Researchers have discovered that mammalian cells can convert RNA sequences back into DNA using Polθ, which challenges long-believed notions surrounding polymerases.
Polymerases were thought to only work in a single direction: DNA into DNA or RNA. This prevents RNA messages from being rewritten back into genomic DNA.
Recently, researchers at Thomas Jefferson University have provided the first evidence that RNA segments can in fact be written back into DNA. Dr. Pomerantz and his team began investigating a unique DNA polymerase-helicase fusion protein, called polymerase theta (Polθ). This polymerase is found in higher eukaryotes.
The Polθ is unlike most DNA polymerase I enzymes, as it is:
- Highly error-prone and promiscuous.
- Performs translesion synthesis (TLS) opposite DNA lesions.
- Facilitates microhomology-mediated end-joining (MMEJ) of double strand breaks (DSBs).
Polθ is not expressed in most tissues, but it is highly expressed in many cancer cells and therefore corresponds to poor clinical outcomes. It also shows resistance to genotoxic cancer therapies and promotes the survival of cells that are deficient in DNA damage response pathways. Consequently, this polymerase has the potential to become a promising cancer drug target.
The researchers at Thomas Jefferson University tested Polθ against the reverse transcriptase from HIV, which acts as a DNA polymerase and can also read RNA back into a DNA strand.
It was discovered that Polθ converted RNA messages into DNA just as well as HIV reverse transcriptase did. In fact, Polθ was actually better at duplicating DNA to DNA. Furthermore, it was found that Polθ became significantly more accurate when using an RNA template to write new DNA messages than when duplicating DNA into DNA. This infers that the probable primary function of the enzyme in the cell is to write RNA sequences into DNA.
The key results highlighted in the paper were:
- Polθ exhibits RNA-dependent DNA synthesis activity.
- The polymerase exhibits higher velocity and fidelity of deoxyribonucleotide incorporation on RNA.
- Ternary structure of Polθ on a DNA/RNA primer-template showed that thumb subdomain undergoes a major reconfiguration.
- Polθ promotes RNA-templated DNA repair.
Overall, the study revealed that Polθ had an extraordinary degree of structural plasticity that enabled it to efficiently transcribe template ribonucleotides. This in turn, accommodated reverse transcriptase activity.
The fact that Polθ showed structural-functional characteristics that have not previously been observed in other DNA polymerases or retroviral reverse transcriptase’s is hugely exciting. These findings raise several other questions and open the door for many follow-up investigations, given that this promiscuous enzyme can act on a variety of different templates and has a unique predominant mechanism of promoting RNA-DNA repair.
Dr Pomerantz explained:
“In healthy cells, the purpose of this molecule may be to enable 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.”
Image credit: chfhonk FreePik