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The major flagellar proteins, including the flagellins and the hook protein, are synthesized periodically in the Caulobacter crescentus cell cycle at the time of flagellum assembly. Although fla genes are regulated at the transcriptional level [Ohta, N., Chen, L.-S., Swanson, E. & Newton, A. (1985) J. Mol. Biol. 186,[107][108][109][110][111][112][113][114][115], the 5' regulatory regions of C. crescentus genes have not been identified. We describe here the results of nuclease Si protection assays that map the 5' ends of mRNAs synthesized in vivo from transcription units II (hook operon) and I1.1 of the hook gene cluster and locate the corresponding promoter regions P11 and PII.1. The two promoters are regulated with different periodicities in the cell cycle and have different genetic requirements for expression.The failure to detect transcripts from either PI or P11 in Escherichia coli suggests that developmentally regulated promoters of C. crescentus have different recognition sequences from those of E. coli. There is little nucleotide sequence homology between P11 and P11.1. There are, however, three regions of homology between PU and the nucleotide sequence 5' to the 29-kDa-flagellin-related gene, and two of these are in regions of dyad symmetry. We discuss the possibility that DNA-protein interactions at homologous nucleotide sequences like those identified in P11 are part of a regulatory gene cascade that participates in timing fia gene expression in the C. crescentus cell cycle.Caulobacter crescentus has a well-defined pattern of asymmetric cell division that has made it an attractive model system for study of the temporal and spatial regulation of cell differentiation. The stalked cell of this Gram-negative bacterium divides repeatedly to form the old stalked cell plus a new, motile swarmer cell with a set of polar surface structures, including the flagellum, the pili, and the DNA phage receptor sites. The temporal regulation of development in C. crescentus is illustrated by the precise and characteristic times at which these structures are formed during the cell cycle (for review, see refs. 1-3).Evidence We report in this paper preliminary results of nuclease S1 protection assays that map the 5' termini of RNAs synthesized in vivo from transcription units II (hook operon) and 11.1 of C. crescentus. The locations of these 5' RNA termini define promoters PII and PII.1. Transcripts from the two promoters are regulated with different periodicities in the cell cycle and they show different genetic requirements for expression. Analysis of promoters PII and PII.1 and of the sequence 5' of a gene encoding a 29-kDa flagellin-homologous protein (12) (hereafter referred to as the 29-kDaflagellin gene) reveals three regions of sequence homology. The possible role of these sequences in the transcriptional regulation of C. crescentus fla genes is discussed. MATERIALS AND METHODSStrains and Growth Conditions. Wild-type C. crescentus strain CB15 (American Type Culture Collection 19089), motility mutants, and mer...

Previous genetic analysis ofCaulobacter crescentuw showed that the periodic synthesis of hook protein, flagellin A, and flagellin B, the major flagellar subunits, is coupled in some way to chromosome replication. To