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Mutated human beta3-adrenergic receptor (Trp64Arg) lowers the response to beta3-adrenergic agonists in transfected
1Department of Biomedical Sciences, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo, Japan. k-kimura@vetmed.hokudai.ac.jp
Abstract:
Wild-type or mutated human beta3-adrenergic receptor (Trp64Arg) cDNAs were stably expressed in mouse 3T3-L1 cells. Saturation binding study using a beta-adrenergic ligand revealed that there was no significant difference in the receptor density and the equilibrium dissociation constant between the two cell lines. However, the ability of the mutant beta3-adrenergic receptor to accumulate cyclic AMP (cAMP) in response to isoproterenol was much reduced and Kact for cAMP accumulation was lowered as compared to the wild type receptor. The amount of alpha subunit of stimulatory GTP-binding protein (GSalpha) and adenylyl cyclase activity in response to forskolin were not different in the two cell lines. The responses of the mutant receptor to epinephrine, norepinephrine and L-755,507, a highly specific agonist for human beta3-adrenergic receptor, were also reduced, but the reduction of Kact for L-755,507 was more evident than other agonists tested. The cAMP accumulation in response to some conventional beta3 agonists was less than 10% of that to isoproterenol even in the cells expressing the wild type receptor. These results suggest that the Trp64Arg mutant beta3-adrenergic receptor has less ability to stimulate adenylyl cyclase, and that lipolytic activity through the beta3-adrenergic receptor by catecholamines in subjects carrying this mutation might be suppressed.
Insights
The Trp64Arg mutation in the beta3-adrenergic receptor impairs its ability to activate cyclic AMP (cAMP) signaling. This suggests suppressed lipolytic activity in individuals with this common beta3 receptor mutation.
Area of Science:
- Pharmacology
- Molecular Biology
- Genetics
Background:
- The beta3-adrenergic receptor plays a role in regulating metabolic processes.
- A common mutation, Trp64Arg, is found in the beta3-adrenergic receptor gene.
Purpose of the Study:
- To investigate the functional consequences of the Trp64Arg mutation on beta3-adrenergic receptor activity.
- To compare the signaling capabilities of wild-type and mutant beta3-adrenergic receptors.
Main Methods:
- Stable expression of wild-type and Trp64Arg mutant human beta3-adrenergic receptors in mouse 3T3-L1 cells.
- Saturation binding assays to determine receptor density and affinity.
- Measurement of cyclic AMP (cAMP) accumulation in response to various adrenergic agonists and forskolin.
Main Results:
- No significant differences in receptor density or binding affinity between wild-type and mutant receptors.
- The Trp64Arg mutant receptor showed significantly reduced cyclic AMP (cAMP) accumulation in response to isoproterenol compared to the wild-type.
- Responses to other agonists, including epinephrine, norepinephrine, and L-755,507, were also reduced, with a more pronounced effect on the activation constant (Kact) for L-755,507.
Conclusions:
- The Trp64Arg mutation in the beta3-adrenergic receptor leads to a diminished ability to stimulate adenylyl cyclase.
- Lipolytic activity mediated by catecholamines through the beta3-adrenergic receptor may be suppressed in individuals carrying the Trp64Arg mutation.