Role of the signal peptide in the synthesis and processing of the glucagon-like peptide-1 receptor.
OBJECTIVE
The glucagon-like peptide-1 receptor (GLP-1R) belongs to Family B of the G protein-coupled receptor superfamily and is a target for treatment of type 2 diabetes. Family B G protein-coupled receptors contain a putative N-terminal signal peptide, but its role in receptor synthesis and trafficking are unclear. Further, the signal peptide is not cleaved in at least one family member.


METHODS
We examined receptor glycosylation and the role of the signal peptide in GLP-1R synthesis and trafficking using constructs containing epitope tags at the N- and/or C-terminus and in which the signal peptide sequence was either present or absent.


RESULTS
The signal peptide was absolutely required for GLP-1R synthesis but could be substituted to some extent by increasing positive charge in the N-terminal region of the receptor flanking the signal peptide. The signal peptide is cleaved during synthesis and processing of the receptor. An enhanced GFP-epitope tag at the N-terminus of the receptor permitted synthesis of the receptor but blocked signal peptide cleavage and prevented trafficking to the plasma membrane. Cleavage site mutation allowed synthesis of a full-length receptor, blocked signal peptide cleavage and caused retention within the endoplasmic reticulum.


CONCLUSIONS
Signal peptide cleavage was not essential for receptor synthesis but was obligatory for processing and trafficking of receptors to the plasma membrane. Further, the GLP-1R is subject to N-linked glycosylation and only the mature, fully glycosylated form of the receptor is present in the plasma membrane. Inhibition of glycosylation prevents processing and cell surface expression of the GLP-1R.
Huang
Y
Y
Department of Cell Physiology and Pharmacology, University of Leicester, Leicester, UK.
Wilkinson
G F
GF
Willars
Gary B
GB
D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D000939:Epitopes; D000067757:Glucagon-Like Peptide-1 Receptor; D006031:Glycosylation; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D021382:Protein Sorting Signals; D021381:Protein Transport; D018027:Receptors, Glucagon / Q000096:biosynthesis / Q000378:metabolism*
D000939:Epitopes; C000593833:GLP1R protein, human; D000067757:Glucagon-Like Peptide-1 Receptor; D021382:Protein Sorting Signals; D018027:Receptors, Glucagon; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins