Lon recognition of the replication initiator DnaA is not confined to a single degron

Abstract

DnaA initiates chromosome replication in bacteria. In Caulobacter crescentus, the Lon protease degrades DnaA to coordinate replication with nutrient availability and to halt the cell cycle during acute stress. Here we characterize the mechanism of DnaA recognition by Lon. We find that the native folded state of DnaA is crucial for its degradation, in contrast to the well-known role of Lon in degrading misfolded proteins. We fail to identify a single degradation motif (degron) sufficient for DnaA degradation, rather we show that both the ATPase domain and a species-specific N-terminal motif are important for productive Lon degradation of DnaA. Mutations in either of these determinants disrupt DnaA degradation in vitro and in vivo. DnaA switches from an inactive to active state depending on its nucleotide state and we find that locking DnaA in an active state inhibits degradation. Our working model is that Lon engages DnaA through at least two elements, one of which anchors DnaA to Lon and the other acting as an initiation site for degradation.