GAGE cancer-germline antigens are recruited to the nuclear envelope by germ cell-less (GCL).

GAGE proteins are highly similar, primate-specific molecules with unique primary structure and undefined cellular roles. They are restricted to cells of the germ line in adult healthy individuals, but are broadly expressed in a wide range of cancers. In a yeast two-hybrid screen we identified the metazoan transcriptional regulator, Germ cell-less (GCL), as an interaction partner of GAGE12I. GCL directly binds LEM-domain proteins (LAP2β, emerin, MAN1) at the nuclear envelope, and we found that GAGE proteins were recruited to the nuclear envelope inner membrane by GCL. Based on yeast two-hybrid analysis and pull-down experiments of GCL polypeptides, GCL residues 209-320 (which includes the BACK domain) were deduced sufficient for association with GAGE proteins. GAGE mRNAs and GCL mRNA were demonstrated in human testis and most types of cancers, and at the protein level GAGE members and GCL were co-expressed in cancer cell lines. Structural studies of GAGE proteins revealed no distinct secondary or tertiary structure, suggesting they are intrinsically disordered. Interestingly GAGE proteins formed stable complexes with dsDNA in vitro at physiological concentrations, and GAGE12I bound several different dsDNA fragments, suggesting sequence-nonspecific binding. Dual association of GAGE family members with GCL at the nuclear envelope inner membrane in cells, and with dsDNA in vitro, implicate GAGE proteins in chromatin regulation in germ cells and cancer cells.