Reverse transcriptases are commonly found in viruses but have been more elusive in mammals, including humans. The polymerases in human cells are generally understood to only be able to transcribe RNA from DNA and not work the other way around, under the central dogma of molecular biology. However, a research team from Thomas Jefferson University has discovered that the human DNA polymerase theta can reverse transcribe DNA just as well as the reverse transcriptase found in HIV.
Researchers used a DNA primer annealed to an RNA template to test the reverse transcription abilities of pol theta compared to HIV reverse transcriptase, as well as DNA polymerase eta, which previous studies found has some RT activity under certain conditions. The study found pol theta was able to reverse transcribe with high efficiency, similarly to HIV RT, while pol eta was unable to reverse transcribe beyond 3 nucleotides under the same conditions.
Researchers also used X-ray crystallography to examine the molecular basis of pol theta’s RT activity and observed how the polymerase changed it structure to accommodate the bulkier RNA molecule, notably by a major reconfiguration of the thumb subdomain, with more than half of thumb subdomain residues refolding from alpha helices to loops.
Similarly to retrovirus RTs, pol theta is highly error-prone when duplicating DNA into DNA, but much more efficient and accurate when using an RNA template, as shown by the study, which was published in Science Advances.
“Our research suggests that polymerase theta’s main function is to act as a reverse transcriptase,” said Richard Pomerantz, lead researcher and associate professor of biochemistry and molecular biology at Thomas Jefferson University. “In healthy cells, the purpose of this molecule may be toward RNA-mediated DNA repair. In unhealthy cells, such as cancer cells, polymerase theta is highly expressed and promotes cancer cell growth and drug resistance. It will be exciting to further understand how polymerase theta’s activity on RNA contributes to DNA repair and cancer-cell proliferation.”
The results of the study suggest that RNA can be used as a template for repairing or re-writing genomic DNA, and could have wide implications affecting many fields of biology and medicine.