Recent advances in understanding the assembly and repair of photosystem II

Background

Photosystem II (PSII) is the light-driven water:plastoquinone oxidoreductase of oxygenic photosynthesis and is found in the thylakoid membrane of chloroplasts and cyanobacteria. Considerable attention is focused on how PSII is assembled in vivo and how it is repaired following irreversible damage by visible light (so-called photoinhibition). Understanding these processes might lead to the development of plants with improved growth characteristics especially under conditions of abiotic stress.

Scope

Here we summarize recent results on the assembly and repair of PSII in cyanobacteria, which are excellent model organisms to study higher plant photosynthesis.

Conclusions

Assembly of PSII is highly co-ordinated and proceeds through a number of distinct assembly intermediates. Associated with these assembly complexes are proteins that are not found in the final functional PSII complex. Structural information and possible functions are beginning to emerge for several of these ‘assembly’ factors, notably Ycf48/Hcf136, Psb27 and Psb28. A number of other auxiliary proteins have been identified that appear to have evolved since the divergence of chloroplasts and cyanobacteria. The repair of PSII involves partial disassembly of the damaged complex, the selective replacement of the damaged sub-unit (predominantly the D1 sub-unit) by a newly synthesized copy, and reassembly. It is likely that chlorophyll released during the repair process is temporarily stored by small CAB-like proteins (SCPs). A model is proposed in which damaged D1 is removed in Synechocystis sp. PCC 6803 by a hetero-oligomeric complex composed of two different types of FtsH sub-unit (FtsH2 and FtsH3), with degradation proceeding from the N-terminus of D1 in a highly processive reaction. It is postulated that a similar mechanism of D1 degradation also operates in chloroplasts. Deg proteases are not required for D1 degradation in Synechocystis 6803 but members of this protease family might play a supplementary role in D1 degradation in chloroplasts under extreme conditions.     

The influence of acetyl phosphate on DspA signalling in the Cyanobacterium <Emphasis Type="Italic">Synechocystis</Emphasis> sp. PCC6803

Background  

The dspA (hik33) gene, coding for a putative sensory histidine kinase, is conserved in plastids (ycf26) and cyanobacteria. It has been linked with a number of different stress responses in cyanobacteria.     

Insights into the complex 3-D architecture of thylakoid membranes in the unicellular cyanobacterium Cyanothece sp. ATCC 51142

In cyanobacteria and chloroplasts, thylakoids are the complex internal membrane system where the light reactions of oxygenic photosynthesis occur. In plant chloroplasts, thylakoids are differentiated into a highly interconnected system of stacked grana and unstacked stroma membranes. In contrast, in cyanobacteria, the evolutionary progenitors of chloroplasts, thylakoids do not routinely form stacked and unstacked regions, and the architecture of the thylakoid membrane systems is only now being described in detail in these organisms. We used electron tomography to examine the thylakoid membrane systems in one cyanobacterium, Cyanothece sp. ATCC 51142. Our data showed that thylakoids form a complicated branched network with a rudimentary quasi-helical architecture in this organism. A well accepted helical model of grana-stroma architecture of plant thylakoids describes an organization in which stroma thylakoids wind around stacked granum in right-handed spirals. Here we present data showing that the simplified helical architecture in Cyanothece 51142 is lefthanded in nature. We propose a model comparing the thylakoid membranes in plants and this cyanobacterium in which the system in Cyanothece 51142 is composed of non-stacked membranes linked by fret-like connections to other membrane components of the system in a limited left-handed arrangement.Key words: cyanobacteria, Cyanothece 51142, thylakoid membrane, electron tomography, chloroplast     

Outer membrane proteins can be simply identified using secondary structure element alignment

Background  

Outer membrane proteins (OMPs) are frequently found in the outer membranes of gram-negative bacteria, mitochondria and chloroplasts and have been found to play diverse functional roles. Computational discrimination of OMPs from globular proteins and other types of membrane proteins is helpful to accelerate new genome annotation and drug discovery.     

The YlmG protein has a conserved function related to the distribution of nucleoids in chloroplasts and cyanobacteria

Background  

Reminiscent of their free-living cyanobacterial ancestor, chloroplasts proliferate by division coupled with the partition of nucleoids (DNA-protein complexes). Division of the chloroplast envelope membrane is performed by constriction of the ring structures at the division site. During division, nucleoids also change their shape and are distributed essentially equally to the daughter chloroplasts. Although several components of the envelope division machinery have been identified and characterized, little is known about the molecular components/mechanisms underlying the change of the nucleoid structure.     

