Embryonic stem (ES) cells are capable of continuous self-renewal and pluripotential differentiation. A "core" set of transcription factors, Oct4, Sox2, and Nanog, maintains the ES cell state, whereas various combinations of factors, invariably including Oct4 and Sox2, reprogram somatic cells to pluripotency. We have sought to define the transcriptional network controlling pluripotency in mouse ES cells through combined proteomic and genomic approaches. We constructed a protein interaction network surrounding Nanog and determined gene targets of the core and reprogramming factors, plus others. The expanded transcriptional network we have constructed forms the basis for further studies of directed differentiation and lineage reprogramming, and a paradigm for comprehensive elucidation of regulatory pathways in other stem cells.