Whole-tissue determination of the rate coefficients of photoinactivation and repair of photosystem II in cotton leaf discs based on flash-induced P700 redox kinetics

Using radioactively labelled amino acids to investigate repair of photoinactivated photosystem II (PS II) gives only a relative rate of repair, while using chlorophyll fluorescence parameters yields a repair rate coefficient for an undefined, variable location within the leaf tissue. Here, we report on a whole-tissue determination of the rate coefficient of photoinactivation k _i, and that of repair k _r in cotton leaf discs. The method assays functional PS II via a P700 kinetics area associated with PS I, as induced by a single-turnover, saturating flash superimposed on continuous background far-red light. The P700 kinetics area, directly proportional to the oxygen yield per single-turnover, saturating flash, was used to obtain both k _i and k _r . The value of k _i, directly proportional to irradiance, was slightly higher when CO₂ diffusion into the abaxial surface (richer in stomata) was blocked by contact with water. The value of k _r, sizable in darkness, changed in the light depending on which surface was blocked by contact with water. When the abaxial surface was blocked, k _r first peaked at moderate irradiance and then decreased at high irradiance. When the adaxial surface was blocked, k _r first increased at low irradiance, then plateaued, before increasing markedly at high irradiance. At the highest irradiance, k _r differed by an order of magnitude between the two orientations, attributable to different extents of oxidative stress affecting repair (Nishiyama et al., EMBO J 20: 5587-5594, 2001). The method is a whole-tissue, convenient determination of the rate coefficient of photoinactivation k _i and that of repair k _r .