Photolyase expression in sea urchin larvae using in situ hybridization

Abstract

Stratospheric ozone depletion (the ozone hole) over the Antarctic, and the resulting increase of ultraviolet-B radiation (UV-B, 280 to 320 nm), poses a major threat to Antarctic marine ecosystems (Wulff, 2008). The ozone hole is now predicted to persist for the next 50 years at least, and with the threat of global climate change extending its life, its direct effects in marine ecosystems and the organisms within are paramount. Planktonic organisms, including the larval life-history stages of many benthic marine invertebrates, are exposed to harmful UV-B while in the upper water column. UV-B causes lethal and sub-lethal damage to DNA, lipids and proteins, reduces photosynthesis and fecundity. UV-B increases 20-fold through the sea ice (Lesser et al., 2004) while the ozone hole is present and penetrates deeper into the water column (Smith, 1992), while near-UV and blue light, associated with photo-reactivation and photo-repair do not proportionally increase, tipping the balance in favour of damage. Cyclobutane pyrimidine dimers (CPDs) are the most common form of DNA damage induced by UV-B, and can be repaired through the production of photolyases that reversibly bind and convert CPDs back to the original bases. Photolyase are catalytic, using near-UV and blue light (320-700 nm) to repair damaged DNA. Photolyase activity has already been shown to increase under ozone depleted conditions (Lamare, et al., 2006; Isely, et al., 2009). This thesis applies in situ hybridisation techniques to examine expression of the DNA repair enzyme photolyase in sea urchin embryos in response to UV-R exposure. From this I was able to see where, and to what extent, damage was occurring in relation to (1) depth (2) sea ice coverage and (3) with overhead ozone conditions in McMurdo Sound during the spring of 2008. I observed an increase in photolyase expression and abnormality with increased exposure to UV-B (either by ozone depletion or open water). Most notably, photolyase expression was significantly greater during periods when the ozone hole was positioned over the study site. The endemic New Zealand sea urchin Evechinus chloroticus was also exposed in situ during the summer of 2009 and the expression of photolyase examined along with levels of CPDs. E. chloroticus larvae expressed significantly higher levels of photolyase when exposed to UV-B radiation at depths less than 2 m compared to larvae protected from UV-B.