The calcium sensing receptor (CaR) transduces local changes in extracellular calcium and metabolites (anrtino acids, polyamines) into intracellular signals, which include acute alterations in cell metabolism and secretion, and long term changes in cell proliferation and differentiation. CaR is critical to maintenance of systemic calcium levels, by controlling parathyroid PTH secretion and the balance of renal calcium excretion/resorption. Cell-specific variability in CaR signaling may in part be due to interaction(s) of CaR with scaffold proteins and the formation of cell-specific signaling complexes. The cytoskeletal protein filamin A is a scaffold for CaR, required for CaR-mediated activation of MARK signaling, and slowing CaR degradation. Preliminary data suggests CaR interactions with arrestins-1 and -2 modulate intracellular calcium responses and MARK signaling. To understand how protein scaffolds regulate CaR-mediated signaling, we will test the following hypotheses: (1) Does state-dependent arrestin-1 binding regulate CaR responsiveness? (2) Is CaR-mediated MARK signaling organized by filamin A and arrestin-2, and does the signaling pathway include transactivation of the EGF receptor? (3) Does filamin A act as a molecular escort to stabilize cellular CaR levels by protecting CaR from proteasome-mediated degradation? We will combine biochemical approaches with intracellular calcium and/or confocal imaging, using human CaR and mutant and deletion constructs. The proposed experiments will increase our understanding of the role of the protein scaffolds arrestin-1 and -2 and filamin A in enhancing the specificity of CaR signaling. More than 40 mutations in human CaR have been linked to disease, revolutionizing the clinical dissection of parathyroidisms. Allosteric modulators of CaR have clinical utility for treatment of primary hyper- and hypo- parathyroidism, secondary changes in parathyroid function resulting from renal disease, and osteoporosis. CaR stimulates cell proliferation as well as PTHrP secretion, suggesting role for CaR in potentiating the pathophysiological consequences of malignancies. Dysregulation of CaR expression and/or acute or long term signaling may, in part, be the result of altered interactions with scaffold proteins which serve to enhance CaR signaling specificity and to regulate CaR stability. Understand the role of scaffolds in CaR function may provide novel, specific sites for pharmacological intervention in treatment of calcium handling diseases.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
3R01GM077563-04S1
Application #
7937316
Study Section
Special Emphasis Panel (ZRG1-MDCN-L (02))
Program Officer
Dunsmore, Sarah
Project Start
2009-09-30
Project End
2010-03-31
Budget Start
2009-09-30
Budget End
2010-03-31
Support Year
4
Fiscal Year
2009
Total Cost
$81,054
Indirect Cost
Name
Geisinger Clinic
Department
Type
DUNS #
079161360
City
Danville
State
PA
Country
United States
Zip Code
17822
Cavanaugh, Alice; Huang, Ying; Breitwieser, Gerda E (2012) Behind the curtain: cellular mechanisms for allosteric modulation of calcium-sensing receptors. Br J Pharmacol 165:1670-1677
Grant, Michael P; Stepanchick, Ann; Cavanaugh, Alice et al. (2011) Agonist-driven maturation and plasma membrane insertion of calcium-sensing receptors dynamically control signal amplitude. Sci Signal 4:ra78
Huang, Ying; Cavanaugh, Alice; Breitwieser, Gerda E (2011) Regulation of stability and trafficking of calcium-sensing receptors by pharmacologic chaperones. Adv Pharmacol 62:143-73
Stepanchick, Ann; Breitwieser, Gerda E (2010) The cargo receptor p24A facilitates calcium sensing receptor maturation and stabilization in the early secretory pathway. Biochem Biophys Res Commun 395:136-40
Stepanchick, Ann; McKenna, Jennifer; McGovern, Olivia et al. (2010) Calcium sensing receptor mutations implicated in pancreatitis and idiopathic epilepsy syndrome disrupt an arginine-rich retention motif. Cell Physiol Biochem 26:363-74
Cavanaugh, Alice; McKenna, Jennifer; Stepanchick, Ann et al. (2010) Calcium-sensing receptor biosynthesis includes a cotranslational conformational checkpoint and endoplasmic reticulum retention. J Biol Chem 285:19854-64
White, Elissa; McKenna, Jennifer; Cavanaugh, Alice et al. (2009) Pharmacochaperone-mediated rescue of calcium-sensing receptor loss-of-function mutants. Mol Endocrinol 23:1115-23
Breitwieser, Gerda E (2008) Extracellular calcium as an integrator of tissue function. Int J Biochem Cell Biol 40:1467-80
Lorenz, Stephan; Frenzel, Romy; Paschke, Ralf et al. (2007) Functional desensitization of the extracellular calcium-sensing receptor is regulated via distinct mechanisms: role of G protein-coupled receptor kinases, protein kinase C and beta-arrestins. Endocrinology 148:2398-404
Huang, Ying; Breitwieser, Gerda E (2007) Rescue of calcium-sensing receptor mutants by allosteric modulators reveals a conformational checkpoint in receptor biogenesis. J Biol Chem 282:9517-25