scholarly journals Proteolytic cleavage of the extracellular domain affects signaling of parathyroid hormone receptor 1

2021 ◽  
Author(s):  
Christoph Klenk ◽  
Leif Hommers ◽  
Martin J. Lohse
Keyword(s):  
Parathyroid Hormone ◽  
Extracellular Domain ◽  
Proteolytic Cleavage ◽  
Protein Sequencing ◽  
Signaling Specificity ◽  
Parathyroid Hormone Receptor ◽  
Amino Terminal ◽  
Class B ◽  
Receptor Mutant

Parathyroid hormone 1 receptor (PTH1R) is a member of the class B family of G protein-coupled receptors, which are characterized by a large extracellular domain required for ligand binding. We have previously shown that the extracellular domain of PTH1R is subject to metalloproteinase cleavage in vivo that is regulated by ligand-induced receptor trafficking and leads to impaired stability of PTH1R. In this work, we localize the cleavage site in the first loop of the extracellular domain using amino-terminal protein sequencing of purified receptor and by mutagenesis studies. We further show, that a receptor mutant not susceptible to proteolytic cleavage exhibits reduced signaling to Gs and increased activation of Gq/11 compared to wild-type PTH1R. These findings indicate that the extracellular domain modulates PTH1R signaling specificity.

scholarly journals Actions of the Small Molecule Ligands SW106 and AH-3960 on the Type-1 Parathyroid Hormone Receptor

2015 ◽  
Vol 29 (2) ◽  
pp. 307-321 ◽  
Author(s):  
Percy H. Carter ◽  
Thomas Dean ◽  
Brijesh Bhayana ◽  
Ashok Khatri ◽  
Raj Rajur ◽  
...  
Keyword(s):  
Parathyroid Hormone ◽  
Small Molecule ◽  
Hormone Receptor ◽  
Transmembrane Domain ◽  
Receptor Activation ◽  
Blood Calcium ◽  
Parathyroid Hormone Receptor ◽  
Peptide Analog ◽  
Amino Terminal ◽  
Small Molecule Ligands

Abstract The parathyroid hormone receptor-1 (PTHR1) plays critical roles in regulating blood calcium levels and bone metabolism and is thus of interest for small-molecule ligand development. Of the few small-molecule ligands reported for the PTHR1, most are of low affinity, and none has a well-defined mechanism of action. Here, we show that SW106 and AH-3960, compounds previously identified to act as an antagonist and agonist, respectively, on the PTHR1, each bind to PTHR1-delNT, a PTHR1 construct that lacks the large amino-terminal extracellular domain used for binding endogenous PTH peptide ligands, with the same micromolar affinity with which it binds to the intact PTHR1. SW106 antagonized PTHR1-mediated cAMP signaling induced by the peptide analog, M-PTH(1–11), as well as by the native PTH(1–9) sequence, as tethered to the extracellular end of transmembrane domain (TMD) helix-1 of the receptor. SW106, however, did not function as an inverse agonist on either PTHR1-H223R or PTHR1-T410P, which have activating mutations at the cytoplasmic ends of TMD helices 2 and 6, respectively. The overall data indicate that SW106 and AH-3960 each bind to the PTHR1 TMD region and likely to within an extracellularly exposed area that is occupied by the N-terminal residues of PTH peptides. Additionally, they suggest that the inhibitory effects of SW106 are limited to the extracellular portions of the TMD region that mediate interactions with agonist ligands but do not extend to receptor-activation determinants situated more deeply in the helical bundle. The study helps to elucidate potential mechanisms of small-molecule binding at the PTHR1.

scholarly journals The CBP Bromodomain and Nucleosome Targeting Are Required for Zta-Directed Nucleosome Acetylation and Transcription Activation

2003 ◽  
Vol 23 (8) ◽  
pp. 2633-2644 ◽  
Author(s):  
Zhong Deng ◽  
Chi-Ju Chen ◽  
Michaela Chamberlin ◽  
Fang Lu ◽  
Gerd A. Blobel ◽  
...  
Keyword(s):  
Complex Formation ◽  
Histone Acetylation ◽  
Protein Sequencing ◽  
Creb Binding Protein ◽  
Barr Virus ◽  
Amino Terminal ◽  
Viral Promoters ◽  
Epstein Barr

