Multiple chromosomes in bacteria: The Yin and Yang of trp gene localization in Rhodobacter sphaeroides 2.4.1

The existence of multiple chromosomes in bacteria has been known for some t ime. Yet the extent of functional solidarity between different chromosomes remains unknown. To examine this question, we have surveyed the well-descri bed genes of the tryptophan biosynthetic pathway in the multichromosomal ph otosynthetic eubacterium Rhodobacter sphaeroides 2.4.1. The genome of this organism was mutagenized using Tn5, and strains that were auxotrophic for t ryptophan (Trp(-)) were isolated. Pulsed-field gel mapping indicated that T n5 insertions in both the large (3 Mb CI) and the small (0.9 Mb CII) chromo somes created a Trp- phenotype. Sequencing the DNA flanking the sites of th e Tn5 insertions indicated that the genes trkE-yibQ-trpGDC were at a locus on CI, while genes trpF-aroR-trpB were at locus on CII. Unexpectedly, trpA was not found downstream of trpB. Instead, it was placed on the CI physical map at a locus 1.23 Mb away from trpE-yibQ-trpGDC. To relate the context o f the R. sphaeroides trp genes to those of other bacteria, the DNA regions surrounding the trp genes on both chromosomes were sequenced. Of particular significance was the finding that rpsA1, which encodes ribosomal protein S 1, and cmkA, which encodes cytidylate monophosphate kinase, were on CII. Th ese genes are considered essential for translation and chromosome replicati on, respectively. Southern blotting suggested that the trp genes and rpsA1 exist in single copy within the genome. To date, this topological organizat ion of the trp "operon" is unique within a bacterial genome. When taken wit h the finding that CII encodes essential housekeeping functions, the overal l impression is one of close regulatory and functional integration between these chromosomes.