Abstract
It is well established that the β2-adrenergic receptor (β2-AR) exhibits a robust ligand-independent activity, whereas this property is considerably weaker in the closely related β 1-AR subtype. To identify the potential domain(s) of β 2-AR responsible for the spontaneous receptor activation, we created three chimeras in which the third intracellular (β1/β 2-Li3 or the carboxyl terminus (β1/β 2-CT) or both domains (β1/β2-Li3CT) of β1-AR are replaced by the corresponding parts of the β2-AR. Using adenoviral gene transfer, we individually expressed these β1/β2-AR chimeras in mouse cardiomyocytes lacking both native β1-AR and β 2-AR (β1/β2 double knockout), and examined their possible spontaneous activities. Overexpression of these β1/β2-AR chimeras markedly elevated basal cAMP accumulation and myocyte contractility in the absence of agonist stimulation compared with those infected by a control adenovirus expressing β-galactosidase or an adenovirus expressing wild type β 1-AR. These effects were fully reversed by a β2-AR inverse agonist, (± )-1-[2,3-(dihydro-7-methyl-1H-inden-4-yl)oxy]-3-[(1-methylethyl)amino] -2-butanol (ICI 118,551; 5 × 10-7 M), regardless of inhibition of Gi with pertussis toxin, but not by a panel of β 1-AR antagonists, including [2-(3-carbamoyl-4-hydroxyphenoxy)-ethylamino] -3-[4-(1-methyl-4-trifluormethyl-2-imidazolyl)-phenoxy] -2-propanolmethanesulfonate (CGP20712A), betaxolol, bisoprolol, and metoprolol. Furthermore, we have shown that the C-terminal postsynaptic density 95/disc-large/ZO-1 (PDZ) motif of β1-AR is not responsible for the lack of β1-AR spontaneous activation, although it has been known that the β1-AR PDZ motif prevents the receptor from undergoing agonist-induced trafficking and Gi coupling in cardiomyocytes. Taken together, the present results indicate that both the third intracellular loop and the C terminus are involved in β2-AR spontaneous activation and that either domain seems to be sufficient to confer the receptor spontaneous activity.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 1048-1058 |
| Number of pages | 11 |
| Journal | Molecular Pharmacology |
| Volume | 64 |
| Issue number | 5 |
| DOIs | |
| State | Published - Nov 2003 |
| Externally published | Yes |
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ASJC Scopus subject areas
- Pharmacology
Cite this
The Third Intracellular Loop and the Carboxyl Terminus of β 2-Adrenergic Receptor Confer Spontaneous Activity of the Receptor. / Chakir, Khalid; Xiang, Yang Kevin; Yang, Dongmei; Zhang, Sheng Jun; Cheng, Heping; Kobilka, Brian K.; Xiao, Rui Ping.
In: Molecular Pharmacology, Vol. 64, No. 5, 11.2003, p. 1048-1058.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - The Third Intracellular Loop and the Carboxyl Terminus of β 2-Adrenergic Receptor Confer Spontaneous Activity of the Receptor
AU - Chakir, Khalid
AU - Xiang, Yang Kevin
AU - Yang, Dongmei
AU - Zhang, Sheng Jun
AU - Cheng, Heping
AU - Kobilka, Brian K.
