Increase in cell-surface localization of parathyroid hormone receptor by cytoskeletal protein 4.1G.
The cell-surface localization of GPCRs (G-protein-coupled receptors) has emerged as one of critical factors of the GPCR-mediated signal transduction. It has been reported that the C-termini of GPCRs contain the sequences for sorting the receptors to cell surface. In the present study, we have searched for proteins that interact with the C-terminus of PTH (parathyroid hormone)/PTH-related protein receptor (PTHR) by using the yeast two-hybrid system, and identified a cytoskeletal protein 4.1G (generaltype 4.1 protein) as an interactant with the C-terminus. Immunohistochemical study revealed that both PTHR and 4.1G were co-localized on plasma membranes, when they were transiently expressed in COS-7 cells. When 4.1G or the C-terminal domain of 4.1G (4.1G-CTD), a dominant-negative form of 4.1G, was co-expressed with PTHR in COS-7 cells, 4.1G, but not 4.1G-CTD, facilitated the cell-surface localization of PTHR, determined by cell-surface biotinylation assay. PTH-(1-34) caused phosphorylation of ERK (extracellular-signal-regulated kinase) 1/2 in PTHR-expressed cells mainly mediated through EGF (epidermal growth factor) receptor. The phosphorylation was enhanced by the expression of 4.1G, but not 4.1G-CTD. PTH-(1-34) elevated [Ca2+]i (intracellular Ca2+ concentration) independent of EGF receptor activation, and the elevation was enhanced by the expression of 4.1G, but not 4.1G-CTD. These data indicate that 4.1G facilitates the cell-surface localization of PTHR through its interaction with the C-terminus of the receptor, resulting in the potentiation of PTHR-mediated signal transduction.
Saito
Masaki
M
Department of Cellular Signaling, Graduate School of Pharmaceutical Sciences, Tohoku University, Aoba 6-3, Aramaki, Aoba-ku, Sendai 980-8578, Japan.
Sugai
Maki
M
Katsushima
Yuriko
Y
Yanagisawa
Teruyuki
T
Sukegawa
Jun
J
Nakahata
Norimichi
N
D000818:Animals; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D066246:ErbB Receptors / Q000378:metabolism; D005786:Gene Expression Regulation; D006801:Humans; D020930:MAP Kinase Kinase Kinases / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D044168:Receptor, Parathyroid Hormone, Type 1 / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D003598:Cytoskeletal Proteins; D008565:Membrane Proteins; D044168:Receptor, Parathyroid Hormone, Type 1; C028841:erythrocyte membrane band 4.1 protein; D066246:ErbB Receptors; D020930:MAP Kinase Kinase Kinases; D002118:Calcium