Evidence for loss of D1 protein during photoinhibition of Chenopodium rubrum L. culture cells

The effects of high-light stress on chlorophyllfluorescence parameters, D1-protein turnover and the actual level of this protein were analysed in nitrogen-deficient and nitrogen-replete cells of Chenopodium rubrum L. Changes in the number of atrazine-binding sites and in the D1-protein immunoblot signal indicated that a net loss of D1 protein occurred in high light and was partly reversible in low light. Nitrogen deficiency did not exacerbate these changes. The involvement of D1-protein turnover was shown in pulse-chase experiments with [³⁵S]-methionine and by the application of a chloroplastic protein-synthesis inhibitor (chloramphenicol). The slowly reversible non-photochemical fluorescence quenching increased pronouncedly when D1 protein was lost at high irradiances, but its increase was only small when a net loss of D1 protein was produced at moderate irradiances by addition of chloramphenicol. The ratio of variable to maximum fluorescence, F_v/F_m, and the number of atrazine-binding sites were correlated but a proportionality between these parameters could not be observed. We conclude from these results that (i) degradation of D1 protein was not always coupled to its resynthesis, (ii) the actual level of D1 protein reflected the balance between degradation and resynthesis of D1 protein and (iii) changes in the level of D1 protein did not depend on a pronounced increase of the slowly reversible non-photochemical quenching. We thank Helga Simper for excellent technical assistance and Stefan Peter for his enthusiastic help in some of the experiments. Furthermore, we thank Professor E. Beck (Lehrstuhl für Pflanzenphysiologie, Bayreuth, FRG) for critical reading of the manuscript, Professor M. Sprinzl (Lehrstuhl für Biochemie, Bayreuth, FRG) for the opportunity to use the laser densitometer and Professor A. Trebst (Lehrstuhl für Biochemie der Pflanzen, Bochum, FRG) for the generous gift of D1 antibody. This work was supported by the Deutsche Forschungsgemeinschaft (SFB 137).