Hakalahti, A. E., Vierimaa, M. M., Lilja, M. K., Kumpula, E.-P., Tuusa, J. T., & Petäjä-Repo, U. E. (2010). Human β1-Adrenergic Receptor Is Subject to Constitutive and Regulated N-terminal Cleavage. Journal of Biological Chemistry, 285(37), 28850–28861. https://doi.org/10.1074/jbc.m110.149989
Human β1-adrenergic receptor is subject to constitutive and regulated N-terminal cleavage
|Author:||Hakalahti, Anna E.1; Vierimaa, Miia M.1; Lilja, Minna K.1;|
1Department of Anatomy and Cell Biology, Institute of Biomedicine, University of Oulu, FI-90014 Oulu, Finland
|Online Access:||PDF Full Text (PDF, 1.8 MB)|
|Persistent link:|| http://urn.fi/urn:nbn:fi-fe2019120445642
American Society for Biochemistry and Molecular Biology,
|Publish Date:|| 2019-12-04
The β₁-adrenergic receptor (β₁AR) is the predominant βAR in the heart, mediating the catecholamine-stimulated increase in cardiac rate and force of contraction. Regulation of this important G protein-coupled receptor is nevertheless poorly understood. We describe here the biosynthetic profile of the human β₁AR and reveal novel features relevant to its regulation using an inducible heterologous expression system in HEK293i cells. Metabolic pulse-chase labeling and cell surface biotinylation assays showed that the synthesized receptors are efficiently and rapidly transported to the cell surface. The N terminus of the mature receptor is extensively modified by sialylated mucin-type O-glycosylation in addition to one N-glycan attached to Asn15. Furthermore, the N terminus was found to be subject to limited proteolysis, resulting in two membrane-bound C-terminal fragments. N-terminal sequencing of the fragments identified two cleavage sites between Arg³¹ and Leu³² and Pro⁵² and Leu⁵³, which were confirmed by cleavage site and truncation mutants. Metalloproteinase inhibitors were able to inhibit the cleavage, suggesting that it is mediated by a matrix metalloproteinase or a disintegrin and metalloproteinase (ADAM) family member. Most importantly, the N-terminal cleavage was found to occur not only in vitro but also in vivo. Receptor activation mediated by the βAR agonist isoproterenol enhanced the cleavage in a concentration- and time-dependent manner, and it was also enhanced by direct stimulation of protein kinase C and adenylyl cyclase. Mutation of the Arg³¹–Leu³² cleavage site stabilized the mature receptor. We hypothesize that the N-terminal cleavage represents a novel regulatory mechanism of cell surface β₁ARs.
Journal of biological chemistry
|Pages:||28850 - 28861|
|Type of Publication:||
A1 Journal article – refereed
|Field of Science:||
This work was supported in part by the Sigrid Juselius Foundation and Grants 107922 and 127199 from the Academy of Finland. Anne E. Hakalahti was supported by the Finnish Foundation for Cardiovascular Research and the Finnish Medical Society Duodecim.
|Academy of Finland Grant Number:||
107922 (Academy of Finland Funding decision)
127199 (Academy of Finland Funding decision)
This research was originally published in the Journal of Biological Chemistry. Hakalahti, A. E., Vierimaa, M. M., Lilja, M. K., Kumpula, E.-P., Tuusa, J. T., & Petäjä-Repo, U. E. Human β1-Adrenergic Receptor Is Subject to Constitutive and Regulated N-terminal Cleavage. J. Biol. Chem. 2010; 285:28850–28861. © by The American Society for Biochemistry and Molecular Biology.