Parathyroid hormone-related peptide (PTHrP) is a mediator of cellular growth and differentiation as well as a cause of malignancy-induced hypercalcemia. Most of the actions of PTHrP have been attributed to its interaction with a specific cell surface receptor that binds the N-terminal domain of the protein. Here we present evidence that PTHrP promotes some of its cellular effects by translocating to the nucleolus. Localization of transiently expressed PTHrP to the nucleolus was dependent on the presence of a highly basic region at the carboxyl terminus of the molecule that bears homology to nucleolar targeting sequences identified within human retroviral (human immunodeficiency virus type 1 and human T-cell leukemia virus type 1) regulatory proteins. Endogenous PTHrP also localized to the nucleolus in osseous cells in vitro and in vivo. Moreover, expression of PTHrP in chondrocytic cells (CFK2) delayed apoptosis induced by serum deprivation, and this effect depended on the presence of an intact nucleolar targeting signal. The present findings demonstrate a unique intracellular mode of PTHrP action and a novel mechanism by which this peptide growth factor may modulate programmed cell death.