The budding yeast Cdc6 protein (Cdc6p) is essential for formation of pre-replicative complexes (pre-RCs) at origins of DNA replication. Regulation of pre-RC assembly plays a key role in making initiation of DNA synthesis dependent upon passage through mitosis and in limiting DNA replication to once per cell cycle. Cdc6p is normally only present at high levels during the G1 phase of the cell cycle. This is partly because the CDC6 gene is only transcribed during G1. In this article we show that rapid degradation of Cdc6p also contributes to this periodicity. Cdc6p degradation rates are regulated during the cell cycle, reaching a peak during late G1/early S phase. Removal of a 47-amino-acid domain near the N-terminus of Cdc6p prevents degradation of Cdc6p. Likewise, mutations in the Cdc4/34/53 pathway involved in ubiquitin-mediated degradation block proteolysis and genetic evidence is presented indicating that the N-terminus of Cdc6p interacts with the Cdc4/34/53 pathway, probably through Cdc4p. A stable Cdc6p mutant which is no longer degraded by the Cdc4/34/53 pathway is, none the less, fully functional. Constitutive overexpression of either wild-type or stable Cdc6p does not induce re-replication and does not induce assembly of pre-replicative complexes after DNA replication is complete.