1. On to Caulobacter! The stalked bacterium Swarmer cell Stalk cell 0.7 um in diameter 2-3 um long

2. Graphical Representations of the Genome 4,016,942 bp 3767 ORFs (CW and CCW)

3. A Sticky Wicket

4. Shaping a Crescent- Shaped Cell: Crescentin 2 um Elongated stationary phase cells Wild-type Vibriod Cells Transposon Insertion Complementation by Normal Gene

5. Cresecentin Filaments Shape Caulobacter optical sections of stationary phase cells

6. Crescentin is an Intermediate Filament Ortholog

7. MreB Directs Wall Synthesis as in E. coli Xylose-regulatable MreB Turned Off After Switch to Xylose-Free Medium

8. MreB Also Positions the Chromosomes Depletion and Overexpression Both Disturb Partitioning

9. And FtsZ is Involved in Division xylose medium glucose medium complementation by a wild-type ftsZ allele Xylose-regulatable FtsZ 40/60 split

10. Prokaryotes Contain Orthologs of All Three Major Filamentous Networks of the Eukaryotic Cytoskeleton EM scale bars 100nm Light microscope bars 2 um

11. The Caulobacter Life Cycle ~150 min

12. Transcriptome Changes During the Cell Cycle Non-synchronized average level Expression of 16% of the Genome is Cell-Cycle Regulated

13. Turning it on, turning it over Pulse/Chase

14. Cell Cycle Regulation Is Mediated by CtrA CtrA controls 38 direct and 144 indirect targets (~7% of the genome)

15. Proteolysis Frees the ORI Resynthesis Represses the ORIs Cell Cycle Expression of CtrA SW ST

16. Asymmetric Localization of Other Key Determinant Molecules Proteolysis In SW Localize to ST Localize to SW Localize to polesand disperses KinasesKinases

17. Localization of DivJ During the Cell Cycle Localization of DivJ::GFP to the ST pole during division Movie

18. MreB Organizes Polarity

19. Another master Regulator, GcrA, Partitions Oppositely from CtrA GcrA is a Positive Regulator of DNA Replication in ST

20. The CtrA/GcrA Master Cycle SW molecular identity ST molecular identity Literally Sorting out molecular identity Taking Different Paths Proteolysis of CtrA releases GcrA expression CtrA represses GcrA expression GcrA expression turns CtrA expression back on CtrA represses expression of itself and GcrA

21. A Few Questions for Thought Describe the Caulobacter life cycle, noting key cellular landmarks of each stage of development. Extend your discussion of the prokaryotic cytoskeleton from E. coli to include Caulobacter’s crescentin. Propose a model based on the Min operon system for how Caulobacter reproducibly divides in an unequal fashion. Discuss the critical role of CtrA in the developmental of Caulobacter, including a sense of when, where and why this protein is active as well as the rich diversity of targets that it regulates (and the consequences of those interactions). Describe how expression of CtrA and GcrA alternate to drive the developmental cycle of Caulobacter. Explain how differential inheritance of transcription factors and kinases can constrain two daughter cells to different developmental fates.