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Light induces rapid changes of the phosphorylation pattern in the cytosol of evacuolated parsley protoplasts

Harter, K., Frohnmeyer, H., Kircher, S., Kunkel, T., Muhlbauer, S., Schafer, E.
Proceedings of the National Academy of Sciences of the United States of America 1994 v.91 no.11 pp. 5038-5042
Petroselinum crispum, plant proteins, phosphorylation, cytosol, protoplasts, photoreceptors, red light, white light, ultraviolet radiation, phytochrome, phosphoprotein phosphatase
The fractionation of cells of a parsley suspension culture [Petroselinum crispum (Mill.) A. Hill] by protoplasting subsequent removal of the vacuoles led to physiologically intact evacuolated protoplasts retaining light inducibility of chalcone synthase expression. Lysis of the evacuolated protoplasts permitted the isolation of a pure, highly concentrated cytosolic fraction containing major cytosolic membranes but only minor contamination by proplastids, mitochondria, and nuclei. Short-time irradiations of the cytosol with red or UV-containing white light resulted in very fast changes of the phosphorylation pattern of 18-, 40-, 48-, 55- to 70-, and 120-kDa proteins. Major differences were observed between the phosphorylation patterns obtained by red or UV-containing white light treatment, indicating a different primary action of the excited photoreceptors in vitro. Separation of the microsomal fraction from the cytosolic matrix established the localization of these proteins. Chase and photoreversibility experiments revealed that phytochrome in vitro regulates the phosphorylation of the 40-kDa protein by modifying a soluble cytosolic kinase/phosphatase system.