ABSTRACT The Epstein-Barr virus (EBV)-encoded lytic activator Zta is a bZIP protein that can stimulate nucleosomal histone acetyltransferase (HAT) activity of the CREB binding protein (CBP) in vitro. We now show that deletion of the CBP bromo- and C/H3 domains eliminates stimulation of nucleosomal HAT activity in vitro and transcriptional coactivation by Zta in transfected cells. In contrast, acetylation of free histones was not affected by the addition of Zta or by deletions in the bromo or C/H3 domain of CBP. Zta stimulated acetylation of oligonucleosomes assembled on supercoiled DNA and dinucleosomes assembled on linear DNA, but Zta-stimulated acetylation was significantly reduced for mononucleosomes. Western blotting and amino-terminal protein sequencing indicated that all lysine residues in the H3 and H4 amino-terminal tails were acetylated by CBP and enhanced by the addition of Zta. Histone acetylation was also dependent upon the Zta basic DNA binding domain, which could not be substituted with the homologous basic region of c-Fos, indicating specificity in the bZIP domain nucleosome binding function. Finally, we show that Zta and CBP colocalize to viral immediate-early promoters in vivo and that overexpression of Zta leads to a robust increase in H3 and H4 acetylation at various regions of the EBV genome in vivo. Furthermore, deletion of the CBP bromodomain reduced stable CBP-Zta complex formation and histone acetylation at Zta-responsive viral promoters in vivo. These results suggest that activator- and bromodomain-dependent targeting to oligonucleosomal chromatin is required for stable promoter-bound complex formation and transcription activity.

Effects of parathyroid hormone and the synthetic 1–34 amino-terminal fragment in rats and dogs

1983 ◽  
Vol 97 (1) ◽  
pp. 21-30
Author(s):  
R. W. Stevenson ◽  
J. A. Parsons
Keyword(s):  
Parathyroid Hormone ◽  
Infusion Rate ◽  
Calcium Absorption ◽  
Biological Activities ◽  
Direct Action ◽  
Plasma Calcium ◽  
Biologically Active ◽  
Amino Terminal ◽  
Active Fragments

Since the structural requirements for all known biological activities of parathyroid hormone (PTH(1–84)) are virtually satisfied by the amino-terminal 34 amino acid fragment, PTH(1–34), we investigated whether this fragment could elaborate the overall actions of the intact hormone in the whole animal by comparing the effects of equimolar infusions of each peptide to dogs and rats. Infusion of bovine PTH(1–84) (bPTH(1–84)) at 17 pmol/kg per h for 20 h to three dogs or at 100 and 200 pmol/kg per h to groups of six rats for 5 days produced greater hypercalcaemia (3·02±0·03, 2·52±0·07 and 3·24±0·11 mmol/l respectively) than equimolar infusions of human PTH(1–34) (hPTH(1–34)) (2·61±0·03, 2·46 ± 0·05 and 2·71 ±0·09 mmol/l respectively). A significant calcium rise was not observed in dogs until after 4 h of PTH infusion. No rise in plasma calcium was apparent in rats, however, until the third day of PTH infusion. Only in parathyroidectomized rats was there a rise in plasma calcium within 24 h of starting an infusion of PTH. The hypercalciuria and plasma phosphate responses in dogs during equimolar infusions of hPTH(1–34) and bPTH(1–84) were not significantly different. However, by day 5 of infusion in rats greater hypercalciuria was produced by bPTH(1–84). Although infusion of hPTH(1–34) and bPTH(1–84) caused rises in urinary cyclic AMP excretion (measured only in the dog) of immediate onset and equal magnitude, bPTH(1–84) tended to produce greater phosphaturia than hPTH(1–34) in both species. If the assumption is correct that the half-lives of hPTH(1–34) and bPTH(1–84) in the circulation are similar and provided that hPTH does not inherently have less biological activity than bPTH, then during equimolar infusions of these peptides into dogs and rats, the greater responses observed with bPTH(1–84) suggest that intact PTH may have a direct action of its own in vivo before being metabolized into smaller biologically active fragments. In additional experiments using parathyroidectomized rats, the infusion rate of bPTH(1–84) required to restore normocalcaemia was 26 pmol/kg per h. Although near-normal calcaemia and intestinal calcium absorption could still be maintained when the infusion rate was increased to 39 pmol/kg per h, hypercalciuria and phosphaturia became apparent.

scholarly journals Agonist-regulated Cleavage of the Extracellular Domain of Parathyroid Hormone Receptor Type 1

2010 ◽  
Vol 285 (12) ◽  
pp. 8665-8674 ◽  
Author(s):  
Christoph Klenk ◽  
Stefan Schulz ◽  
Davide Calebiro ◽  
Martin J. Lohse
Keyword(s):  
Parathyroid Hormone ◽  
Hormone Receptor ◽  
Extracellular Domain ◽  
Receptor Type ◽  
Parathyroid Hormone Receptor

In Vivo Demonstration of Receptors in Rat Liver to the Amino-Terminal Region of Parathyroid Hormone*

Endocrinology ◽  
1981 ◽  
Vol 109 (5) ◽  
pp. 1552-1559 ◽  
Author(s):  
J. J. M. BERGERON ◽  
S. TCHERVENKOV ◽  
M. F. ROULEAU ◽  
M. ROSENBLATT ◽  
D. GOLTZMAN
Keyword(s):  
Parathyroid Hormone ◽  
Rat Liver ◽  
Terminal Region ◽  
Amino Terminal

scholarly journals Differential effects of parathyroid hormone fragments on collagen gene expression in chondrocytes.

