Parathyroid hormone-related peptide and parathyroid hormone-related peptide receptor in breast cancer mcf7 cells
Parathyroid hormone-related peptide and parathyroid hormone-related peptide receptor in breast cancer mcf7 cells
The aim of this work was to clarify the nature and the affinity state of the PTHrP receptor in breast cancer MCF7 cells compared to the classical PTH/PTHrP receptor expressed in the SaOS-2 osteosarcoma cell line.
Quantitative equilibrium ligand binding assays and competitive binding assays using 125I-PTHrP-1-34Tyrosine demonstrated that PTHrP bound with high affinity in both cell lines, MCF7 and SaOS-2. However, KD values were not the same in both cell types. Ligand-receptor labelling followed by SDS-PAGE analysis revealed the presence of a single 85,000 Dalton band in both cell lines, which was thought to be the PTH/PTHrP receptor, since ligand binding was specifically inhibited by excess of unlabelled ligand.
The substitution at position 23 of PTHrP caused an increase in the KD values for equilibrium ligand binding from 2.0 nM to 2.75 nM in MCF7 cells and from 5.0 nM to 5.4 nM in SaOS-2 cells. However, when both amino acids at positions 5 and 23 (His5 and Phe23) in the PTHrP molecule were substituted for those in PTH (Ile5 and Trp23), the affinity of the PTHrP receptor decreased to 4.1nM in MCF7 cells similar to that for PTH (4.25 nM) in MCF7 cells.
The PTHrP receptor expression in MCF7 cells was increased following exposure to EGF or E2, whereas 1,25 DHCC inhibited the receptor expression in MCF7 cells, while DEX had no effect. In contrast, 1,25 DHCC and E2 treatment increased PTH/PTHrP receptor expression in SaOS-2 cells whereas EGF did not, while DEX inhibited the PTH/PTHrP receptor expression in SaOS-2 cells. This suggests that receptor expression is differently regulated in the two cell lines.
The results suggest that the PTHrP receptor in MCF7 cells has many characteristics which distinguish from the receptor in SaOS-2 cells. The control of both PTHrP receptor expression and peptide production also differs between these two cell lines. These data show for the first time that there are higher affinity receptors for PTHrP in MCF7 breast cancer cells.
University of Southampton
Alokail, Majed Saleh Abdullah
284cdde2-2d1a-4928-a304-5d3f97843499
1999
Alokail, Majed Saleh Abdullah
284cdde2-2d1a-4928-a304-5d3f97843499
Alokail, Majed Saleh Abdullah
(1999)
Parathyroid hormone-related peptide and parathyroid hormone-related peptide receptor in breast cancer mcf7 cells.
University of Southampton, Doctoral Thesis.
Record type:
Thesis
(Doctoral)
Abstract
The aim of this work was to clarify the nature and the affinity state of the PTHrP receptor in breast cancer MCF7 cells compared to the classical PTH/PTHrP receptor expressed in the SaOS-2 osteosarcoma cell line.
Quantitative equilibrium ligand binding assays and competitive binding assays using 125I-PTHrP-1-34Tyrosine demonstrated that PTHrP bound with high affinity in both cell lines, MCF7 and SaOS-2. However, KD values were not the same in both cell types. Ligand-receptor labelling followed by SDS-PAGE analysis revealed the presence of a single 85,000 Dalton band in both cell lines, which was thought to be the PTH/PTHrP receptor, since ligand binding was specifically inhibited by excess of unlabelled ligand.
The substitution at position 23 of PTHrP caused an increase in the KD values for equilibrium ligand binding from 2.0 nM to 2.75 nM in MCF7 cells and from 5.0 nM to 5.4 nM in SaOS-2 cells. However, when both amino acids at positions 5 and 23 (His5 and Phe23) in the PTHrP molecule were substituted for those in PTH (Ile5 and Trp23), the affinity of the PTHrP receptor decreased to 4.1nM in MCF7 cells similar to that for PTH (4.25 nM) in MCF7 cells.
The PTHrP receptor expression in MCF7 cells was increased following exposure to EGF or E2, whereas 1,25 DHCC inhibited the receptor expression in MCF7 cells, while DEX had no effect. In contrast, 1,25 DHCC and E2 treatment increased PTH/PTHrP receptor expression in SaOS-2 cells whereas EGF did not, while DEX inhibited the PTH/PTHrP receptor expression in SaOS-2 cells. This suggests that receptor expression is differently regulated in the two cell lines.
The results suggest that the PTHrP receptor in MCF7 cells has many characteristics which distinguish from the receptor in SaOS-2 cells. The control of both PTHrP receptor expression and peptide production also differs between these two cell lines. These data show for the first time that there are higher affinity receptors for PTHrP in MCF7 breast cancer cells.
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Published date: 1999
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Local EPrints ID: 463751
URI: http://eprints.soton.ac.uk/id/eprint/463751
PURE UUID: 2bce1a38-80e4-49b2-a310-f858b43ae893
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Date deposited: 04 Jul 2022 20:56
Last modified: 23 Jul 2022 02:15
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Author:
Majed Saleh Abdullah Alokail
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