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Significance Primary endosymbiosis allowed the evolution of complex life on Earth. In this process, a prokaryote was engulfed and retained in the cytoplasm of another microbe, where it developed into a new organelle (mitochondria and plastids). During organelle evolution, genes from the endosymbiont are transferred to the host nuclear genome, where they must become active despite differences in the genetic nature of the “partner” organisms. Here, we show that in the amoeba Paulinella micropora , which harbors a nascent photosynthetic organelle, the “copy-paste” mechanism of retrotransposition allowed domestication of endosymbiont-derived genes in the host nuclear genome. This duplication mechanism is widespread in eukaryotes and may be a major facilitator for host–endosymbiont integration and the evolution of organelles.

Significance Most forms of life on Earth cannot exist without photosynthesis. Our food and atmosphere depend on it. To obtain high photosynthetic yields, light energy must be efficiently coupled to the fixation of CO2 into organic molecules. Suboptimal environmental conditions can severely impact the conversion of light energy to biomass and lead to reactive oxygen production, which in turn can cause cellular damage and loss of productivity. Hence, plants, algae, and photosynthetic bacteria have evolved a network of alternative outlets to sustain the flow of photosynthetically derived electrons. Our work is focused on the nature and integration of these outlets, which will inform a rational engineering of crop plants and algae to optimize photosynthesis and meet the increased global demand for food., Environmental stresses dramatically impact the balance between the production of photosynthetically derived energetic electrons and Calvin–Benson–Bassham cycle (CBBC) activity; an imbalance promotes accumulation of reactive oxygen species and causes cell damage. Hence, photosynthetic organisms have developed several strategies to route electrons toward alternative outlets that allow for storage or harmless dissipation of their energy. In this work, we explore the activities of three essential outlets associated with Chlamydomonas reinhardtii photosynthetic electron transport: (i) reduction of O2 to H2O through flavodiiron proteins (FLVs) and (ii) plastid terminal oxidases (PTOX) and (iii) the synthesis of starch. Real-time measurements of O2 exchange have demonstrated that FLVs immediately engage during dark-to-light transitions, allowing electron transport when the CBBC is not fully activated. Under these conditions, we quantified maximal FLV activity and its overall capacity to direct photosynthetic electrons toward O2 reduction. However, when starch synthesis is compromised, a greater proportion of the electrons is directed toward O2 reduction through both the FLVs and PTOX, suggesting an important role for starch synthesis in priming/regulating CBBC and electron transport. Moreover, partitioning energized electrons between sustainable (starch; energetic electrons are recaptured) and nonsustainable (H2O; energetic electrons are not recaptured) outlets is part of the energy management strategy of photosynthetic organisms that allows them to cope with the fluctuating conditions encountered in nature. Finally, unmasking the repertoire and control of such energetic reactions offers new directions for rational redesign and optimization of photosynthesis to satisfy global demands for food and other resources.

The maintenance of functional chloroplasts in photosynthetic eukaryotes requires real-time coordination of the nuclear and plastid genomes. Tetrapyrroles play a significant role in plastid-to-nucleus retrograde signaling in plants to ensure that nuclear gene expression is attuned to the needs of the chloroplast. Well-known sites of synthesis of chlorophyll for photosynthesis, plant chloroplasts also export heme and heme-derived linear tetrapyrroles (bilins), two critical metabolites respectively required for essential cellular activities and for light sensing by phytochromes. Here we establish that Chlamydomonas reinhardtii, one of many chlorophyte species that lack phytochromes, can synthesize bilins in both plastid and cytosol compartments. Genetic analyses show that both pathways contribute to iron acquisition from extracellular heme, whereas the plastid-localized pathway is essential for light-dependent greening and phototrophic growth. Our discovery of a bilin-dependent nuclear gene network implicates a widespread use of bilins as retrograde signals in oxygenic photosynthetic species. Our studies also suggest that bilins trigger critical metabolic pathways to detoxify molecular oxygen produced by photosynthesis, thereby permitting survival and phototrophic growth during the light period.

