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The involvement of HSP70B in the photosystem II damage and repair process in Dunaliella salina was investigated. A full-length cDNA of the D. salina hsp70B gene was cloned and sequenced. Expression patterns of the hsp70B gene were investigated upon shifting a D. salina culture from low-light to high-light growth conditions, designed to significantly accelerate the rate of PSII photodamage. Northern blot analyses and nuclear run-on transcription assays revealed a significant but transient induction of hsp70B gene transcription, followed by a subsequent increase in HSP70B protein synthesis and accumulation. Mild detergent solubilization of photoinhibited thylakoid membranes, in which photodamaged PSII centers had accumulated, followed by native gel electrophoresis revealed the formation of a 320 kDa protein complex that contained, in addition to the HSP70B, the photodamaged but as yet undegraded D1 protein as well as D2 and CP47. Evidence suggested that the 320 kDa complex is a transiently forming PSII repair intermediate. Denaturing solubilization of the 320 kDa PSII repair intermediate by SDS-urea resulted in cross-linking of its polypeptide constituents, yielding a 160 kDa protein complex. The role of the HSP70B in the repair of photodamaged PSII centers, e.g. in stabilizing the disassembled PSII-core complex and in facilitating the D1 degradation and replacement process, is discussed.

In addition to glutathione (y-GluCysGly), many species of the family Poaceae have another tripeptide which has the amino acid sequence y-GluCysSer. This thiol was isolated from etiolated leaves of wheat (Triticum aestivum L. cv. Star). Its structure was elucidated by quantitative amino acid analysis after total hydrolysis and by partial hydrolysis with carboxypeptidase A and y-glutamyltranspeptidase. The content of y-GluCysSer in the leaves ofT. aestivum is increased by incubation with sulfate and is severely diminished by incubation with buthionine sulfoximine, a specific inhibitor of y-glutamylcysteine synthetase. Oxidized y-GluCysSer is reduced by yeast glutathione reductase with a rate somewhat lower than for glutathione, but the new tripeptide is not a substrate of glutathione-S-transferase from equine liver.

The putative alpha-galactosidase gene HvSF11 of barley, previously shown to be expressed during dark induced senescence, is expressed in the growing/elongating zone of primary foliage leaves of barley. The amino acid sequence deduced from the full length HvSF11 cDNA contains a hydrophobic signal sequence at the N-terminus. Phylogenetic relationship of the HvSF11 encoded barley alpha-galactosidase to other alpha-galactosidases revealed high homology with the alpha-galactosidase encoded by the gene At5g08370 from Arabidopsis thaliana. We have isolated two independent heterozygous At5g08370 T-DNA insertion mutants from Arabidopsis thaliana, both of which have a higher number of rosette leaves with a curly surface leaf morphology and delayed flowering time in comparison to wildtype plants. Localization of the Arabidopsis alpha-galactosidase protein via GUS-tag revealed that the protein is associated with the cell wall. This result was confirmed by immunological detection of the orthologous barley protein in a protein fraction derived from cell walls of barley leaves. It is concluded that the alpha-galactosidase proteins from barley and Arabidopsis might fulfill an important role in leaf development by functioning in cell wall loosening and cell wall expansion.

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