Translesion synthesis (TLS) is one of the DNA damage tolerance strategies, which have evolved to enable organisms to replicate their genome despite the presence of unrepaired damage. The process of TLS has the propensity to produce mutations, a potential origin of cancer, and is therefore of medical interest. Significant progress in our understanding of TLS has come primarily from studies of the bacterium Escherichia coli, the budding yeast Saccharomyces cerevisiae and, more recently, human cells. Results from these analyses indicate that the fundamental mechanism of TLS and the proteins involved have been conserved throughout evolution from bacteria to humans.