Molecular analysis of mutations in Agrobacterium tumefaciens under selection pressure

Abstract

Mutations may occur as point mutations and insertion mutations in a cell. Bacteria often rely on mutations to survive a harsh environment. However, it is not clear how point and insertion mutations are coordinated in the event of selection pressure. In this study, we developed a genetic assay that can monitor both types of mutations simultaneously in Agrobacterium tumefaciens. It was found that A. tumefaciens gave rise to tetracycline resistance (Tcr) at a high frequency; both point mutation and transposon insertion at tetR may give rise to Tcr. It is found that a mutation at the sensor protein ChvG led to a dramatic increase in insertional mutation frequency but a dramatic decrease in point mutation frequency. ChvG affected the mutation to Tcr independent of the DNA repair protein MutS, since the chvG-mutS double mutant did not show the phenotype of chvG or mutS. Generation and screening of the mutant libraries led to the identification of two other mutants 715 and Tcm3, which also exhibited a dramatic increase in insertional mutation frequency but a dramatic decrease in point mutation frequency. This study provides the first piece of evidence that point mutation is coupled with insertion mutation in A. tumefaciens. his implies that bacteria may have a regulatory system to monitor the success of mutational events.