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Plastids harbor a multi-subunit RNA polymerase (RNAP) highly similar to eubacterial RNA polymerase, encoded by the plastid rpoA, rpoB and rpoC genes. The rpo genes have been found in all plant species so far investigated, with the exception of the parasite Epifagus virginiana. The eubacterial-like plastid RNA polymerase has been designated PEP (= plastid encoded polymerase). Several lines of evidence implicate the existence of a second RNA polymerase in the plastids of higher plants which is encoded in the nucleus (NEP = nuclear encoded polymerase. The plastid enzyme, NEP, functions in a coordinated manner, which can be described as „division of labour“, together with the plastid encoded PEP [4, 10, 14, 17].

A unique protein phosphorylation system regulated by light is present in the chloroplast thylakoid membrane (1,2). The light-induced phosphorylation occurs at the outer membrane surface on Thr residues of chlorophyll a/b-binding proteins and several subunits of photosystem II including the reaction centre proteins D1 and D2. This reversible phosphorylation regulates the balance of excitation energy between the two photosystems as well as the degradation and turnover of the central functional proteins of photosystem 11 (3). Very little is known about the enzymology of this reversible protein phosphorylation, in particular with respect to the control of thylakoid protein phosphatase activity. The enzymes involved is highly specific towards thylakoid phosphoproteins or mimicking phosphopeptides (4) and was proposed to be unrelated to conventional Ser/Thr phosphatases because of the failure to obtain inhibition by okadaic acid or microcystin in membrano experiments (5,6). In this work we demonstrate that upon purification, the intrinsic thylakoid membrane protein phosphatase reveals the properties characteristic of conventional eukaryotic protein phosphatases and that it is associated with and regulated by a recently discovered cyclophilin-like thylakoid lumenal prolyl isomerase of 40 kDa (TLP40) (7).

The main emphasis in photosynthesis research has so far been devoted to structural and functional aspects of electron/proton transport and light-harvesting. The progress has been significant, and the mechanism of photosynthetic energy conversion and the participating protein complexes can now be described at a very refined molecular level. In contrast, very little is known about the regulatory processes of photosynthesis, which maintain an optimal photosynthetic efficiency during fluctuating and stressful environmental conditions to the plant. The characterization of such regulatory or auxiliary processes, including the responsible enzymology and signal transduction mechanisms will be crucial topics for future photosynthesis research. Examples of such auxilary components or enzymes; are processing enzymes and chaperones involved in biogenesis of photosynthetic complexes [1], enzymes responsible for protein acylation [2] and isoprenylation [3], desaturases controlling membrane lipid composition and dynamics [4], stress induced proteins such as the ELIPs [5], as well as several enzymatic activities involved in various forms of regulatory proteolysis [6,7]. Protein phosphorylation is of importance for regulation of gene-expression and metabolic control and there is an increasing interest in thylakoid kinases and phosphatases [8].

As „open systems” organisms do not only add compounds (influx of matter), they also constantly exchange matter with their environment to modify structures and functions for optimization, maintenance, acclimatization, stress protection, and repair. Molecular mechanisms controlling this efflux of matter must contribute with comparable quality to the regulation of supramolecular structures as those generating them. Often such processes involve different regulatory components, such as kinases, phosphatases, or proteases, operating at various time scales that range from seconds to days.

The distribution of light energy between the two photosystems as well as the light-induced turnover of PSII proteins are regulated by the reversible phosphorylation of LHCII and the PSII-core proteins. The thylakoid protein kinase(s) is activated by a signal transduction system involving the interaction of reduced plastoquinone with the quinol oxidation site of the cytochrome bf complex [1]. Phosphorylation of the mobile pool of LHCII induces dissociation of this antenna from PSII and allows its interaction with the PSI in the stroma exposed membranes (state transition)[21. Dephosphorylation of LHCII by a membrane -bound phosphatase appears to be regulated by a cyclophilinlike protein located in the thylakoid lumen [1, 31. So far, it was assumed that the role of light in the above processes is only to drive the photosynthetic electron flow thus reducing plastoquinone and triggering the signal transduction system resulting in the protein kinase(s) activation. In this work we have used an in vitro reconstituted system consisting of partially purified, solubilized thylakoid protein-kinase(s) and isolated native as well as recombinant LHCII. We demonstrate that light induces conformational changes in the LHCII structure exposing the specific threonine phosphorylation site to the kinase activity, thus suggesting a regulation of protein phosphorylation also at the substrate level.

