The mutations that give rise to melanoma result from a chemical conversion in DNA fueled by sunlight and not just a DNA copying error as previously believed, according to a study (“The major mechanism of melanoma mutations is based on deamination of cytosine in pyrimidine dimers as determined by circle damage sequencing”) by Van Andel Institute scientists in Science Advances.

The findings upend long-held beliefs about the mechanisms underlying the disease, reinforce the importance of prevention efforts and offer a path forward for investigating the origins of other cancer types, according to the researchers.

“Sunlight-associated melanomas carry a unique C-to-T mutation signature. UVB radiation induces cyclobutane pyrimidine dimers (CPDs) as the major form of DNA damage, but the mechanism of how CPDs cause mutations is unclear. To map CPDs at single-base resolution genome wide, we developed the circle damage sequencing (circle-damage-seq) method,” write the investigators.

“In human cells, CPDs form preferentially in a tetranucleotide sequence context (5’-Py-T<>Py-T/A), but this alone does not explain the tumor mutation patterns. To test whether mutations arise at CPDs by cytosine deamination, we specifically mapped UVB-induced cytosine-deaminated CPDs. Transcription start sites (TSSs) were protected from CPDs and deaminated CPDs, but both lesions were enriched immediately upstream of the TSS, suggesting a mutation-promoting role of bound transcription factors. Most importantly, the genomic dinucleotide and trinucleotide sequence specificity of deaminated CPDs matched the prominent mutation signature of melanomas.

“Our data identify the cytosine-deaminated CPD as the leading premutagenic lesion responsible for mutations in melanomas.”

“Cancers result from DNA mutations that allow defective cells to survive and invade other tissues. However, in most cases, the source of these mutations is not clear, which complicates development of therapies and prevention methods,” said Gerd Pfeifer, PhD, a VAI professor and the study’s corresponding author. “In melanoma, we’ve now shown that damage from sunlight primes the DNA by creating ‘premutations’ that then give way to full mutations during DNA replication.”

Most mutations of any cancer

Previous large-scale sequencing studies have shown that melanoma has the most DNA mutations of any cancer. Like other skin cancers, melanoma is linked to sun exposure, specifically UVB radiation which damages skin cells as well as the DNA within cells.

Most cancers are thought to begin when DNA damage directly causes a mutation that is then copied into subsequent generations of cells during normal cellular replication. In the case of melanoma, however, Pfeifer and his team found a different mechanism that produces disease-causing mutations — the introduction of a chemical base not normally found in DNA that makes it prone to mutation.

In melanoma, the problem occurs when UVB radiation hits certain sequences of bases (CC, TT, TC, and CT) causing them to chemically link together and become unstable. The resulting instability causes a chemical change to cytosine that transforms it into uracil, which is found in mRNA but not in DNA. This change, called a “premutation,” primes the DNA to mutate during normal cell replication, thereby causing alterations that underlie melanoma.

These mutations may not cause disease right away; instead, they may lay dormant for years. They also can accumulate as time goes on and a person’s lifetime exposure to sunlight increases, resulting in a tough-to-treat cancer that evades many therapeutic options.

“Safe sun practices are very important. In our study, 10–15 minutes of exposure to UVB light was equivalent to what a person would experience at high noon, and was sufficient to cause premutations,” Pfeifer said. “While our cells have built-in safeguards to repair DNA damage, this process occasionally lets something slip by. Protecting the skin is generally the best bet when it comes to melanoma prevention.”

The findings were made possible using a method developed by Pfeifer’s lab called Circle Damage Sequencing, which allows scientists to “break” DNA at each point where damage occurs. They then coax the DNA into circles, which are replicated thousands of times using PCR. Once they have enough DNA, they use next-generation sequencing to identify which DNA bases are present at the breaks. Going forward, Pfeifer and colleagues plan to use this powerful technique to investigate other types of DNA damage in different kinds of cancer.