AU - Xiao, Rui Ping
PY - 2003/11
Y1 - 2003/11
N2 - It is well established that the β2-adrenergic receptor (β2-AR) exhibits a robust ligand-independent activity, whereas this property is considerably weaker in the closely related β 1-AR subtype. To identify the potential domain(s) of β 2-AR responsible for the spontaneous receptor activation, we created three chimeras in which the third intracellular (β1/β 2-Li3 or the carboxyl terminus (β1/β 2-CT) or both domains (β1/β2-Li3CT) of β1-AR are replaced by the corresponding parts of the β2-AR. Using adenoviral gene transfer, we individually expressed these β1/β2-AR chimeras in mouse cardiomyocytes lacking both native β1-AR and β 2-AR (β1/β2 double knockout), and examined their possible spontaneous activities. Overexpression of these β1/β2-AR chimeras markedly elevated basal cAMP accumulation and myocyte contractility in the absence of agonist stimulation compared with those infected by a control adenovirus expressing β-galactosidase or an adenovirus expressing wild type β 1-AR. These effects were fully reversed by a β2-AR inverse agonist, (± )-1-[2,3-(dihydro-7-methyl-1H-inden-4-yl)oxy]-3-[(1-methylethyl)amino] -2-butanol (ICI 118,551; 5 × 10-7 M), regardless of inhibition of Gi with pertussis toxin, but not by a panel of β 1-AR antagonists, including [2-(3-carbamoyl-4-hydroxyphenoxy)-ethylamino] -3-[4-(1-methyl-4-trifluormethyl-2-imidazolyl)-phenoxy] -2-propanolmethanesulfonate (CGP20712A), betaxolol, bisoprolol, and metoprolol. Furthermore, we have shown that the C-terminal postsynaptic density 95/disc-large/ZO-1 (PDZ) motif of β1-AR is not responsible for the lack of β1-AR spontaneous activation, although it has been known that the β1-AR PDZ motif prevents the receptor from undergoing agonist-induced trafficking and Gi coupling in cardiomyocytes. Taken together, the present results indicate that both the third intracellular loop and the C terminus are involved in β2-AR spontaneous activation and that either domain seems to be sufficient to confer the receptor spontaneous activity.
AB - It is well established that the β2-adrenergic receptor (β2-AR) exhibits a robust ligand-independent activity, whereas this property is considerably weaker in the closely related β 1-AR subtype. To identify the potential domain(s) of β 2-AR responsible for the spontaneous receptor activation, we created three chimeras in which the third intracellular (β1/β 2-Li3 or the carboxyl terminus (β1/β 2-CT) or both domains (β1/β2-Li3CT) of β1-AR are replaced by the corresponding parts of the β2-AR. Using adenoviral gene transfer, we individually expressed these β1/β2-AR chimeras in mouse cardiomyocytes lacking both native β1-AR and β 2-AR (β1/β2 double knockout), and examined their possible spontaneous activities. Overexpression of these β1/β2-AR chimeras markedly elevated basal cAMP accumulation and myocyte contractility in the absence of agonist stimulation compared with those infected by a control adenovirus expressing β-galactosidase or an adenovirus expressing wild type β 1-AR. These effects were fully reversed by a β2-AR inverse agonist, (± )-1-[2,3-(dihydro-7-methyl-1H-inden-4-yl)oxy]-3-[(1-methylethyl)amino] -2-butanol (ICI 118,551; 5 × 10-7 M), regardless of inhibition of Gi with pertussis toxin, but not by a panel of β 1-AR antagonists, including [2-(3-carbamoyl-4-hydroxyphenoxy)-ethylamino] -3-[4-(1-methyl-4-trifluormethyl-2-imidazolyl)-phenoxy] -2-propanolmethanesulfonate (CGP20712A), betaxolol, bisoprolol, and metoprolol. Furthermore, we have shown that the C-terminal postsynaptic density 95/disc-large/ZO-1 (PDZ) motif of β1-AR is not responsible for the lack of β1-AR spontaneous activation, although it has been known that the β1-AR PDZ motif prevents the receptor from undergoing agonist-induced trafficking and Gi coupling in cardiomyocytes. Taken together, the present results indicate that both the third intracellular loop and the C terminus are involved in β2-AR spontaneous activation and that either domain seems to be sufficient to confer the receptor spontaneous activity.
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UR - http://www.scopus.com/inward/citedby.url?scp=0142148104&partnerID=8YFLogxK
U2 - 10.1124/mol.64.5.1048
DO - 10.1124/mol.64.5.1048
M3 - Article
C2 - 14573753
AN - SCOPUS:0142148104
VL - 64
SP - 1048
EP - 1058
JO - Molecular Pharmacology
JF - Molecular Pharmacology
SN - 0026-895X
IS - 5
ER -