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Abstract

The regulation of peptidoglycan (PG) remodeling has been studied intensively in rod-shaped and coccoid model bacteria such as Escherichia coli and Bacillus subtilis, but the question of how shape arises in bacteria with more complex morphologies remains incompletely understood. Among the morphologically complex species is Caulobacter crescentus, a crescent-shaped stalked α-proteobacterium, which is characterized by a biphasic life cycle and an asymmetric cell division. Cell wall remodeling critically depends on the action of peptidoglycan-degrading enzymes, whose enzymatic activity must be tightly regulated in time and space to prevent cell lysis. In several organisms, including the model E. coli, it has been found that proteins with catalytically inactive LytM domains act as regulators of PG lytic enzymes. C. crescentus possesses two such LytM factors with degenerate LytM domains, DipM and LdpF. These two factors have been individually studied and the data suggest that they act in different stages of cell division and are responsible of the recruitment of different PG lytic enzymes. However, these findings only give a partial explanation of the function of these proteins. The results obtained in this work indicate that while LdpF appears to connect FtsEX with AmiC, DipM interacts in vivo with multiple autolysins (SdpA, SdpB, AmiC and CrbA) and FtsN. We confirmed in vitro the interactions between DipM and SdpA, SdpB, AmiC and FtsN and our data supports the notion that the interaction surface with all these factors is shared. Moreover, we show that the previously reported mid-cell localization dependency of SdpA and SdpB on DipM is most probably indirect and not mediated by their direct interaction. Additionally, DipM was able to stimulate the enzymatic activity of SdpA, SdpB and AmiC in vitro. Finally, our data also suggest a novel mechanism to regulate the activity of AmiC based on its dimerization.