An increasing challenge in the control of tuberculosis (TB) worldwide is the emergence and spread of drug resistance in Mycobacterium tuberculosis (Mtb), the causative agent of TB. Evolution of drug resistance in Mtb is associated with chromosomal mutations, not with the acquisition of resistance plasmids or transferred resistance genes. DnaE2 is required for induced mutagenesis in Mtb and belongs to the C-family of bacterial DNA polymerases that are responsible for genome replication. dnaE2 is a component of the imuA-imuB-dnaE2 cassette and all three genes are essential for DNA damage-induced mutagenesis. The DnaE2-ImuA-ImuB protein complex is proposed to associate with the sliding clamp processivity factor (the ?-subunit of DNA pol III) through interactions with the ImuB subunit, allowing DnaE2 access to sites of replication where it can perform error-prone DNA synthesis. However, this enzyme complex has not been well characterized. We hypothesize that error prone DNA synthesis by the DnaE2-ImuA-ImuB polymerase complex contributes to drug resistance during TB infection. Here, we will (Aim 1) evaluate the importance of the complex for Mtb survival, mutagenesis and drug resistance in host-associated stress conditions, and (Aim 2) characterize the assembly, catalytic activity and mutational properties of this putative error-prone polymerase. These studies are essential steps towards understanding the mechanistic bases of drug resistance emergence in Mtb and the potential use of this error-prone DNA polymerase complex as a target for novel adjunctive therapies to combat drug- resistance in Mtb.
Tuberculosis (TB) is a major global health problem, and new intervention strategies are urgently needed to reduce its deadly impact. Increasing levels of drug resistance especially threaten to derail TB control, making it critical to understand and treat the underlying mechanisms of resistance evolution. This work will develop robust tools to characterize mechanisms underlying evolution of drug resistance in Mtb, while specifically evaluating the relative contributions of the host environment and an error prone DNA polymerase in Mtb, which may have potential as a new adjunctive therapeutic target for TB intervention.
