Mu opioid receptor activation enhances regulator of G protein signaling 4 association with the mu opioid receptor/G protein complex in a GTP‐dependent manner
Abstract
The interaction of Regulator of G protein Signaling 4 (RGS4) with the rat mu opioid receptor (MOR)/G protein complex was investigated. Solubilized MOR from rat brain membranes was immunoprecipitated in the presence of RGS4 with antibodies against the N‐terminus of MOR (anti‐MOR10–70). Activation of MOR with [D‐Ala2, N‐Me‐Phe4, Gly5‐ol] enkephalin (DAMGO) during immunoprecipitation caused a 150% increase in Goα and a 50% increase in RGS4 in the pellet. When 10 μM GTP was included with DAMGO, there was an additional 72% increase in RGS4 co‐immunoprecipitating with MOR (p = 0.003). Guanosine 5′‐O‐(3‐thiotriphosphate) (GTPγS) increased the amount of co‐precipitating RGS4 by 93% (compared to DAMGO alone, p = 0.008), and the inclusion of GTPγS caused the ratio of MOR to RGS4 to be 1 : 1 (31 fmoles : 28 fmoles, respectively). GTPγS also increased the association of endogenous RGS4 with MOR. In His6RGS4/Ni2+‐NTA agarose pull down experiments, 0.3 μM GTPγS tripled the binding of Goα to His6RGS4, whereas the addition of 100 μM GDP blocked this effect. Importantly, activation of solubilized MOR with DAMGO in the presence of 100 μM GDP and 0.3 μM GTPγS increased Goα binding to His6RGS4/Ni2+‐NTA agarose (p = 0.001).
Document Details
- Document Type
- Pub Defense Publication
- Publication Date
- Jul 16, 2015
- Source ID
- 10.1111/jnc.13222
Entities
People
- Alicia Tamara Crowder
- Brian M Cox
- Rema Santhappan
- Shawn Gouty
- Thomas E. Cote
Organizations
- Uniformed Services University of the Health Sciences