The Evolution of Light Stress Proteins in Photosynthetic Organisms

The Elip (early light-inducible protein) family in pro- and eukaryotic photosynthetic organisms consists of more than 100 different stress proteins. These proteins accumulate in photosynthetic membranes in response to light stress and have photoprotective functions. At the amino acid level, members of the Elip family are closely related to light-harvesting chlorophyll a/b-binding (Cab) antenna proteins of photosystem I and II, present in higher plants and some algae. Based on their predicted secondary structure, members of the Elip family are divided into three groups: (a) one-helix Hlips (high light-induced proteins), also called Scps (small Cab-like proteins) or Ohps (one-helix proteins); (b) two-helix Seps (stress-enhanced proteins); and (c) three-helix Elips and related proteins. Despite having different physiological functions it is believed that eukaryotic three-helix Cab proteins evolved from the prokaryotic Hlips through a series of duplications and fusions. In this review we analyse the occurrence of Elip family members in various photosynthetic prokaryotic and eukaryotic organisms and discuss their evolutionary relationship with Cab proteins.


Introduction
Plants and cyanobacteria respond to light stress by transient accumulation of light stress proteins from the Elip (early light-induced proteins) family [1]. These proteins are integrally located in photosynthetic membranes, spanning the membrane with either one (e.g. Ohps, one-helix proteins, called also Hlips; high light-induced proteins or Scps; small Cab-like proteins in prokaryotic organisms), two (e.g. Seps, stress-enhanced proteins) or three (e.g. Elips) α-helices [1,6,10,15]. The members of the Elip family are closely related to chlorophyll a/b-binding (Cab) or fucoxanthin-chlorophyll a/cbinding proteins that form antenna systems around photosystems I (PS I) and II (PS II) in chlorophytes (green algae, mosses, ferns and higher plants) or chromophytes, respectively [4,19].
According to sequence conservation between Elip and Cab family members, all of these proteins are assumed to share a common evolutionary origin [7,19] and presumably have a similar structure. The three-dimensional structure of one Cab family member has been determined at 3.4Å resolution by electron crystallography of two-dimensional crystals [12] showing that two of the three transmembrane α-helices are held together by ion pairs formed by charged residues. Thus, it is expected that Elip family members will have a similar twofold symmetry structure, since helices I and III of Elips and Cab proteins and the helix I of Seps and Ohps/Hlips/Scps are highly conserved in their amino acid composition. However, in order to form such structures, two-helix Seps and onehelix Ohps/Hlips/Scps need to form homo-or heterodimers.
In addition to similarities between Elip and Cab proteins there are also very pronounced differences. In contrast to Cab family members that are the most abundant proteins of the thylakoid membranes, Elips accumulate only transiently in substoichiometric amounts in response to physiological stress [1]. Moreover, a very unusual pigment composition and pigment-binding characteristics were reported for isolated Elips, such as a weak excitonic coupling between chlorophyll a molecules and an extremely high lutein content as compared with other chlorophyll-binding proteins [2]. Based on these features, a non-lightharvesting function has been proposed for Elip family members. It is believed that these proteins fulfil a photoprotective role within thylakoid membranes under light stress conditions, either by transient binding of free chlorophyll molecules and preventing the formation of free radicals, and/or by participating in energy dissipation [1,16].
In this review we analyse the evolutionary relationship of various members of Elip family in proand eukaryota and discuss the origin of eukaryotic Cab proteins.

