Scholars@Duke publication: Protein kinase A and G protein-coupled receptor kinase phosphorylation mediates beta-1 adrenergic receptor endocytosis through different pathways.

Protein kinase A and G protein-coupled receptor kinase phosphorylation mediates beta-1 adrenergic receptor endocytosis through different pathways.

Publication , Journal Article

Rapacciuolo, A; Suvarna, S; Barki-Harrington, L; Luttrell, LM; Cong, M; Lefkowitz, RJ; Rockman, HA

Published in: J Biol Chem

September 12, 2003

Agonist-induced phosphorylation of beta-adrenergic receptors (beta ARs) by G protein-coupled receptor kinases (GRKs) results in their desensitization followed by internalization. Whether protein kinase A (PKA)-mediated phosphorylation of beta ARs, particularly the beta 1AR subtype, can also trigger internalization is currently not known. To test this, we cloned the mouse wild type beta 1AR (WT beta 1AR) and created 3 mutants lacking, respectively: the putative PKA phosphorylation sites (PKA-beta 1AR), the putative GRK phosphorylation sites (GRK-beta 1AR), and both sets of phosphorylation sites (PKA-/GRK-beta 1AR). Following agonist stimulation, both PKA-beta 1AR and GRK-beta 1AR mutants showed comparable increases in phosphorylation and desensitization. Saturating concentrations of agonist induced only 50% internalization of either mutant compared with wild type, suggesting that both PKA and GRK phosphorylation of the receptor contributed to receptor sequestration in an additive manner. Moreover, in contrast to the WT beta 1AR and PKA-beta 1AR, sequestration of the GRK-beta 1AR and PKA-/GRK-beta 1AR was independent of beta-arrestin recruitment. Importantly, clathrin inhibitors abolished agonist-dependent internalization for both the WT beta 1AR and PKA-beta 1AR, whereas caveolae inhibitors prevented internalization only of the GRK-beta 1AR mutant. Taken together, these data demonstrate that: 1) PKA-mediated phosphorylation can trigger agonist-induced internalization of the beta 1AR and 2) the pathway selected for beta 1AR internalization is primarily determined by the kinase that phosphorylates the receptor, i.e. PKA-mediated phosphorylation directs internalization via a caveolae pathway, whereas GRK-mediated phosphorylation directs it through clathrin-coated pits.

Duke Scholars

Author Robert J. Lefkowitz Medicine, Cardiology

Author Howard Allan Rockman Medicine, Cardiology

Published In

J Biol Chem

DOI

10.1074/jbc.M305675200

ISSN

0021-9258

Publication Date

September 12, 2003

Volume

278

Issue

Start / End Page

35403 / 35411

Location

United States

Related Subject Headings

Transfection
Sequence Deletion
Recombinant Proteins
Receptors, Adrenergic, beta-2
Receptors, Adrenergic, beta-1
Receptor Protein-Tyrosine Kinases
Protein Structure, Secondary
Phosphorylation
Models, Molecular
Mice

Citation

APA

Chicago

ICMJE

MLA

NLM

Rapacciuolo, A., Suvarna, S., Barki-Harrington, L., Luttrell, L. M., Cong, M., Lefkowitz, R. J., & Rockman, H. A. (2003). Protein kinase A and G protein-coupled receptor kinase phosphorylation mediates beta-1 adrenergic receptor endocytosis through different pathways. J Biol Chem, 278(37), 35403–35411. https://doi.org/10.1074/jbc.M305675200

Rapacciuolo, Antonio, Shayela Suvarna, Liza Barki-Harrington, Louis M. Luttrell, Mei Cong, Robert J. Lefkowitz, and Howard A. Rockman. “Protein kinase A and G protein-coupled receptor kinase phosphorylation mediates beta-1 adrenergic receptor endocytosis through different pathways.” J Biol Chem 278, no. 37 (September 12, 2003): 35403–11. https://doi.org/10.1074/jbc.M305675200.

Rapacciuolo A, Suvarna S, Barki-Harrington L, Luttrell LM, Cong M, Lefkowitz RJ, et al. Protein kinase A and G protein-coupled receptor kinase phosphorylation mediates beta-1 adrenergic receptor endocytosis through different pathways. J Biol Chem. 2003 Sep 12;278(37):35403–11.

Rapacciuolo, Antonio, et al. “Protein kinase A and G protein-coupled receptor kinase phosphorylation mediates beta-1 adrenergic receptor endocytosis through different pathways.” J Biol Chem, vol. 278, no. 37, Sept. 2003, pp. 35403–11. Pubmed, doi:10.1074/jbc.M305675200.

Rapacciuolo A, Suvarna S, Barki-Harrington L, Luttrell LM, Cong M, Lefkowitz RJ, Rockman HA. Protein kinase A and G protein-coupled receptor kinase phosphorylation mediates beta-1 adrenergic receptor endocytosis through different pathways. J Biol Chem. 2003 Sep 12;278(37):35403–35411.