Large Putative PEST-like Sequence Motif at the Carboxyl Tail of Human Calcium Receptor Directs Lysosomal Degradation and Regulates Cell Surface Receptor Level

Zhuang, Xiaolei; Northup, John K.; Ray, Kausik K.

doi:10.1074/jbc.m111.271528

Cited by 29 publications

(43 citation statements)

References 38 publications

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“…CaSR is the closest related mammalian receptor of GPRC6A and has proven highly resistant to desensitization, which most likely reflects a persistent exposure to the endogenous agonist Ca 2ϩ (50,51). Constitutive internalization of CaSR has been observed in some studies (52,53), and moreover, a distinct regulatory mechanism for CaSR has been reported in which agonist stimulation triggers further insertion of receptors in the cell membrane from intracellular stores, thereby increasing the amount of receptors available for activation. This mechanism has been named agonist-driven insertional signaling (54).…”

Section: Discussionmentioning

confidence: 99%

The GPRC6A receptor displays constitutive internalization and sorting to the slow recycling pathway

Jacobsen

Ammendrup-Johnsen

Jansen

et al. 2017

Journal of Biological Chemistry

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The class C G protein-coupled receptor GPRC6A is a putative nutrient-sensing receptor and represents a possible new drug target in metabolic disorders. However, the specific physiological role of this receptor has yet to be identified, and the mechanisms regulating its activity and cell surface availability also remain enigmatic. In the present study, we investigated the trafficking properties of GPRC6A by use of both a classical antibody feeding internalization assay in which cells were visualized using confocal microscopy and a novel internalization assay that is based on real-time measurements of fluorescence resonance energy transfer. Both assays revealed that GPRC6A predominantly undergoes constitutive internalization, whereas the agonist-induced effects were imperceptible. Moreover, postendocytic sorting was investigated by assessing the co-localization of internalized GPRC6A with selected Rab protein markers. Internalized GPRC6A was mainly co-localized with the early endosome marker Rab5 and the long loop recycling endosome marker Rab11 and to a much lesser extent with the late endosome marker Rab7. This suggests that upon agonist-independent internalization, GPRC6A is recycled via the Rab11-positive slow recycling pathway, which may be responsible for ensuring a persistent pool of GPRC6A receptors at the cell surface despite chronic agonist exposure. Distinct trafficking pathways have been reported for several of the class C receptors, and our results thus substantiate that non-canonical trafficking mechanisms are a common feature for the nutrient-sensing class C family that ensure functional receptors in the cell membrane despite prolonged agonist exposure.The G protein-coupled receptor, class C, group 6, member A (GPRC6A), 2 is a nutrient-sensing G protein-coupled receptor (GPCR) belonging to class C. It is activated by basic L-␣-amino acids and divalent cations, which trigger coupling to the G q protein and thereby an increase in the intracellular levels of the second messengers inositol trisphosphate and Ca 2ϩ (1-8). Other ligands and signaling pathways have been reported (3, 9 -13); however Rueda et al. (8) and we (7) have not been able to confirm these findings. The GPRC6A receptor displays a broad but low expression profile in human, mouse, and rat (2,5,10,11,14,15), thus giving little indication to the physiological role of the receptor. Several groups have conducted studies using GPRC6A knock-out mice to elucidate the physiological function of the receptor, and although results differ between knockout mouse models, they altogether suggest an involvement in metabolism and endocrine regulation (6, 10, 11, 16 -18).Over the years, it has become evident that GPCR signaling is much more complex than once believed. For most receptors, a variety of ligands and intracellular signaling pathways are available, and numerous regulatory mechanisms are thus required for obtaining specific biological responses (19).

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Section: Discussionmentioning

confidence: 99%

The GPRC6A receptor displays constitutive internalization and sorting to the slow recycling pathway

Jacobsen

Ammendrup-Johnsen

Jansen

et al. 2017

Journal of Biological Chemistry

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“…A number of reports suggest that CaSR endocytosis is rapid, constitutive, and facilitated by Rab7 and Rab11a. 68,84 Recycling of the CaSR has been noted in some studies 68,84 but not in others, 63 suggesting that the constitutive receptor internalization may require a different endocytic machinery. The CaSR endocytosis is essential for the transactivation of epidermal growth factor receptor that leads to the MAP kinase signaling cascade and links receptor signaling to PTH-related peptide secretion via a Rab11a-dependent and associated molecule with the SH3 domain of STAM (AMSH)-sensitive mechanism.…”

Section: Endocytosis and Recycling Of Casrmentioning

confidence: 94%

Calcium-Sensing Receptor

Ray¹

2015

Progress in Molecular Biology and Translational Science

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“…A long PEST-like sequence motif (rich in proline, glutamine, serine and threonine) within amino acids 920-970 of the C-tail directs lysosomal degradation and regulates cell surface receptor level (30) with the latter property also influenced by an arginine-rich motif, R890-R898 (31). A leucine 1013, leucine 1014 motif in the CASR ICD interacts with the multi-subunit adaptor protein 2 (AP-2) that mediates endocytosis of the CASR.…”

Section: Discussionmentioning

confidence: 99%

Novel calcium-sensing receptor cytoplasmic tail deletion mutation causing autosomal dominant hypocalcemia: molecular and clinical study

Obermannová

Šumnı́k

Dušátková

et al. 2016

European Journal of Endocrinology

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Objective: Autosomal dominant hypocalcemia (ADH) is a rare disorder caused by activating mutations of the calcium-sensing receptor (CASR). The treatment of ADH patients with 1a-hydroxylated vitamin D derivatives can cause hypercalciuria leading to nephrocalcinosis. Design and methods: We studied a girl who presented with hypoparathyroidism and asymptomatic hypocalcemia at age 2.5 years. Mutations of CASR were investigated by DNA sequencing. Functional analyses of mutant and WT CASRs were done in transiently transfected human embryonic kidney (HEK293) cells. Results: The proband and her father are heterozygous for an eight-nucleotide deletion c.2703_2710delCCTTGGAG in the CASR encoding the intracellular domain of the protein. Transient expression of CASR constructs in kidney cells in vitro suggested greater cell surface expression of the mutant receptor with a left-shifted extracellular calcium dose-response curve relative to that of the WT receptor consistent with gain of function. Initial treatment of the patient with calcitriol led to increased urinary calcium excretion. Evaluation for mosaicism in the paternal grandparents of the proband was negative. Conclusions: We describe a novel naturally occurring deletion mutation within the CASR that apparently arose de novo in the father of the ADH proband. Functional analysis suggests that the cytoplasmic tail of the CASR contains determinants that regulate the attenuation of signal transduction. Early molecular analysis of the CASR gene in patients with isolated idiopathic hypoparathyroidism is recommended because of its relevance to clinical outcome and treatment choice. In ADH patients, calcium supplementation and low-dose cholecalciferol avoids hypocalcemic symptoms without compromising renal function.

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Large Putative PEST-like Sequence Motif at the Carboxyl Tail of Human Calcium Receptor Directs Lysosomal Degradation and Regulates Cell Surface Receptor Level

Cited by 29 publications

References 38 publications

The GPRC6A receptor displays constitutive internalization and sorting to the slow recycling pathway

The GPRC6A receptor displays constitutive internalization and sorting to the slow recycling pathway

Calcium-Sensing Receptor

Novel calcium-sensing receptor cytoplasmic tail deletion mutation causing autosomal dominant hypocalcemia: molecular and clinical study

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