The goal of this proposal is to define the regulatory mechanisms that control the bacterial cell cycle and to understand how these mechanisms function within an integrated system. We have shown that the control of the Caulobacter crescentus cell cycle incorporates discrete transcription patterns, controlled proteolytic events that clear the cell of critical regulatory proteins at defined times in the cell cycle, and dynamic localization of polar phospho- signaling proteins that orchestrates the activity of cell cycle regulators. A transcriptional network governed by three global regulators (CtrA, GcrA, DnaA) that oscillate out of phase temporally and spatially has been shown to regulate the temporal expression of approximately 200 genes. This transcriptional network influences and is influenced by the subcellular organization of the cell and its progressive changes during the cell cycle. Cellular compartmentalization preceding cell division is a critical signal for the differential activation of specific proteolytic events in the daughter cells. The CIpXP protease, responsible for the timely degradation of the CtrA global regulator, must be localized at the cell pole with its CtrA substrate for this function. Further, the timing of CIpXP localization is controlled by signaling from the transiently localized CckA histidine kinase that also mediates the activation of CtrA at the correct time in the cell cycle, thus creating a robust transcriptional network. We will focus on identifying the factors responsible for protein localization and the dynamic regulation of the proteolytic mechanisms that play a fundamental role in cell cycle progression, and identifying the factors that contribute to and integrate cell cycle regulatory networks. In addition, because free living organisms must readily adapt to a changing environment, it is critical to understand how this regulatory circuitry works not only under optimal conditions, but also how it is re-programmed when the cell is challenged;and how, when conditions are favorable again, it begins anew. We will explore individual regulatory mechanisms in depth while determining how they function in an integrated system that operates in time and space to carry out the functions of a living cell.