Analysis of extracts of rat adrenal glands showed that the activity of DNA polymerase beta was preferentially decreased upon hypophysectomy, whereas that of DNA polymerase alpha or gamma stayed relatively constant. The adrenal polymerase beta from hypophysectomized rats sedimented at 5-7S through sucrose gradients, whereas that from sham-operated controls sedimented at 3.3-3.6S. The large form of DNA polymerase beta was stable in 0.1 M KCl or 0.5% Nonidet P-40 but was converted into a 3.3S form in 0.5 M KCl. These changes in adrenal DNA polymerase beta were reversed by the daily injection of adrenocorticotropic hormone but were not reversed by exogenous glucocorticoid.