Two distinct cDNAs encoding β subunits of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) were cloned from the cDNA library constructed for the pituitary of the red-bellied newt, Cynops pyrrhogaster, and sequenced. The newt FSHβ and LHβ cDNAs encode polypeptides of 129 and 131 amino acids, including signal peptides of 20 and 19 amino acids, respectively. The number and position of cysteine and N-glycosylation in each of the β subunits of FSH and LH, which are considered essential for assembly of the α subunit, are well conserved between the newt and other tetrapods. The high homology (41.6%) between the β subunits of newt FSH and LH imply less specificity of FSH and LH in gonadal function. One cDNA encoding the common polypeptide chain α subunit of FSH and LH was also isolated from the newt pituitary gland. The mRNAs of FSHβ, LHβ, and the α subunit were expressed only in the pituitary gland among various newt tissues. Double-staining with in situ hybridization and immunohistochemistry revealed coexpression of FSHβ and LHβ in the same newt pituitary cells. Ovariectomy induced a significant increase in FSHβ mRNA levels, but there was no significant change in LHβ or α subunit mRNA levels compared with those in control animals. Taken together, these data suggest that two kinds of gonadotropins, namely FSH and LH, are expressed in the same gonadotropin-producing cells in the pars distalis of the newt as well as in other tetrapods and that the expression of FSHβ is negatively regulated by the ovaries.