Reversible protein phosphorylation on serine and threonine residues plays a crucial role for the regulation of numerous cellular functions and signal transduction pathways. Control of this process is achieved by the modulation of the activities and substrate specificities of corresponding protein kinases and protein phosphatases. In thylakoids, a substantial number of proteins can be reversibly phosphorylated [I], and an intrinsic, redox-controlled kinase/phosphatase system is involved to adjust the energy distribution between the photosystems I and II [2]. Transient changes in light intensity and quality lead frequently to an imbalance of the excitation rates of the two photosystems which can result in a reduced photosynthetic efficiency. Plants harboring chlorophyll a/b antenna systems respond to this imbalance by modulating the association and energy transfer between the major light-harvesting antenna (LHCII) and the two photosystems via phosphorylation/dephosphorylation of LHCPs. This „state transition“ process is redox-controlled and activated by changes in the redox state of the thylakoid plastoquinone pool [3]. The exact mechanism of the phosphorylation-related reversible energy distribution of the LHCII antenna between the photosystems is not yet well understood. The signal transduction system appears to include a redox-controlled protein kinase of approx. 65 kDa molecular mass, the cytochrome b6f complex, induced conformational substrate (LHCII) changes (Zer et al., these Proceedings), and an immunophilin of the complex type (TLP40) located in the thylakoid lumen that appears to interact with the protein dephosphorylation signal transduction chain and to link it with protein degradation [4]. Attempts to isolate thylakoid-located kinase(s) and their cDNAs or genes have not been successful so far.

Two subunits of the fight-harvesting complex II (LHCII) and four of the photosystem II (PSII): the D1 and D2 core proteins, CP43 and a 9 kDa polypeptide constitute the principal phosphoproteins of the thylakoid membrane. Protein phosphorylation ensures the energy balance between the two photosystems [1] and regulates protein turnover during the repair of photoinhibitory damage to PSII [2,3]. Both thylakoid-bound [4–6] and soluble [7] protein phosphatases are present in chloroplasts, but only little is known about their specificity, location and regulation. In the present study we show that elevated temperature stimulates the thylakoid-bound protein phosphatase which dephosphorylates the reaction centre proteins D1 and D2. This activation is paralleled by a dissociation of the thylakoid cyclophilin TLP40 (thylakoid lumen PPIase of 40 kDa) [8, see also Andersson et al.; Vener et al. these Proceedings] from the inner thylakoid face, a possible regulator of thylakoid dephosphorylation.

Thylakoid membranes are specialized and highly versatile biomembranes, capable of converting solar energy to utilizable high-energy chemical metabolites. The invention of biological photosynthesis, which occurs in various forms in pro- and eukaryotes, ensured that life could survive; photosynthesis also provides the only renewable energy source of our planet. Thylakoid membranes are lipid bilayers into which several supramolecular protein complexes are intercalated in a vectorially oriented way. In oxygenic photosynthesis, they cooperate in the conversion of radiant energy together with mobile electron carriers in the stroma (or cytoplasm), lumen, and in the lipid bilayer.

The light harvesting apparatus of higher plant photosystem II (PSII) is a highly complex and heterogenous structure consisting of several chlorophyll-binding proteins including the minor chlorophyll a/b complexes, CP29, CP27 and CP24. CP24 was first identified by Dunahay and Staehlin (1) and is estimated to contain only about 5% of the PSII chlorophyll. It is proposed to contain one (2) or several polypeptides in the range of 20–24 kDa (1). CP24 has been suggested to act as a linker protein between the light-harvesting antenna and the core of PSII (1). Others have suggested it to be associated with the light-harvesting apparatus of PSII (3,4).