Receptors activate adenylyl cyclases through the Gα_s subunit. Previous studies from our laboratory have shown in certain cell types that express adenylyl cyclase 6 (AC6), heterologous desensitization included reduction of the capability of adenylyl cyclases to be stimulated by Gα_s. Here we further analyze protein kinase A (PKA) effects on adenylyl cyclases. PKA treatment of recombinant AC6 in insect cell membranes results in a selective loss of stimulation by high (>10 nM) concentrations of Gα_s. Similar treatment of AC1 or AC2 did not affect Gα_s stimulation. Conversion of Ser-674 in AC6 to an Ala blocks PKA phosphorylation and PKA-mediated loss of Gα_s stimulation. A peptide encoding the region 660–682 of AC6 blocks stimulation of AC6 and AC2 by high concentrations of Gα_s. Substitution of Ser-674 to Asp in the peptide renders the peptide ineffective, indicating that the region 660–682 of AC6 is involved in regulation of signal transfer from Gα_s. This region contains a conserved motif present in most adenylyl cyclases; however, the PKA phosphorylation site is unique to members of the AC6 family. These observations suggest a mechanism of how isoform selective regulatory diversity can be obtained within conserved regions involved in signal communication.