Mutation of the third intracellular loop of the cAMP receptor, cAR1, of Dictyostelium yields mutants impaired in multiple signaling pathways.
Seven-membrane span receptors transduce a wide range of signals across the plasma membrane. One member of this family, the cAMP receptor, cAR1, of Dictyostelium, mediates some responses (e.g. adenylyl cyclase activation, multicellular aggregation) which require G-proteins and others (e.g. Ca2+ influx, loss of ligand binding, cAR1 phosphorylation) which appear to be G-protein-independent. In this study, we randomly mutagenized the NH2-terminal eight amino acids of the third intracellular loop of cAR1 and examined the ability of these mutants to exhibit the three G-protein-independent responses listed above. Most mutants (classes I, II) exhibited wild-type or midly defective responses. Several mutants (class III), however, were severely impaired in all three processes but not in cAMP binding. Furthermore, these mutants failed to couple productively with G-proteins and could not replace cAR1 in a car1- cell. For these reasons, we propose that class III mutations interfere with the formation of an "active" conformation of the receptor.
Caterina
M J
MJ
Department of Biological Chemistry, Johns Hopkins University, School of Medicine, Baltimore, Maryland 21205-2185.
Milne
J L
JL
Devreotes
P N
PN
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001692:Biological Transport; D002118:Calcium / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D004252:DNA Mutational Analysis; D004023:Dictyostelium; D019204:GTP-Binding Proteins / Q000502:physiology; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D010766:Phosphorylation; D017354:Point Mutation; D017434:Protein Structure, Tertiary; D011953:Receptors, Cyclic AMP / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D013329:Structure-Activity Relationship
D009838:Oligodeoxyribonucleotides; D011953:Receptors, Cyclic AMP; D000242:Cyclic AMP; D019204:GTP-Binding Proteins; D002118:Calcium