Cloning of type 1 cannabinoid receptor in Rana esculenta reveals differences between genomic sequence and cDNA.
The endocannabinoid system is a conserved system involved in the modulation of several physiologic processes, from the activity of the central nervous system to reproduction. Type 1 cannabinoid receptor (CNR1) cDNA was cloned from the brain and testis of the anuran amphibian, the frog Rana esculenta. Nucleotide identity ranging from 62.6% to 81.9% is observed among vertebrates. The reading frame encoded a protein of 462 amino acids (FCNR1) with all the properties of a membrane G-coupled receptor. Alignments of FCNR1 with those of other vertebrates revealed amino acid identity ranging from 61.9% to 88.1%; critical domains for CNR1 functionality were conserved in the frog. As nucleotide differences of cnr1 cDNA were observed in brain and testis, the genomic sequence of the cnr1 gene was also determined in the same tissue preparations. Nucleotide changes in codons 5, 30, 70, 186, 252 and 408 were observed when cDNA and genomic DNA were compared; the nucleotide differences did not affect the predicted amino acid sequences, except for changes in codons 70 and 408. Interestingly, the predicted RNA folding was strongly affected by different nucleotide sequences. Comparison of cnr1 mRNA sequences available in GenBank with the corresponding genomic sequences revealed that also in human, rat, zebrafish and pufferfish, nucleotide changes between mRNA and genomic sequences occurred. Furthermore, amino acid sequences deduced from both mRNA and the genome were compared among vertebrates, and also in pufferfish the nucleotide changes corresponded to modifications in the amino acid sequence. The present results indicate for the first time that changes in nucleotides may occur in cnr1 mRNA maturation and that this phenomenon might not be restricted to the frog.
Meccariello
Rosaria
R
Dipartimento di Studi delle Istituzioni e dei Sistemi Territoriali, Università di Napoli Parthenope, Naples, Italy.
Chianese
Rosanna
R
Cobellis
Gilda
G
Pierantoni
Riccardo
R
Fasano
Silvia
S
D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D015139:Blotting, Southern; D001923:Brain Chemistry; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry*; D016678:Genome; D008297:Male; D008969:Molecular Sequence Data; D010802:Phylogeny; D011893:Rana esculenta; D043884:Receptor, Cannabinoid, CB1 / Q000235:genetics*; D016415:Sequence Alignment; D013737:Testis / Q000737:chemistry
D018076:DNA, Complementary; D043884:Receptor, Cannabinoid, CB1