The C-terminal tail of CRTH2 is a key molecular determinant that constrains Galphai and downstream signaling cascade activation.
Prostaglandin D(2) activation of the seven-transmembrane receptor CRTH2 regulates numerous cell functions that are important in inflammatory diseases, such as asthma. Despite its disease implication, no studies to date aimed at identifying receptor domains governing signaling and surface expression of human CRTH2. We tested the hypothesis that CRTH2 may take advantage of its C-tail to silence its own signaling and that this mechanism may explain the poor functional responses observed with CRTH2 in heterologous expression systems. Although the C terminus is a critical determinant for retention of CRTH2 at the plasma membrane, the presence of this domain confers a signaling-compromised conformation onto the receptor. Indeed, a mutant receptor lacking the major portion of its C-terminal tail displays paradoxically enhanced Galpha(i) and ERK1/2 activation despite enhanced constitutive and agonist-mediated internalization. Enhanced activation of Galpha(i) proteins and downstream signaling cascades is probably due to the inability of the tail-truncated receptor to recruit beta-arrestin2 and undergo homologous desensitization. Unexpectedly, CRTH2 is not phosphorylated upon agonist-stimulation, a primary mechanism by which GPCR activity is regulated. Dynamic mass redistribution assays, which allow label-free monitoring of all major G protein pathways in real time, confirm that the C terminus inhibits Galpha(i) signaling of CRTH2 but does not encode G protein specificity determinants. We propose that intrinsic CRTH2 inhibition by its C terminus may represent a rather unappreciated strategy employed by a GPCR to specify the extent of G protein activation and that this mechanism may compensate for the absence of the classical phosphorylation-dependent signal attenuation.
Schröder
Ralf
R
Institute of Pharmaceutical Biology, University of Bonn, Nussallee 6, 53115 Bonn, Germany.
Merten
Nicole
N
Mathiesen
Jesper Mosolff
JM
Martini
Lene
L
Kruljac-Letunic
Anamarija
A
Krop
Friederike
F
Blaukat
Andree
A
Fang
Ye
Y
Tran
Elizabeth
E
Ulven
Trond
T
Drewke
Christel
C
Whistler
Jennifer
J
Pardo
Leonardo
L
Gomeza
Jesús
J
Kostenis
Evi
E
D000595:Amino Acid Sequence; D019390:Arrestins / Q000378:metabolism; D002460:Cell Line; D000242:Cyclic AMP / Q000378:metabolism; D066246:ErbB Receptors / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D010766:Phosphorylation; D015230:Prostaglandin D2 / Q000494:pharmacology; D011485:Protein Binding; D011971:Receptors, Immunologic / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011982:Receptors, Prostaglandin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D015398:Signal Transduction; D013379:Substrate Specificity; D013997:Time Factors; D015533:Transcriptional Activation / Q000187:drug effects; D000071557:beta-Arrestins
D019390:Arrestins; D011971:Receptors, Immunologic; D011982:Receptors, Prostaglandin; D000071557:beta-Arrestins; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D000242:Cyclic AMP; D066246:ErbB Receptors; D048049:Extracellular Signal-Regulated MAP Kinases; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D015230:Prostaglandin D2; C036293:prostaglandin D2 receptor