Cloning of a putative G-protein-coupled receptor from Arabidopsis thaliana.
We have cloned and characterized a cDNA from Arabidopsis thaliana that most likely encodes a novel member of the vast superfamily of G-protein-coupled receptor proteins (GPCRs). By taking advantage of amino acid sequence similarities between plant expressed sequence tags (ESTs) and established G-protein-coupled receptor sequences, a probe was obtained which was used for the screening of an Arabidopsis cDNA library. The cDNA which was found is very infrequently represented in the cDNA library, suggesting a low and/or spatially restricted expression. A region of the translated sequence of the cDNA shows the highest similarity to cAMP receptors from the slime mold Dictyostelium discoideum. The same region is also similar to that in members of the animal calcitonin family of receptors. Another region of the putative receptor, however, is similar to sequences of serotonin receptors and other receptors of the so-called rhodopsin family of GPCRs. The rhodopsin family has numerous members in higher vertebrate species. Alignments and phylogenetic analyses of the regions of similarity yielded results in accordance with other evolutionary considerations. Our cDNA thus occurred on a distinct major branch in relation to the rest of the rhodopsin family. In relation to the calcitonin family, our cDNA and cAMP receptors occurred together on a distinct major branch but appear to have diverged from each other shortly after their divergence from the rest of the calcitonin family. Other features further argue for a tentative identification of it as a GPCR. It displays seven discrete and strongly predicted transmembrane domains when analyzed in hydropathy plots. The preferred orientation is with the amino terminus towards the outside. It has one Cys residue in extracellular loop 1 and another in extracellular loop 2. Cys residues in these loops are known to form disulfide bridges in many other GPCRs. Finally, it has several fully conserved amino acids that belong to the most conserved in previously known GPCRs, that occur in the above regions of similarity.