Previous work has identified the parathyroid hormone–related protein (PTHrP) nucleolar targeting signal (NTS) as both necessary and sufficient for localization of PTHrP to the nucleus and nucleolus of a variety of cells where it is believed to participate in the regulation of cell proliferation, differentiation, and apoptotic cell death. The mechanism whereby a secreted peptide, such as PTHrP, gains access to the nuclear compartment remains a question of debate. The current work examines the possibility that exogenous PTHrP is internalized and transported to the nuclear compartment by a mechanism that is dependent on preservation of the PTHrP NTS. Transiently expressed, PTHrP(1–141) was detected at the cell surface as well as in the cytoplasmic and nuclear compartments of COS‐1 cells. Deletion of the NTS, or mutation of the conserved GxKKxxK motif within the NTS, effectively prevented both cell‐surface binding and nuclear/nucleolar accumulation of PTHrP(1–141). A biotinylated peptide corresponding to the PTHrP NTS (PTHrP‐NTS‐biotin) was internalized and translocated to the nucleus and nucleolus in a time‐, temperature‐, and concentration‐dependent manner, whereas a peptide representing a similar bipartite NTS from Nucleolin was not. Internalization and nucleolar targeting of PTHrP‐NTS‐biotin were indistinguishable in CFK2 cells, which express the common PTH/PTHrP receptor, and in 27m21 cells, which do not. In addition, pretreatment with a saturating dose of synthetic PTHrP(74–113) was capable of abrogating nucleolar accumulation of the PTHrP‐NTS peptide, whereas pretreatment with PTHrP(1–34) or PTHrP(67–86) was not. These observations demonstrate that binding of exogenous, full‐length PTHrP to the cell surface is mediated through a conserved motif embedded in the NTS and suggest that internalization and nucleolar targeting of an NTS peptide are mediated through binding to a cell surface protein distinct from the PTH/PTHrP receptor. In total, the data support the hypothesis that secreted PTHrP(1–141) can be endocytosed and targeted to the nucleolus through a mechanism that is dependent on preservation of a core motif within the PTHrP NTS.