The Coding and Noncoding Architecture of the Caulobacter crescentus Genome

Abstract

Caulobacter crescentus undergoes an asymmetric cell division controlled by a genetic circuit that cycles in space and time. We provide a universal strategy for defining the coding potential of bacterial genomes by applying ribosome profiling, RNA-seq, global 5′-RACE, and liquid chromatography coupled with tandem mass spectrometry (LC-MS) data to the 4-megabase C. crescentus genome. We mapped transcript units at single base-pair resolution using RNA-seq together with global 5′-RACE. Additionally, using ribosome profiling and LC-MS, we mapped translation start sites and coding regions with near complete coverage. We found most start codons lacked corresponding Shine-Dalgarno sites although ribosomes were observed to pause at internal Shine-Dalgarno sites within the coding DNA sequence (CDS). These data suggest a more prevalent use of the Shine-Dalgarno sequence for ribosome pausing rather than translation initiation in C. crescentus. Overall 19% of the transcribed and translated genomic elements were newly identified or significantly improved by this approach, providing a valuable genomic resource to elucidate the complete C. crescentus genetic circuitry that controls asymmetric cell division.

Author Summary

Caulobacter crescentus is a model system for studying asymmetric cell division, a fundamental process that, through differential gene expression in the two daughter cells, enables the generation of cells with different fates. To explore how the genome directs and maintains asymmetry upon cell division, we performed a coordinated analysis of multiple genomic and proteomic datasets to identify the RNA and protein coding features in the C. crescentus genome. Our integrated analysis identifies many new genetic regulatory elements, adding significant regulatory complexity to the C. crescentus genome. Surprisingly, 75.4% of protein coding genes lack a canonical translation initiation sequence motif (the Shine-Dalgarno site) which hybridizes to the 3′ end of the ribosomal RNA allowing translation initiation. We find Shine-Dalgarno sites primarily inside of genes where they cause translating ribosomes to pause, possibly allowing nascent proteins to correctly fold. With our detailed map of genomic transcription and translation elements, a systems view of the genetic network that controls asymmetric cell division is within reach.