Molecular cloning and expression of a novel type V adenylyl cyclase from rabbit myocardium

Abstract

A cDNA of a novel form of type V adenylyl cyclase has been cloned from rabtut myocardmm using ohgonucleotlde probes denved from peptides that were produced by enzymatic cleavage of punfied heart cyclase A correspondmg mRNA (6 kb) has been detected m rabtnt myocarchal tissue by Northern blot analysis The cDNA encodes a protein of 1,264 ammo acids exhlbltmg 12 putatwe membrane-spanning regons m its hydroplnhclty profile Sequence comparison to two other previously published type V adenylyl cyclases reveals ammo-terminal domams of different length and low correlative homology, whereas the rest of the sequences IS almost identical The nonconserved ammo-terminal region of the subtype consists of 214 ammo acids and exceeds the length of the others by 40 and 80 residues, respectively Its presence m membrane preparations from different tissues has been confirmed immunologically usmg an antibody directed against a synthetic peptide The cloned adenylyl cyclase was functionally expressed m COS-1 cells to attam an enzymatrc actlvlty 3 S-to 1Cfold above control m the presence of forskohn

Identity of adenylyl cyclase isoform determines the G protein mediating chronic opioid‐induced adenylyl cyclase supersensitivity

“…However, such subtype-specific differences appear plausible as comparison of the primary sequences of a putative G i a1 binding region on AC type V (C1 region) revealed some structural heterogeneity between AC types I, V and VI (Tang et al 1991;Wallach et al 1994;Dessauer et al 1998). In this respect, G o a has been previously shown to specifically interact with AC type I (Taussig et al 1994).…”

“…The following DNAs were used: rat l-OR (Chen et al 1993), rabbit AC type V (Wallach et al 1994), rat AC type VI (Krupinski et al 1992), and rat G z a (Wong et al 1992). All cDNAs were subcloned into expression vector pcDNA3.1 (Invitrogen, Karlsruhe, Germany) using standard molecular biology techniques.…”

Functional Role of Tryptophan Residues in the Fourth Transmembrane Domainof the CB2 Cannabinoid Receptor

“…Rabbit AC type V (AC-V) and ␤-galactosidase cDNAs in the pXMD1 vector (Wallach et al, 1994) were described previously (Avidor-Reiss et al, 1997;Rhee et al, 1998). The human CB 2 cDNA in pCDM8 was kindly provided by Dr. S. Munro (Cambridge, U.K.).…”

Determination and cellular localization of adenylyl cyclase isozymes expressed in embryonic chick heart

“…As mammalian types V and VI adenylyl cyelase are inhibited by submicromolar Ca 2+ concentrations [7][8][9][10], and appear to be the maj or adenylyl cyclase isozymes expressed in mammalian heart [7,9,11], we postulated the embryonic chick myocytes express avian type V and/or type VI isozymes [14]. We now report that: (i) RT-PCR detects type V and VI messages in mRNA isolated from preparations of isolated myocytes, and (ii) RNase protection assays suggest that type V message in 'myocyte RNA' is 4-5 times that of type VI message.…”

“…One was chick AC V (2 clones) which had 97.2% identity with all current known mammalian type V isoforrns at the amino acid level [9][10][11]19], and 86.1% identity with all current known mammalian type VI isoforms [7,8,11,20]. The other was chick AC VI (15 clones), which had 91.7% identity with all current known mammalian type VI isoforms at the amino acid level [7,8,11,20] and had 84.7% identity with all current known mammalian type V isoforms [9][10][11]19]. The identity between the PCR amplified sequences of chick AC V and AC VI was 86.1% at the amino acid level and 75.1% at the nucleotide acid level.…”