As shown in the section on GPCR dimerization, it is generally believed that GPCRs have to reach the cell surface to be fully functional. However, I will show in the following pages that this is not necessarily the case! One invaluable aspect of studying the ghrelin receptor, was the discovery that it is a constitutively-active receptor. That means that the ghrelin receptor has some activity even in the absence of exogenous ghrelin as the normal stimulus. We used this property of the ghrelin receptor to isolate subcellular fractions of cells transfected to over-express GHS-R1a and see if we could detect endogenous cell signalling activity.
On the next few pages you will see the methods used to isolate subcellular fractions of cells transfected to express GHS-R1a and/or GHS-R1b. Because the ghrelin receptor constitutively activates the mitogen-activated protein kinase (MAPK) known as ERK1/2, we used a Western Blot to detect activated ERK1/2 in the different subcellular fractions.
From the following work we discovered that the ghrelin receptor was capable of signalling from subcellular fractions associated with the endoplasmic reticulum (ER). Determining the consequence of this cell signalling activity in the ER is something we could not pursue with the tools and resources available, so this is the end of my contribution to the story of ghrelin receptor signalling.