Dystroglycan (DG) is an extracellular matrix receptor implicated in muscular dystrophies and cancers. DG belongs to the membrane-tethered mucin family and is composed of extracellular (alpha-DG) and transmembrane (beta-DG) subunits stably coupled at the cell surface. These two subunits are generated by autoproteolysis of a monomeric precursor within a distinctive protein motif called sea urchin-enterokinase-agrin (SEA) domain, yet the purpose of this cleavage and heterodimer creation is uncertain. In this study, we identify a functional nuclear localization signal within beta-DG and show that, in addition to associating with alpha-DG at the cell surface, the full-length and glycosylated beta-DG autonomously traffics to the cytoplasm and nucleoplasm in a process that occurs independent of alpha-DG ligand binding. The trafficking pattern of beta-DG mirrors that of MUC1-C, the transmembrane subunit of the related MUC1 oncoprotein, also a heterodimeric membrane-tethered mucin created by SEA autoproteolysis. We show that the transmembrane subunits of both MUC1 and DG transit the secretory pathway prior to nuclear targeting and that their monomeric precursors maintain the capacity for nuclear trafficking. A screen of breast carcinoma cell lines of distinct pathophysiological origins revealed considerable variability in the nuclear partitioning of beta-DG, indicating that nuclear localization of beta-DG is regulated, albeit independent of extracellular ligand binding. These findings point to novel intracellular functions for beta-DG, with possible disease implications. They also reveal an evolutionarily conserved role for SEA autoproteolysis, serving to enable independent functions of mucin transmembrane subunits, enacted by a shared and poorly understood pathway of segregated subunit trafficking.