**5. Conclusions and perspectives**

The heme and chlorophyll biosynthetic pathways branch at protoporphyrin IX (Prot IX). GTR, glutamine tRNA reductase, is subjected to feedback inhibition by heme and FLU. In most land plants, conversion of PChlide (protochlorophyllide) to Chlide is light-dependent (L, in *Chlamydomonas* this conversion also occurs in the dark, *D*). Through its negative feedback on GTR, FLU prevents overaccumulation of PChlide in the dark. The steps affected by the *gun* and *hy* mutations which affect retrograde signaling are indicated. GSA, glutamate 1-semial‐ dehyde; ALA, 5-aminolevulinic acid; Chl, chlorophyll; Fc, ferrochelatase, Hmox1, heme oxygenase; BV, biliverdin; Pcy, bilin reductase; PCB, phytocyanobilin, CHLH, CHLI and

Additional evidence for the involvement of tetrapyrroles in retrograde signaling comes from the identification of a functional bilin biosynthesis pathway in *Chlamydomonas*[130,131]. In this pathway, protoporphyrin IX is converted to protoheme and Mg-Proto by Fe- and Mgchelatase, respectively. While heme is used as prosthetic group for many hemoproteins, a portion of heme is converted to biliverdin IXa by heme oxygenase (Hmox1) and in the next step by a ferredoxin-dependent phytochromobilin synthase (PcyA) to phytochromobilin which serves as chromophore of phytochromes (**Figure 5**). However, *Chlamydomonas* as well as other chlorophytes do not produce phytochromes, raising questions on the role of this pathway in these algae. Some clues came from the analysis of a mutant of *Chlamydomonas* deficient in Hmox1 whose phototrophic growth is compromised and in which the increase of chlorophyll upon a dark-to-light transition no longer occurs [131]. Comparative transcriptomic studies of wild-type and *hmox1* cells revealed a set of nuclear genes that are up-regulated by bilins and that comprise several oxygen-dependent redox enzymes such as mono-and dioxygenases, proteins with redox cofactors and enzymes of oxidative amino acid metabolism. These results raise the possibility that bilins operate within a retrograde signaling pathway that evolved in chlorophytes for the detoxification of ROS generated during sudden dark to light shifts. It remains to be seen whether bilins assume additional roles in chlorophytes besides ensuring smooth daily transitions from dark to light with minimal photooxidative damage.

A further striking example of the action of tetrapyrroles as mediators for plastid-to-nucleuscommunication is the identification of a tetrapyrrole-regulated ubiquitin ligase for cell cycle coordination from organelle to nuclear DNA replication in the red alga *Cyanidioschyzon*

Redox changes within the photosynthetic electron transport chain occur upon changes in light quality and quantity, CO2 levels, nutrient availability and elevated temperature. As a result of unequal excitation of PSI and PSII or of insufficient electron acceptor capacity on the PSI acceptor side, the redox state of the plastoquinone pool is altered. In this case, chloroplast gene expression is affected in land plants [135] although the evidence is less convincing in algae. However, in these organisms, there is unambiguous evidence that nuclear gene expression is affected [136]. A possible candidate for sensing the redox state of the plastoquinone pool is the chloroplast protein kinase Stt7/STN7 which is known to be activated when plastoquinol occupies the Qo site of the Cyt*b*6*f* complex [21,22]. During experiments in which plants were shifted from light preferentially absorbed by PSI to light preferentially absorbed by PSII, the

*merolae* [132–134].

38 Applied Photosynthesis - New Progress

CHLD are subunits of Mg-chelatase. Reproduced from Ref. 126 with permission.

The photosynthetic apparatus is a complex machinery consisting of several large protein– pigment complexes whose components are encoded by both nuclear and chloroplast genes. Thus, the biosynthesis of this system involves two distinct genetic systems which act in a coordinate manner. In nature, photosynthetic organisms are subjected to continuous environ‐ mental changes and need to adapt so as to maintain optimal photosynthetic activity and to protect themselves from photooxidative damage. These processes can be grouped in shortterm and long-term responses. The first occurs in the second-to-minute range and involves light-induced protein conformational changes, posttranslational protein modifications, cell compartment–specific pH changes and ion fluxes across the chloroplast and thylakoid membranes. The second occurs in the minute-to-hour range and involves changes in gene expression and protein accumulation, which depend on an intricate bilateral communication system between chloroplasts and nucleus. Many nuclear genes encoding chloroplast proteins have been identified which are required for chloroplast gene expression and act mainly at posttranscriptional steps. Some of these factors appear to act constitutively while others assume a regulatory role because they have short half-lives and their level varies greatly upon changes in environmental cues including light, temperature and nutrient availability. How‐ ever, the molecular mechanisms underlying the intercompartmental communication between chloroplast, mitochondria and nucleus are still largely unknown although several retrograde signals have been identified. They involve specific compounds such as tetrapyrroles and isoprenoids as well as plastid protein synthesis, the redox state of the photosynthetic electron transport chain and ROS generated under specific stress conditions. Moreover, a complex signaling network is operating within chloroplasts comprising several protein kinases and phosphatases, ion channels, and specific metabolites which act as signals and for the commu‐ nication between chloroplast and nucleus. However, the signaling chains connecting these different components are still largely unknown and their identification remains an important challenge for future research.

The flexibility of the thylakoid membrane is truly remarkable. Although it is crowded with proteins, it still allows for efficient remodeling of the photosynthetic complexes within the thylakoid membrane especially in response to changes in the quality and quantity of light. Among these responses, state transitions and NPQ have been studied extensively and some of the underlying molecular mechanisms have been elucidated. However, many questions remain open. We still do not fully understand how the Stt7/STN7 kinase that plays a central role in state transitions and chloroplast signaling is activated and inactivated as a result of perturbations of the chloroplast redox poise. From an evolutionary point of view, it is partic‐ ularly interesting to compare these adaptive responses in different photosynthetic organisms such as plants, fresh water and marine algae and cyanobacteria. In this respect, NPQ, the dissipation of excess excitation energy as heat in the light-harvesting systems of the photo‐ systems, is of great importance and it is widely used in the plant kingdom. Recent studies on NPQ in different photosynthetic organisms raise several questions regarding the evolution of this essential photoprotective mechanism. For example, it is not clear why the Lhcsr proteins were lost during the transition from aquatic to land plants. Moreover, the qE process in most algae derived by secondary endosymbiosis from a red algal ancestor differs from that in extant red algae. All of these derived algae possess a xanthophyll cycle and Lhcsr-related proteins which are apparently absent in red algae [87] and which have been suggested to be derived from green algae [143,144]. It will clearly be important and challenging to elucidate these evolutionary puzzles.