1996 ◽  
Vol 135 (4) ◽  
pp. 1179-1191 ◽  
Author(s):  
S Erdmann ◽  
W Müller ◽  
S Bahrami ◽  
S I Vornehm ◽  
H Mayer ◽  
...  
Keyword(s):  
Gene Expression ◽  
Parathyroid Hormone ◽  
Cartilage Matrix ◽  
Functional Domain ◽  
Collagen Gene ◽  
Terminal Portion ◽  
Amino Terminal ◽  
Collagen Gene Expression ◽  
Matrix Metabolism

The effect of parathyroid hormone (PTH) in vivo after secretion by the parathyroid gland is mediated by bioactive fragments of the molecule. To elucidate their possible role in the regulation of cartilage matrix metabolism, the influence of the amino-terminal (NH2-terminal), the central, and the carboxyl-terminal (COOH-terminal) portion of the PTH on collagen gene expression was studied in a serum free cell culture system of fetal bovine and human chondrocytes. Expression of alpha1 (I), alpha1 (II), alpha1 (III), and alpha1 (X) mRNA was investigated by in situ hybridization and quantified by Northern blot analysis. NH2-terminal and mid-regional fragments containing a core sequence between amino acid residues 28-34 of PTH induced a significant rise in alpha1 (II) mRNA in proliferating chondrocytes. In addition, the COOH-terminal portion (aa 52-84) of the PTH molecule was shown to exert a stimulatory effect on alpha1 (II) and alpha1 (X) mRNA expression in chondrocytes from the hypertrophic zone of bovine epiphyseal cartilage. PTH peptides harboring either the functional domain in the central or COOH-terminal region of PTH can induce cAMP independent Ca2+ signaling in different subsets of chondrocytes as assessed by microfluorometry of Fura-2/AM loaded cells. These results support the hypothesis that different hormonal effects of PTH on cartilage matrix metabolism are exerted by distinct effector domains and depend on the differentiation stage of the target cell.

Actions of parathyroid hormone-related protein on the rat kidney in vivo

1989 ◽  
Vol 122 (1) ◽  
pp. 229-235 ◽  
Author(s):  
H. Zhou ◽  
D. D. Leaver ◽  
J. M. Moseley ◽  
B. Kemp ◽  
P. R. Ebeling ◽  
...  
Keyword(s):  
Parathyroid Hormone ◽  
Infusion Rate ◽  
Rat Kidney ◽  
Physiological Activity ◽  
Calcium Excretion ◽  
Related Protein ◽  
Renal Handling ◽  
Amino Terminal ◽  
Parathyroid Hormone Related Protein

ABSTRACT Peptides containing residues 1–34 of parathyroid hormone-related protein (PTHrP) and of bovine parathyroid hormone (bPTH), and recombinant full-length PTHrP(1–141) were infused i.v. into anaesthetized thyroparathyroidectomized rats to compare their action and potency on the renal handling of calcium, phosphate and cyclic AMP (cAMP) in vivo. All three peptides decreased the excretion of calcium and increased the excretion of phosphate and cAMP in the urine, with PTHrP(1–34) and PTHrP(1–141) having virtually equipotent effects. Thus the essential requirements for the major physiological activity of PTHrP on the kidney are contained within the 34 amino-terminal amino acids. For all three peptides, the lowest infusion rate that increased phosphate and cAMP excretion was 0·01 nmol/kg per h, whereas the lowest infusion rate that decreased calcium excretion was 0·025 nmol/kg per h for the PTHrP peptides and 0·1 nmol/kg per h for bPTH(1–34). The response to the PTHrP peptides was maximal at an infusion rate of 01 nmol/kg per h for both calcium and phosphate. Since the kidney is either equally sensitive to PTHrP and bPTH(1–34), or more sensitive to PTHrP than to bPTH(1–34), the hypercalcaemia of humoral hypercalcaemia of malignancy may develop because uncontrolled secretion of PTHrP increases the renal reabsorption of calcium to such an extent that even a modest increase in the inflow of calcium into the blood raises plasma calcium concentration. Journal of Endocrinology (1989) 122, 229–235

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