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ENVIRONMENTAL STRESS AND ADAPTATION TO STRESS

Localization and Role of Manganese Superoxide Dismutase in a Marine Diatom^1,[OA]

Environmental Biophysics and Molecular Ecology Program, Institute of Marine and Coastal Sciences (F.W.-S., O.S., P.G.F.), and Department of Environmental Science (J.R.R.), Rutgers University, New Brunswick, New Jersey 08901; and Department of Cell Biology and Neuroscience (V.S.), and Department of Geological Science, Wright Geological Laboratory (P.G.F.), Rutgers University, Piscataway, New Jersey 08854

Superoxide dismutase (SOD) catalyzes the transformation of superoxide to molecular oxygen and hydrogen peroxide. Of the four known SOD isoforms, distinguished by their metal cofactor (iron, manganese [Mn], copper/zinc, nickel), MnSOD is the dominant form in the diatom Thalassiosira pseudonana. We cloned the MnSOD gene, sodA, using the expression vector pBAD, overexpressed the product in Escherichia coli, and purified the mature protein (TpMnSOD). This recombinant enzyme was used to generate a polyclonal antibody in rabbit that recognizes MnSOD in T. pseudonana. Based on quantitative immunoblots, we calculate that in vivo concentrations of TpMnSOD are approximately 0.9 amol cell^–1 using the recombinant protein as a standard. Immunogold staining indicates that TpMnSOD is localized in the chloroplasts, which is in contrast to most other eukaryotic algae (including chlorophytes and embryophytes) where MnSOD is localized exclusively in mitochondria. Based on the photosynthetic Mn complex in photosystem II, cellular Mn budgets cannot account for 50% to 80% of measured Mn within diatom cells. Our results reveal that chloroplastic MnSOD accounts for 10% to 20% of cellular Mn, depending on incident light intensity and cellular growth rate. Indeed, our analysis indicates that TpMnSOD accounts for 1.4% (±0.2%) of the total protein in the cell. The TpMnSOD has a rapid turnover rate with an apparent half-life of 6 to 8 h when grown under continuous light. TpMnSOD concentrations increase relative to chlorophyll, with an increase in incident light intensity to minimize photosynthetic oxidative stress. The employment of a Mn-based SOD, linked to photosynthetic stress in T. pseudonana, may contribute to the continued success of diatoms in the low iron regions of the modern ocean.

² Present address: Metallomics Laboratory, Department of Chemistry and Biochemistry, Arizona State University, Tempe, AZ 85287.

The author responsible for distribution of materials integral to the findings presented in this article in accordance with the policy described in the Instructions for Authors (www.plantphysiol.org) is: Paul G. Falkowski (falko{at}imcs.rutgers.edu).

^[OA] Open Access articles can be viewed online without a subscription.

www.plantphysiol.org/cgi/doi/10.1104/pp.106.088963

^* Corresponding author; e-mail fwolfe{at}asu.edu; fax 480–965–2747.

Received September 1, 2006; accepted October 16, 2006; published October 20, 2006.

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