Rasmussen U B, Vouret-Craviari V, Jallat S, Schlesinger Y, Pagès G, Pavirani A, Lecocq J P, Pouysségur J, Van Obberghen-Schilling E
Transgene, S.A., Strasbourg, France.
FEBS Lett. 1991 Aug 19;288(1-2):123-8. doi: 10.1016/0014-5793(91)81017-3.
The serine protease alpha-thrombin (thrombin) potently stimulates G-protein-coupled signaling pathways and DNA synthesis in CCL39 hamster lung fibroblasts. To clone a thrombin receptor cDNA, selective amplification of mRNA sequences displaying homology to the transmembrane domains of G-protein-coupled receptor genes was performed by polymerase chain reaction. Using reverse transcribed poly(A)+ RNA from CCL39 cells and degenerate primers corresponding to conserved regions of several phospholipase C-coupled receptors, three novel putative receptor sequences were identified. One corresponds to an mRNA transcript of 3.4 kb in CCL39 cells and a relatively abundant cDNA. Microinjection of RNA transcribed in vitro from this cDNA in Xenopus oocytes leads to the expression of a functional thrombin receptor. The hamster thrombin receptor consists of 427 amino acid residues with 8 hydrophobic domains, including one at the extreme N-terminus that is likely to represent a signal peptide. A thrombin consensus cleavage site is present in the N-terminal extracellular region of the receptor sequence followed by a negatively charged cluster of residues present in a number of proteins that interact with the anion-binding exosite of thrombin.