LysoPC acyltransferase/PC transacylase activities in plant plasma membrane and plasma membrane-associated endoplasmic reticulum

Background  

The phospholipids of the plant plasma membrane are synthesized in the endoplasmic reticulum (ER). The majority of these lipids reach the plasma membrane independently of the secretory vesicular pathway. Phospholipid delivery to the mitochondria and chloroplasts of plant cells also bypasses the secretory pathway and here it has been proposed that lysophospholipids are transported at contact sites between specific regions of the ER and the respective organelle, followed by lysophospholipid acylation in the target organelle. To test the hypothesis that a corresponding mechanism operates to transport phospholipids to the plasma membrane outside the secretory pathway, we investigated whether lysolipid acylation occurs also in the plant plasma membrane and whether this membrane, like the chloroplasts and mitochondria, is in close contact with the ER.     

Specific role of phosphatidylglycerol and functional overlaps with other thylakoid lipids in Arabidopsis chloroplast biogenesis

Key message

With phosphate deficiency, the role of phosphatidylglycerol is compensated by increased glycolipid content in thylakoid membrane biogenesis but not photosynthetic electron transport in Arabidopsis chloroplasts.

Abstract

In plants and cyanobacteria, anionic phosphatidylglycerol (PG) is the only major phospholipid in thylakoid membranes, where neutral galactolipids monogalactosyldiacylglycerol (MGDG) and digalactosyldiacylglycerol (DGDG) are predominant. In addition to provide a lipid bilayer matrix, PG plays a specific role in photosynthetic electron transport. Non-phosphorous sulfoquinovosyldiacylglycerol (SQDG) is another anionic lipid in thylakoids; it substitutes for PG under phosphate (Pi) deficiency to maintain proper balance of anionic charge in thylakoid membranes. Although the crucial role of PG in photosynthesis has been deeply analyzed in cyanobacteria, its physiological function in seed plants other than photosynthesis remains unclear. To reveal specific roles of PG and functional overlaps with other thylakoid lipids, we characterized a PG-deficient Arabidopsis mutant (pgp1-2) under Pi-controlled conditions. Under Pi-sufficient conditions, the proportion of PG and other thylakoid lipids was decreased in pgp1-2, which led to severe disruption of thylakoid membrane biogenesis. Under Pi-deficient conditions, the proportion of all glycolipids in the mutant was greatly increased, with that of PG further decreased. In Pi-deficient pgp1-2, thylakoid membranes remarkably developed, which was accompanied by a change in nucleoid morphology and restored expression of nuclear- and plastid-encoded photosynthesis genes. Increase in glycolipid content with Pi deficiency may compensate for the loss of PG in terms of thylakoid membrane biogenesis. Although Pi deficiency increased chlorophyll and photosynthesis protein content in pgp1-2, it critically decreased photochemical activity in PSII. Further deprivation of PG in photosynthesis complexes may abolish the PSII activity in Pi-deficient pgp1-2, which suggests that glycolipids cannot replace PG in photosynthesis.

     

Fluorescence staining of live cyanobacterial cells suggest non-stringent chromosome segregation and absence of a connection between cytoplasmic and thylakoid membranes

Background  

In spite of their abundance and importance, little is known about cyanobacterial cell biology and their cell cycle. During each cell cycle, chromosomes must be separated into future daughter cells, i.e. into both cell halves, which in many bacteria is achieved by an active machinery that operates during DNA replication. Many cyanobacteria contain multiple identical copies of the chromosome, but it is unknown how chromosomes are segregated into future daughter cells, and if an active or passive mechanism is operative. In addition to an outer and an inner cell membrane, cyanobacteria contain internal thylakoid membranes that carry the active photosynthetic machinery. It is unclear whether thylakoid membranes are invaginations of the inner cell membrane, or an independent membrane system.     

Towards a synthetic chloroplast

Background

The evolution of eukaryotic cells is widely agreed to have proceeded through a series of endosymbiotic events between larger cells and proteobacteria or cyanobacteria, leading to the formation of mitochondria or chloroplasts, respectively. Engineered endosymbiotic relationships between different species of cells are a valuable tool for synthetic biology, where engineered pathways based on two species could take advantage of the unique abilities of each mutualistic partner.

Results

We explored the possibility of using the photosynthetic bacterium Synechococcus elongatus PCC 7942 as a platform for studying evolutionary dynamics and for designing two-species synthetic biological systems. We observed that the cyanobacteria were relatively harmless to eukaryotic host cells compared to Escherichia coli when injected into the embryos of zebrafish, Danio rerio, or taken up by mammalian macrophages. In addition, when engineered with invasin from Yersinia pestis and listeriolysin O from Listeria monocytogenes, S. elongatus was able to invade cultured mammalian cells and divide inside macrophages.

Conclusion

Our results show that it is possible to engineer photosynthetic bacteria to invade the cytoplasm of mammalian cells for further engineering and applications in synthetic biology. Engineered invasive but non-pathogenic or immunogenic photosynthetic bacteria have great potential as synthetic biological devices.     

