GPCRGProteins

A large number of GPCRs localize in membrane rafts/caveolae either before (e.g., 6-adrenergic receptor sub-types) or after treatment with agonists (e.g., certain muscarinic cholinergic and bradykinin receptors, Table 1). Agonists of certain GPCRs can promote the entry of receptors into raft/caveolar domains, while for other

Table 1 Examples of G-protein-coupled receptors that localize in lipid raft/caveo-lae prior to ("pre-agonist") and/or after ("post-agonist") treatment with agonists

Pre-agonist Post-agonist

Table 1 Examples of G-protein-coupled receptors that localize in lipid raft/caveo-lae prior to ("pre-agonist") and/or after ("post-agonist") treatment with agonists

Pre-agonist Post-agonist

Endothelin (ETA, ETB)

+

+

Cholecystokinin (CCK)

+

Muscarinic cholinergic

+

+

Bradykinin (B1 and B2)

+

+

Lysophosphatidic acid (LPA-1)

+

Angiotensin II (AT-1)

+

ß1- and ß2- adrenergic

+

P2Y (PY1)

+

Adenosine A1

+

+

Sphingosine 1-phosphate (EDG-1)

+

+

Smoothened/patched

+

Serotonin (5HT2A)

+

Calcium-sensing receptor

+

^-adrenergic (a )

+

Chemokine CCR2

+

Metabotropic glutamate (mGlu1)

+

Gonadotrophin-releasing hormone (GnRH)

+

Oxytocin

+

Growth-hormone releasing-hormone

+

Dopamine (D1; D(1A) )

+

+

Mu-opioid receptor

+

Neurokinin 1

+

GPCRs, agonists promote the exit from these domains, perhaps as a prelude to receptor internalization/endocytosis and desensitization (Cohen et al. 2004; van Deurs et al. 2003). While certain GPCR internalize in response to agonist activation following phosphorylation by G-protein-receptor kinase, interaction with ß-arrestins and internalization via clathrin-coated pits, other GPCR are phosphorylated by protein kinase A (PKA) and then internalized from caveolae (Morris et al. 2004). The precise contribution of rafts/caveolae to GPCR desensitization and inter-nalization, in terms of the full-range of GPCR and cell types, has not yet been defined (Chini and Parenti 2004). The angiotensin-1 receptor is an example of a GPCR for which receptor-caveolin interaction appears to be important for sorting and delivery to the plasma membrane (Wyse et al. 2003). Other such examples have not been reported, although palmitoylation of a GPCR, for example, of the 5-HT1A receptor, can target it to rafts and facilitate receptor-mediated signal transduction (Renner et al. 2007).

As implied by the caveolin-signaling hypothesis, these microdomains bring "downstream" effectors and receptors together so as to facilitate receptor-, tissue- and cell-specific signal transduction (Cohen et al. 2004; Insel and Patel 2007; Ostrom and Insel 2004). The interaction of effectors with caveolins (via the CSD) or other caveo-lae-associated proteins are thought to be key to their targeting to caveolae. In addition, in response to stimulation, reversible palmitoylation may aid in caveolar localization of proteins by helping to regulate the movement of molecules into and out of caveolae (Lee et al. 2001; Parat and Fox 2001; Song et al. 1997).

Heterotrimeric G-proteins localize to caveolae and directly interact with caveolin (Insel et al. 2005; Ostrom and Insel 2004) in a manner that helps maintain Gaproteins in an inactive, GDP-bound state (Couet et al. 1997; Li et al. 1995, 1996; Murthy and Makhlouf 2000). Agonist stimulation can promote exchange of GTP for GDP and Ga redistribution to the cytoplasm, an effect blocked by the CSD (Murthy and Makhlouf 2000), suggesting that binding to the CSD regulates G-pro-tein function. Localization of Ga-proteins in caveolae may facilitate coupling to particular signaling pathways (Bhatnagar et al. 2004; Head et al. 2005; Iiri et al. 1996; Li et al. 1995; Oh and Schnitzer 2001). Certain heterotrimeric G-protein subunits, such as Gas, show a diffuse distribution on sucrose density gradients that are used to isolate caveolin-rich fractions; the Ga proteins can be detected in both caveolin-enriched (buoyant) and non-buoyant fractions (Cho et al. 2003; Cohen et al. 2004; Ostrom and Insel 2006). Heterotrimeric G-protein subunits can differentially localize to caveolae vs. membrane rafts (Head et al. 2005; Li et al. 1995; Oh and Schnitzer 2001). Palmitoylation enhances targeting of Ga subunits to caveolae (Song et al. 1997); Gßy can increase the palmitoylation of Ga, suggesting that this covalent modification may enhance interaction of caveolae and Ga (Iiri et al. 1996). Localization of Ga subunits in caveolae and interaction with the CSD is thus likely important for heterotrimeric G-protein signaling (Oh and Schnitzer 2001).

Certain small GTP-binding proteins (e.g., H-ras) also reside in caveolae, perhaps via palmitoylation of the C-terminal hypervariable region (CAAX motif) (Prior et al. 2001; Roy et al. 2005) and/or via interaction with the CSD (Li et al. 1995; Song et al. 1996). Activating mutations in H-Ras (e.g., G12V) prevent its interaction with caveolin and maintain an active conformation of the protein; such mutations are found in human cancers, suggesting that agents able to mimic this inhibitory action of caveolin have therapeutic potential in Ras-expressing cancers (Cavallo-Medved et al. 2005; Engelman et al. 1998).

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