Sloppy Repair Helps Tuberculosis Bug Resist Drugs

WASHINGTON -- Shoddy work by a DNA-repair enzyme allows tuberculosis-causing bacteria to develop antibiotic resistance, scientists at the National Institute of Allergy and Infectious Diseases (NIAID) have discovered. Reported in the current issue of the journal Cell, the finding by Clifton E. Barry, III, PhD, and his colleagues in South Africa, could lead to new ways to treat TB without risking the development of drug resistance.

"Tuberculosis takes the lives of almost two million people each year, and eight million people develop active TB annually," says NIAID Director Anthony S. Fauci, MD. "Especially alarming is the upsurge in cases of multidrug-resistant tuberculosis. A clearer understanding of how TB bacteria acquire drug resistance is essential if we are to control this disease," he adds.

Invading microbes, including the TB agent Mycobacterium tuberculosis (MTb), must withstand attacks by the host's immune system, adverse physical conditions, and, often, assaults by antibiotics and other drugs. Bacterial DNA damaged in the fray can be repaired by enzymes. Some bacterial DNA repair enzymes, though, are error-prone and often introduce mutations into the DNA strand. These mutations increase the odds of generating strains resistant to antibiotics.

During her research stints in both Barry's lab and the laboratory of Valerie Mizrahi, PhD, at the University of Witwatersrand in South Africa, co-author Helena Boshoff, PhD, worked to uncover details of MTb's DNA repair process. She used ultraviolet light to mimic the DNA damage suffered by MTb as it invades its host, and then she examined how MTb responded. Boshoff determined that MTb uses a DNA polymerase, DnaE2, to repair its DNA.

"We were surprised to find that MTb uses an enzyme from the major DNA polymerase replication family. In other organisms, including humans, such DNA polymerases are responsible for making perfect copies of DNA before the cell divides. Other DNA polymerases in this family are like straight-A students; they perform almost flawlessly. This is the first error-prone DNA polymerase from this family to be identified," says Barry.

MTb has two copies of the gene encoding DnaE enzyme. Previously, it was a mystery why the bacterium needed multiple copies of its DNA replication enzyme gene. With the realization that MTb relies on DnaE2 enzyme to introduce mutations into its DNA, thereby increasing the chance that drug resistance will result, this riddle is solved.

To learn what role the newly identified enzyme plays in animals, the researchers infected mice with either normal MTb or MTb lacking the DnaE2 gene. Mice infected with the normal MTb died quickly, while mice infected with the mutant germ contained the infection more successfully, indicating a role for DnaE2 in helping MTb persist in the host's cells. Finally, the researchers used mice to evaluate DnaE2's role in the evolution of drug resistance. Confirming findings from the test-tube experiments, mice infected with wild-type MTb developed resistance to a common antibiotic, while mice infected with strains lacking the gene - and hence, the error-prone repair enzyme - did not develop antibiotic resistance.

This new insight into the emergence of MTb drug resistance suggests ways to intervene with drugs targeted specifically at MTb's vital DNA repair enzyme. "For example," notes Barry, "therapies targeted at DnaE2 could block the development of drug resistance in people infected with drug-susceptible bacteria. Such a drug might also more efficiently clear non-replicating MTb."

NIAID is a component of the National Institutes of Health (NIH), which is an agency of the Department of Health and Human Services. NIAID supports basic and applied research to prevent, diagnose, and treat infectious and immune-mediated illnesses, including HIV/AIDS and other sexually transmitted diseases, illness from potential agents of bioterrorism, tuberculosis, malaria, autoimmune disorders, asthma and allergies.

Reference: H I M Boshoff et al. DnaE2-mediated inducible mutagenesis plays a role in vivo persistence and the emergence of drug resistance in Mycobacterium tuberculosis. Cell 113(2):183-93 (2003).