Exposure to solar radiation can cause mortality in natural communities of pico-phytoplankton, both at the surface and to a depth of at least 30 m. DNA damage is a significant cause of death, mainly due to cyclobutane pyrimidine dimer formation, which can be lethal if not repaired. While developing a UV mutagenesis protocol for the marine cyanobacterium Prochlorococcus, we isolated a UV-hyper-resistant variant of high light-adapted strain MED4. The hyper-resistant strain was constitutively upregulated for expression of the mutT-phrB operon, encoding nudix hydrolase and photolyase, both of which are involved in repair of DNA damage that can be caused by UV light. Photolyase (PhrB) breaks pyrimidine dimers typically caused by UV exposure, using energy from visible light in the process known as photoreactivation. Nudix hydrolase (MutT) hydrolyses 8-oxo-dGTP, an aberrant form of GTP that results from oxidizing conditions, including UV radiation, thus impeding mispairing and mutagenesis by preventing incorporation of the aberrant form into DNA. These processes are error-free, in contrast to error-prone SOS dark repair systems that are widespread in bacteria. The UV-hyper-resistant strain contained only a single mutation: a 1 bp deletion in the intergenic region directly upstream of the mutT-phrB operon. Two subsequent enrichments for MED4 UV-hyper-resistant strains from MED4 wild-type cultures gave rise to strains containing this same 1 bp deletion, affirming its connection to the hyper-resistant phenotype. These results have implications for Prochlorococcus DNA repair mechanisms, genome stability and possibly lysogeny.

Summary 23The marine cyanobacterium Prochlorococcus, the smallest and most abundant 24 oxygenic phototroph, has an extremely streamlined genome and a high rate of protein 25 evolution. High-light adapted strains of Prochlorococcus in particular have seemingly 26 inadequate DNA repair systems, raising the possibility that inadequate repair may lead 27 to high mutation rates. Prochlorococcus mutation rates have been difficult to determine, 28 in part because traditional methods involving quantifying colonies on solid selective 29 media are not straightforward for this organism. Here we used a liquid dilution method 30 to measure the approximate number of antibiotic-resistant mutants in liquid cultures of 31Prochlorococcus strains previously unexposed to antibiotic selection. Several antibiotics 32 for which resistance in other bacteria is known to result from a single base pair change 33 were used. The resulting frequencies of antibiotic resistance in Prochlorococcus 34 cultures allowed us to then estimate maximum spontaneous mutation rates, which were 35 similar to those in organisms such as E. coli (~5.4x10 -7 per gene per generation). 36 Therefore, despite the lack of some DNA repair genes, it appears unlikely that the 37 Prochlorcoccus genomes studied here are currently being shaped by unusually high 38 mutation rates.