Binding of Filamin to the C-terminal Tail of the Calcitonin Receptor Controls Recycling

Abstract

Many G protein-coupled receptors undergo endocytosis, but the mechanisms involved in endocytic sorting and recycling remain to be fully elucidated. We found that the G protein-coupled calcitonin receptor (CTR) undergoes tonic internalization and accumulates within the cell. Using a fluorescence loss in photobleaching assay, we classified these vesicles functionally as recycling vesicles. In a two-hybrid screening, we found that the actin-binding protein filamin interacted with the C-terminal tail of the CTR. The degradation of the receptor was profoundly increased in the absence of filamin or the CTR-filamin interaction. The absence of filamin was also associated with a marked decrease in recycling of the receptor from the endosomes to the cell surface. In contrast, calcitonin-induced inhibition of spontaneous filamin proteolysis was associated with increased recycling of the receptor to the cell surface and decreased degradation of the CTR, suggesting an important role for filamin in the endocytic sorting and recycling of the internalized CTR.

Chronic morphine treatment up-regulates mu opioid receptor binding in cells lacking filamin A

“…Filamin A is known to bind to various membrane and signaling molecules, see reviews (Stossel et al, 2001;van der Flier and Sonnenberg, 2001)}. More recently different laboratories, including our own, demonstrated that filamin A associates with various GPCRs, such as the D2 and D3 dopamine receptors (Li et al, 2000;Lin et al, 2001), the calcium-sensing receptor (CaR) (Awata et al, 2001;Hjalm et al, 2001), the metabotropic glutamate receptor type 7 (Enz, 2002), alpha1-adrenergic receptors (Zhang et al, 2004), calcitonin receptor (Seck et al, 2003) and the mu opioid receptor (MOP) (Onoprishvili et al, 2003). In some cases, such as the calcium sensing receptor and the MOP, the binding of filamin A is to the carboxyl tail, while for other receptors, such as the D2 and D3 dopamine receptors, the binding is to the third cytoplasmic loop.…”

Direct Interaction between the Actin-binding Protein Filamin-A and the Inwardly Rectifying Potassium Channel, Kir2.1

“…The reason for this is unclear, although the presence of a correctly organized submembranous actin cytoskeleton may be essential for the insertion or removal of proteins into or out of the plasma membrane. In support of this, binding of filamin to the carboxyl tail of the calcitonin receptor has been shown to be important in the recycling of the receptors back to the membrane after internalization (40).…”

“…We confirmed this finding by use of a double-tagged Kir2.1 construct that allows us to monitor surface expression as a fraction of total expression. A number of other filamin-interacting proteins including ␤-integrins (28), K v 4.2 voltage-gated potassium channels (8), D2 dopamine receptors (7), and calcitonin receptors (40) show markedly decreased levels of functional cell surface expression in the absence of filamin. The reason for this is unclear, although the presence of a correctly organized submembranous actin cytoskeleton may be essential for the insertion or removal of proteins into or out of the plasma membrane.…”

“…The majority of filamin-interacting proteins bind to a region just upstream of the dimerization site. The calcitonin receptor (40), the voltage-gated K ϩ channel K v 4.2 (8), SEK-1 (also known as MKK-4, a member of the stress-activated Jun kinase cascade (10)), TRAF-2 (tumor necrosis factor receptorassociated factor-2 (41)), the small GTPases Rac, Rho, CDC42, and RalA (9), protein kinase C␣ (11), and now Kir2.1 all bind in the region between repeats 20 and 24 that includes the second hinge region.…”

Filamin structure, function and mechanics: are altered filamin-mediated force responses associated with human disease?

“…FLNA is a substrate for multiple kinases (reviewed in Sarkisian et al 2008) depending on whether or not its actin bound (Kawamoto and Hidaka 1984), with its cross-linking activity itself regulated by phosphorylation (Cukier et al 2007). FLN is a substrate for the Ca 2+ -activated protease calpain (Gorlin et al 1990) and for the ASB2 ubiquitin ligase (Heuze et al 2008) which provide irreversible regulatory mechanisms for severing FLN cross-links in processes requiring membrane and cytoskeleton remodelling such as cell migration (Marzia et al 2006;Baldassarre et al 2009) and receptor cycling (Seck et al 2003). FLN's role in cell motility and spreading appears to be cell-type dependent.…”