The distribution of Elip family members in photosynthetic organisms
To resolve the evolutionary relationship between one-two-and three-helix members from the Elip family and to investigate their relation with Cab proteins, we performed BLAST searches in various databases (http://www.tigr.org; http://megasun. bch.umontreal.ca/ogmp/projects/other/cp list. html; http://mips.gsf.de/proj/sputnik/oryza; http://www.jgi.doe.gov/JGI microbial/html) using the Elip consensus motif ERINGRLAMIGFVAA-LAVE, located in the first conserved transmembrane α-helix [1,10], or full-length sequences of different Elip family members. Table 1 shows the distribution of the Elip family members across cyanobacteria, photosynthetic protists and plants, their sizes and their predicted secondary structures.
Searches in the databases of cyanobacteria revealed that multigene Elip families composed of eight Hlip/Scp members are present in the genome of Nostoc sp. PCC7120 or Synechococcus sp. PCC7942. Prochlorococcus marinus contained 10 predicted Hlip/Scp genes (Table 1). In contrast, only one member of Hlips/Scps was found in the Glaucocystophyta Cyanophora paradoxa or in red algae Porphyra purpurea and Cyanidium caldarium. The Cryptophyta Guillardia theta contained two Hlip/Scp members. All Hlips/Scps found in red algae or in Cryptophyta are encoded by the plastid genome (Table 1). In Glaucocystophyta the Hlip/Scp gene is located on a cyanelle genome (VL Stirewalt, CB Michalowski, W Löffelhardt, HJ Bohnert and D.A. Bryant, 1995; direct GenBank submission). Cyanelles are plastid-like organelles that resemble cyanobacteria in morphology, in the organization of their photosynthetic apparatus and in the presence of the peptidoglycan wall [14]. The recent acquisition of complete plastid genome sequences of Glaucocystophyta, Rhodophyta and Cryptophyta allowed us to search for Elip family members in these organisms. However, one should be aware that more Hlips/Scps may be discovered in the nuclear genomes of these algae.
To investigate whether the type of the photosynthetic antenna correlates with the number and/or type of Elip family members present in these organisms, we compared the antenna systems of red algae, Cryptophyta and the cyanobacteria Nostoc and Synechococcus. Both algal groups and cyanobacteria have a chlorophyll a-containing antenna complex functionally associated with PS I and phycobilisomes, which serve as a lightharvesting antenna in PS II [9,19]. While in cyanobacteria chlorophylls are bound directly to the A and B subunits of the PS I reaction centre, in red algae and Cryptophyta proteins related to Cab family members participate in chlorophyll a-binding [19]. In Prochlorophyta the lightharvesting antenna consists of both chlorophylls a and b [17] bound to proteins fundamentally different from Cab proteins. These proteins are encoded by the IsiA (iron stress-induced) gene [13] and are related to the CP43 protein of PS II core complex in higher plants. Interestingly, all these organisms, independently of the antenna-type, contain only one helix Hlips/Scps (Table 1 and Figure 1A). No Seps or Elips were found in these algae or cyanobacteria, suggesting that two-or three-helix Elip family members arose more recently.
Three-helix Elip proteins appeared for the first time in the green algae Dunaliella bardawil and Chlamydomonas reinhardtii, which contained one or six Elips, respectively (Table 1, Figure 1A). In the moss Tortula ruralis and the fern Onoclea sensibilis, two or one Elips were found in the nuclear genomes. A dicotyledon, Arabidopsis thaliana, contained two three-helix Elips, five two-helix Seps and two Ohps. The genome of  Figure 1). All Elip family members in green algae, mosses, ferns and higher plants investigated so far are nuclear-encoded proteins posttranslationally imported into chloroplasts [1]. This indicates that during evolution Elip genes have undergone translocation from plastids to the nucleus and have further evolved into topologically different proteins. It is generally accepted that chloroplasts are derived from a single cyanobacterial ancestor. Of  Table 1 the 3000 genes present in the cyanobacterium Synechocystis PCC6803, only 100-200 are found on plastid genomes, indicating that a massive transfer of genes to the nucleus occurred following endosymbiosis and establishment of the plastid [3].
The antenna system in green algae and higher plants is composed of Cab proteins associated with PS I and PS II that are encoded by multigene families consisting of 21 different members, as was reported for A. thaliana [11]. The Cab family in higher plants also includes the PsbS protein with four transmembrane helices, which is located in PS II [5]. Searches in the databases revealed that one PsbS gene has been annotated in the A. thaliana