Complexity of DNA Repair From Alcohol Damage Revealed

Hematologists have long known of alcohol’s negative effects on blood cells and hematopoiesis — effects that were thought to be reversible.^1,2  However, is the latter strictly true? While alcohol is not typically listed as an environmental agent causing hematologic malignancies, it is metabolized to acetaldehyde, a carcinogen that damages DNA, and chronic alcohol use is certainly associated with solid tumors, including breast, liver, colon, and esophageal.³  Indeed, an insidious action of alcohol on the bone marrow was revealed when Japanese patients with Fanconi anemia (FA) and elevated acetaldehyde from aldehyde dehydrogenase 2 (ALDH2) deficiency were found to have an accelerated progression to bone marrow failure.⁴  However, the interplay between alcohol, acetaldehyde, DNA damage, and its subsequent repair in hematopoietic stem cells (HSCs) has not been well understood until now.

In a recent Nature article, Dr. Juan I. Garaycoechea and colleagues found that acetaldehyde damages the DNA of mouse bone marrow HSCs, and its correction requires multiple different mechanisms of DNA repair. A wealth of prior research has taught us that the FA repair pathway protects against cumulative DNA cross-link damage. By studying patients with this rare disease and through experiments on the different FA genes, we know that this pathway is complex and involves very different DNA repair mechanisms including homologous recombination, nucleotide excision repair, and translesion synthesis. The authors previously looked at mice with a strong FA phenotype.⁵  The mice lack FANCD2, an important FA protein involved in the recruitment of downstream DNA repair. When these mice were also deficient in aldehyde dehydrogenase 2 (Aldh2^-/-Fancd2^-/-), the HSCs were more susceptible to acetaldehyde.

Now in an elegant series of sequential genetic deletion experiments, the authors have defined which of the DNA repair mechanisms are needed to protect from acetaldehyde-induced DNA damage. First, they showed that Aldh2-/-Fancd2-/- mouse bone marrow cells had chromosomes with increased reciprocal transfer of genetic material between sister chromatids. This phenomenon, known as sister chromatid exchange, is often used as a surrogate marker for homologous recombination events. The number of sister chromatid exchanges is potentiated on exposure to alcohol and indicated that acetaldehyde induces homologous recombination DNA repair. They then confirmed this by showing that DT40 cells lacking both homologous recombination and FA genes were more sensitive to acetaldehyde than cells deficient in only one.

Next, they used micronuclei in enucleated erythrocytes and multiplex FISH as markers for DNA damage. In Aldh2^-/-Fancd2^-/- mice, there is a basal increase in DNA damage, and this was exacerbated fourfold by exposure to ethanol. This shows that in the presence of normally functioning homologous recombination repair machinery, the FA pathway is also required to repair chromosome breakage from acetaldehyde. They then looked at mice lacking the FA repair gene Fanca and the nonhomologous end-joining repair (NHEJ) protein Ku70. These mice were anemic, had fewer HSCs, had more DNA damage, and were more sensitive to acetaldehyde. This suggested that NHEJ repair is also important for repair of acetaldehyde-induced DNA damage, at least in the absence of the FA pathway.

Finally, serial transplantation experiments with either one or five HSCs showed that Aldh2^-/-Fancd2^-/- mice have HSCs that are less able to engraft and contribute to hematopoiesis in recipient mice. By sequencing the recipient mouse bone marrows, they showed that acetaldehyde causes specific structural DNA damage involving insertions, deletions, translocations,and rearrangements. Aldh2^-/-Fancd2^-/- mice also had hematopoietic and progenitor stem cells with increased expression of p53 and the poor bone marrow engraftment improved on a p53 null background.