The presence of gsp mutations at codons 201 and 227 in the gene coding for the alpha subunit of the GTP-binding Gs protein which stimulates adenylyl cyclase (AC) has been investigated in 31 samples of differentiated thyroid tumours, which had been previously characterized with respect to their adenylyl cyclase activity (ACA) before and after stimulation by thyroid-stimulating hormone (TSH). Polymerase chain reaction (PCR) amplification of DNA extracted from these tumours, followed by high stringency oligonucleotide probing, enabled the detection of mutations in three samples originating from tumours with high constitutive ACA, which was not significantly further stimulated by TSH. Two mutations were at codon 227 and replaced Gln227 by His or Lys, and one was at codon 201, with the substitution of Arg201 by Ser. Because thyrocytes belong to the subset of differentiated cells which are programmed to proliferate in response to elevated cAMP levels, the gsp mutations observed in some differentiated thyroid carcinomas probably contributed to their tumorigenic phenotype.