Endosymbiotic acquisition of bacteria by a protist, with subsequent evolution of the bacteria into mitochondria and plastids, had a transformative impact on eukaryotic biology. Reconstructing events that created a stable association between endosymbiont and host during the process of organellogenesis—including establishment of regulated protein import into nascent organelles—is difficult because they date back more than 1 billion years. The amoeba Paulinella chromatophora contains nascent photosynthetic organelles of more recent evolutionary origin (∼60 Mya) termed chromatophores (CRs). After the initial endosymbiotic event, the CR genome was reduced to approximately 30% of its presumed original size and more than 30 expressed genes were transferred from the CR to the amoebal nuclear genome. Three transferred genes— psaE , psaK1 , and psaK2 —encode subunits of photosystem I. Here we report biochemical evidence that PsaE, PsaK1, and PsaK2 are synthesized in the amoeba cytoplasm and traffic into CRs, where they assemble with CR-encoded subunits into photosystem I complexes. Additionally, our data suggest that proteins routed to CRs pass through the Golgi apparatus. Whereas genome reduction and transfer of genes from bacterial to host genome have been reported to occur in other obligate bacterial endosymbioses, this report outlines the import of proteins encoded by such transferred genes into the compartment derived from the bacterial endosymbiont. Our study showcases P . chromatophora as an exceptional model in which to study early events in organellogenesis, and suggests that protein import into bacterial endosymbionts might be a phenomenon much more widespread than currently assumed.

Xanthophyll pigments have critical structural and functional roles in the photosynthetic light-harvesting complexes of algae and vascular plants. Genetic dissection of xanthophyll metabolism in the green alga Chlamydomonas reinhardtii revealed functions for specific xanthophylls in the nonradiative dissipation of excess absorbed light energy, measured as nonphotochemical quenching of chlorophyll fluorescence. Mutants with a defect in either the α- or β-branch of carotenoid biosynthesis exhibited less nonphotochemical quenching but were still able to tolerate high light. In contrast, a double mutant that was defective in the synthesis of lutein, loroxanthin (α-carotene branch), zeaxanthin, and antheraxanthin (β-carotene branch) had almost no nonphotochemical quenching and was extremely sensitive to high light. These results strongly suggest that in addition to the xanthophyll cycle pigments (zeaxanthin and antheraxanthin), α-carotene-derived xanthophylls such as lutein, which are structural components of the subunits of the light-harvesting complexes, contribute to the dissipation of excess absorbed light energy and the protection of plants from photo-oxidative damage.

We have isolated, from the prokaryotic cyanobacterium Synechococcus sp. strain PCC 7942, a gene encoding a protein of 72 amino acids [designated high light inducible protein (HLIP)] with similarity to the extended family of eukaryotic chlorophyll a/b binding proteins (CABs). HLIP has a single membrane-spanning alpha-helix, whereas both the CABs and the related early light inducible proteins have three membrane-spanning helices. Hence, HLIP may represent an evolutionary progenitor of the eukaryotic members of the CAB extended family. We also show that the gene encoding HLIP is induced by high light and blue/UV-A radiation. The evolution, regulation, and potential function of HLIP are discussed.

