Cu/Zn superoxide dismutases in developing cotton fibers

Authors: KIM, HEE - UNIV OF NEW ORLEANS; KATO, NAOHIRO - LA STATE UNIV; Kim, Sunran; Triplett, Barbara

Submitted to: Planta
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/25/2008
Publication Date: 6/15/2008
Citation: Kim, H.J., Kato, N., Kim, S., Triplett, B.A. 2008. Cu/Zn superoxide dismutases in developing cotton fibers. Planta. 228:281-292.

Interpretive Summary: Growth of all plant cells, including cotton fiber, is dependent on dual capacities of the plant cell wall; the wall must have enough structural integrity to withstand high pressures generated within each cell and, at the same time, be elastic enough to allow incorporation of new materials into the expanding structure. Recent advances in the biochemistry of plant cell walls suggest the involvement of a chemical, hydrogen peroxide, in both cell wall expansion and cell wall rigidity, therefore, generation of this chemical within the cell wall must be precisely regulated. There are several processes that can form hydrogen peroxide in plants, including a reaction catalyzed by the enzyme superoxide dismutase. In this study, we have discovered a distinct form of superoxide dismutase enzyme that functions in the cell wall of cotton fibers and compare this form with other superoxide dismutase forms that exist in the fiber’s cytoplasm and in chloroplasts, subcellular structures that function in photosynthesis. Using three different methods of localizing superoxide dismutase enzymes coded by the cotton genome, we show that only one type is localized to the cotton fiber cell wall. Additionally, this manuscript identifies potential pitfalls in using a commonly used procedure, antibody localization using transmission electron microscopy, for localizing plant cell wall proteins. Our team is now directly testing the function of cell wall forms of superoxide dismutase in model plants that have had the gene function “knocked out”. If a direct association can be made between superoxide dismutase activity in plant cell walls and cell wall physical properties, the cell wall form of superoxide dismutase will become a candidate gene for genetic manipulation in cotton, biomass crops, and wood-forming species.

Technical Abstract: Hydrogen peroxide (H2O2) and other reactive oxygen species (ROS) are important signaling molecules in diverse physiological processes. Previously, we discovered superoxide dismutase (SOD) activity in extracellular protein preparations from fiber-bearing cotton (Gossypium hirsutum L.) seeds. We show here by use of an inhibitor and immunoreactivity that the enzyme is a CuZn-SOD (CSD). CSD enzyme activity in the apoplast was present throughout all stages of cottonseed development, reaching a peak at 25 days post-anthesis. Five cotton CSD cDNAs were cloned from cotton fiber and classified into three subfamilies (Group 1: GhCSD1; Group 2: GhCSD2a and GhCSD2b; Group 3: GhCSD3 and GhCSD3s). Members of group 1 and 2 are expressed throughout fiber development, but predominant during the cell elongation stage. Group 3 CSDs are also expressed throughout fiber development, but transiently increase in abundance at the transition period between cell elongation and secondary cell wall synthesis. Each of the three GhCSDs also has distinct patterns of expression in tissues other than fiber. Immunogold localization shows that GhCSD3 localizes to secondary cell walls; however, easily soluble forms may be lost during fixation. Overexpression of cotton CSDs fused to green fluorescent protein (GFP) in transgenic arabidopsis demonstrated that GhCSD1 is localized to the cytosol, GhCSD2a localizes to plastids, and GhCSD3 is translocated to the apoplast. Subcellular fractionation of proteins from transgenic arabidopsis seedlings confirmed that c-myc epitope-tagged GhCSD3 co-purifies with cell wall proteins. Apoplastic CSDs and other enzymes that regulate the ROS content of plant cell walls are potential regulators of plant cell growth and development.