynamic interaction of Gαi1 subunits with adenylate cyclases. We established a FRET-based assay between G-protein subunits and AC5 (type 5 adenylate cyclase) and monitored receptor-stimulated interactions between Gαi1 and AC5 in single intact cells with high temporal resolution. We observed that FRET between Gαi1 and AC5 developed at much lower concentration of agonist compared with the overall Gi-protein activity resulting in a left-shift of the concentration–response curve by approximately one order of magnitude. Furthermore, Gi1-protein-mediated attenuation of AC5-dependent increases in cAMP occurred at comparable low concentrations of agonist. On analysing the dynamics we found the dissociation of the Gαi1 subunits and AC5 to occur significantly slower than the G-protein deactivation and to be insensitive to RGS4 (regulator of G-protein signalling type 4) expression. This led us to the conclusion that AC5, by binding active Gαi1, interferes with G-protein deactivation and reassembly and thereby might sensitize its own regulation.
Skip Nav Destination
Article navigation
September 2013
-
Cover Image
Cover Image
- PDF Icon PDF LinkFront Matter
- PDF Icon PDF LinkTable of Contents
- PDF Icon PDF LinkEditorial Board
Research Article|
August 29 2013
Dynamics of Gαi1 interaction with type 5 adenylate cyclase reveal the molecular basis for high sensitivity of Gi-mediated inhibition of cAMP production
Markus Milde;
Markus Milde
*Institute of Pharmacology and Clinical Pharmacy, Universität Marburg, 35032 Marburg, Germany
Search for other works by this author on:
Andreas Rinne;
Andreas Rinne
*Institute of Pharmacology and Clinical Pharmacy, Universität Marburg, 35032 Marburg, Germany
Search for other works by this author on:
Frank Wunder;
Frank Wunder
†Lead Discovery Wuppertal, Bayer HealthCare AG, Pharma Research Center, Aprather Weg 18a, 42096 Wuppertal, Germany
Search for other works by this author on:
Stefan Engelhardt;
Stefan Engelhardt
‡Institute of Pharmacology and Toxicology, Technische Universität München, 80802 Munich, Germany
Search for other works by this author on:
Moritz Bünemann
Moritz Bünemann
1
*Institute of Pharmacology and Clinical Pharmacy, Universität Marburg, 35032 Marburg, Germany
1To whom correspondence should be addressed (email [email protected]).
Search for other works by this author on:
Publisher: Portland Press Ltd
Received:
April 22 2013
Revision Received:
July 01 2013
Accepted:
July 10 2013
Accepted Manuscript online:
July 10 2013
Online ISSN: 1470-8728
Print ISSN: 0264-6021
© The Authors Journal compilation © 2013 Biochemical Society
2013
Biochem J (2013) 454 (3): 515–523.
Article history
Received:
April 22 2013
Revision Received:
July 01 2013
Accepted:
July 10 2013
Accepted Manuscript online:
July 10 2013
Citation
Markus Milde, Andreas Rinne, Frank Wunder, Stefan Engelhardt, Moritz Bünemann; Dynamics of Gαi1 interaction with type 5 adenylate cyclase reveal the molecular basis for high sensitivity of Gi-mediated inhibition of cAMP production. Biochem J 15 September 2013; 454 (3): 515–523. doi: https://doi.org/10.1042/BJ20130554
Download citation file:
Sign in
Don't already have an account? Register
Sign in to your personal account
You could not be signed in. Please check your email address / username and password and try again.
Could not validate captcha. Please try again.