The polypeptide composition of the phycobilisome, the major light-harvesting complex of prokaryotic cyanobacteria and certain eukaryotic algae, can be modulated by different light qualities in cyanobacteria exhibiting chromatic adaptation. We have identified genomic fragments encoding a cluster of phycobilisome polypeptides (phycobiliproteins) from the chromatically adapting cyanobacterium Fremyella diplosiphon using previously characterized DNA fragments of phycobiliprotein genes from the eukaryotic alga Cyanophora paradoxa and from F. diplosiphon. Characterization of two lambda-EMBL3 clones containing overlapping genomic fragments indicates that three sets of phycobiliprotein genes--the alpha- and beta-allophycocyanin genes plus two sets of alpha- and beta-phycocyanin genes--are clustered within 13 kilobases on the cyanobacterial genome and transcribed off the same strand. The gene order (alpha-allophycocyanin followed by beta-allophycocyanin and beta-phycocyanin followed by alpha-phycocyanin) appears to be a conserved arrangement found previously in a eukaryotic alga and another cyanobacterium. We have reported that one set of phycocyanin genes is transcribed as two abundant red light-induced mRNAs (1600 and 3800 bases). We now present data showing that the allophycocyanin genes and a second set of phycocyanin genes are transcribed into major mRNAs of 1400 and 1600 bases, respectively. These transcripts are present in RNA isolated from cultures grown in red and green light, although lower levels of the 1600-base phycocyanin transcript are present in cells grown in green light. Furthermore, a larger transcript of 1750 bases hybridizes to the allophycocyanin genes and may be a precursor to the 1400-base species.

In vivo labeling of eukaryotic phycobilisomes in the presence of inhibitors of translation on 70S and 80S ribosomes demonstrates that some of the polypeptides of this light-harvesting complex are synthesized in the cytoplasm while others are synthesized in the chloroplast. The major pigmented polypeptides, the α and β subunits of the biliproteins (molecular weights between 15,000 and 20,000) and the anchor protein (molecular weight about 90,000) are translated on 70S ribosomes. This suggests that these polypeptides are made within the algal chloroplast. Because the α and β subunits comprise a group of closely related polypeptides, the genes encoding these polypeptides may reside in the plastid genome as a multigene family. Other prominent phycobilisome polypeptides, including a nonpigmented polypeptide that may be involved in maintaining the structural integrity of the complex, are synthesized on cytoplasmic ribosomes. Because the synthesis of phycobilisomes appears to require the expression of genes in two subcellular compartments, this system may be an excellent model for: ( i ) examining interaction between nuclear and plastid genomes: ( ii ) elucidating the molecular processes involved in the evolution of plastid genes: ( iii ) clarifying the events in the synthesis and assembly of macromolecular complexes in the chloroplast.

The unicellular green alga Chlamydomonas reinhardtii synthesizes carbonic anhydrase in response to low levels of CO 2 (i.e., air levels of CO 2 ). This enzyme, localized predominantly in the periplasmic space of the alga (or associated with the cell wall), is an important component of the machinery required for the active accumulation of inorganic carbon by C. reinhardtii and the saturation of ribulose-1,5-bisphosphate carboxylase at low extracellular carbon concentrations. We have begun to examine the synthesis and compartmentalization of carbonic anhydrase in C. reinhardtii . The monomeric species associated with carbonic anhydrase activity is synthesized as a precursor on 80S cytoplasmic ribosomes. This precursor can be detected immunologically in the profiles of translation products when a reticulocyte lysate, cell-free system is primed with poly(A)-RNA from either air-grown C. reinhardtii or cells shifted from growth on 5% CO 2 to air for 12 hr. It is not synthesized when the in vitro system is primed with poly(A)-RNA from CO 2 -grown algae. Since translatable RNA for the polypeptide responsible for carbonic anhydrase activity was only present in cells that experienced low levels of CO 2 , the adaptation process either involves the regulation of transcription of the carbonic anhydrase gene (and perhaps other genes involved in adaptation) or the post-transcriptional processing of the messenger RNA. Furthermore, the appearance of the mature polypeptide associated with carbonic anhydrase activity in the periplasmic space of C. reinhardtii is inhibited by tunicamycin, an antibiotic that prevents core glycosylation of polypeptides on the endoplasmic reticulum. Together, these results suggest that the biosynthesis of this extracellular algal enzyme involves the translation of mRNA for the carbonic anhydrase monomer on ribosomes bound to the endoplasmic reticulum, the cleavage of a signal sequence during transport of the nascent polypeptide into the lumen of the endoplasmic reticulum, and subsequent glycosylation events prior to export across the plasmalemma.