Functional analysis of mutations in the transcription terminator T1 that suppress two dnaG alleles in Escherichia coli.

The mutations dnaG2903 and parB are both temperature-sensitive conditional lethal alleles of the Escherichia coli dnaG gene, which encodes the protein primase. The lesions are located in the 3' end of the gene, 9 basepairs apart, and both cause Glu-to-Lys substitutions in the carboxy terminus of primase. Previously, it was shown that dnaG2903 can be suppressed by point mutations in the rho-independent transcription terminator T1, which is located just upstream of dnaG in the rpsU-dnaG-rpoD macromolecular synthesis operon. We report here that parB can also be suppressed by point mutations in T1, demonstrating that parB can be suppressed in the same manner as dnaG2903. We also identified additional suppressors of dnaG2903 that are point mutations in T1, suggesting that defective transcription termination leading to overexpression of dnaG2903 and parB suppresses the temperature-sensitive phenotype of strains harboring these mutations. Utilizing two mutant rpoB alleles whose transcription termination phenotypes at rho-independent terminators have been previously characterized, we demonstrate that defective transcription termination leading to the overexpression of dnaG does indeed suppress dnaG2903 and parB. The point mutations in T1 identified in this study were analyzed for their effects on termination efficiency at T1. Our results indicate that the thermodynamic stability of the hairpin structures may not be the sole determinant of termination efficiency in vivo.