Fluorescence microscopy and radiolabeling of C3 and C4 chloroplasts using diisothiocyanatostilbene disulfonic acid as a marker for the phosphate translocator

The usefulness of 4,4-diisothiocyanatostilbene-2,2-disulfonic acid (DIDS) for in-situ studies of the chloroplast phosphate translocator was evaluated by fluorescence microscopy and radiolabeling of spinach (Spinacia oleracea L.) (C₃ plant) and maize (Zea mays L.) (C₄ plant) chloroplasts. In maize mesophyll and bundle-sheath chloroplasts and in spinach chloroplasts that were either intact, broken or swollen, DIDS fluorescence was only associated with the chloroplast envelope. Intact chloroplasts often had fluorescent patches corresponding to concave regions of the chloroplast which we assume to be regions enriched in DIDS-binding sites.Incubation of intact or broken spinach chloroplasts or maize mesophyll chloroplasts with [³H₂]DIDS resulted in the labeling of a single polypeptide (relative molecular mass, M_r, 30 kDa) in the envelope fraction, in each case. Label in the stromal fraction was not detected when intact chloroplasts were incubated with [³H₂]DIDS. However, when broken chloroplasts were incubated with [³H₂]DIDS, several polypeptides of various molecular masses were labeled, but not the 30×31-kDa polypeptide. In thylakoid fractions from both broken and intact chloroplasts, a single 30×31-kDa polypeptide was labeled inconsistently. When a mixture of intact maize mesophyll and bundle-sheath chloroplasts was labeled with [³H₂]DIDS, extracts of whole chloroplasts displayed radioactivity only in the 30×31-kDa band.We conclude that DIDS is a valuable probe for the in-situ identification and characterization of the 30-kDa protein — the presumptive phosphate translocator — in C₃ and C₄ chloroplasts since DIDS (1) does not penetrate the inner membrane of the envelope of intact chloroplasts and, therefore, (2) does not bind internal sites in intact chloroplasts, and (3) only binds the 30-kDa protein in the inner membrane of the envelope.Abbreviations CBB Coomassie brilliant blue - DIC differential interference contrast optics - DIDS 4,4-diisothiocyanatostilbene-2,2-disulfonic acid - [³H₂]DIDS 1,2-ditritio-1,2-(2,2-disulfo-4,4-diisothiocyano)diphenylethane - kDa kilodalton - M_r relative molecular mass - PGA 3-phosphoglycerate - Pitranslocator phosphate translocator - SDS sodium dodecyl sulfate     

Complete DNA sequences of the plastid genomes of two parasitic flowering plant species, <Emphasis Type="Italic">Cuscuta reflexa</Emphasis> and <Emphasis Type="Italic">Cuscuta gronovii</Emphasis>

Background  

The holoparasitic plant genus Cuscuta comprises species with photosynthetic capacity and functional chloroplasts as well as achlorophyllous and intermediate forms with restricted photosynthetic activity and degenerated chloroplasts. Previous data indicated significant differences with respect to the plastid genome coding capacity in different Cuscuta species that could correlate with their photosynthetic activity. In order to shed light on the molecular changes accompanying the parasitic lifestyle, we sequenced the plastid chromosomes of the two species Cuscuta reflexa and Cuscuta gronovii. Both species are capable of performing photosynthesis, albeit with varying efficiencies. Together with the plastid genome of Epifagus virginiana, an achlorophyllous parasitic plant whose plastid genome has been sequenced, these species represent a series of progression towards total dependency on the host plant, ranging from reduced levels of photosynthesis in C. reflexa to a restricted photosynthetic activity and degenerated chloroplasts in C. gronovii to an achlorophyllous state in E. virginiana.     

Trade-off between C and N recycling and N<Subscript>2</Subscript>O emissions of soils with summer cover crops in subtropical agrosystems

Aims

Biological soil crusts (biocrusts) are soil-surface communities in drylands, dominated by cyanobacteria, mosses, and lichens. They provide key ecosystem functions by increasing soil stability and influencing soil hydrologic, nutrient, and carbon cycles. Because of this, methods to reestablish biocrusts in damaged drylands are needed. Here we test the reintroduction of field-collected vs. greenhouse-cultured biocrusts for rehabilitation.

Methods

We collected biocrusts for 1) direct reapplication, and 2) artificial cultivation under varying hydration regimes. We added field-collected and cultivated biocrusts (with and without hardening treatments) to bare field plots and monitored establishment.

Results

Both field-collected and cultivated cyanobacteria increased cover dramatically during the experimental period. Cultivated biocrusts established more rapidly than field-collected biocrusts, attaining ~82% cover in only one year, but addition of field-collected biocrusts led to higher species richness, biomass (as assessed by chlorophyll a) and level of development. Mosses and lichens did not establish well in either case, but late successional cover was affected by hardening and culture conditions.

Conclusions

This study provides further evidence that it is possible to culture biocrust components from later successional materials and reestablish cultured organisms in the field. However, more research is needed into effective reclamation techniques.

     

Fatty Acid Activation in Cyanobacteria Mediated by Acyl-Acyl Carrier Protein Synthetase Enables Fatty Acid Recycling

In cyanobacteria fatty acids destined for lipid synthesis can be synthesized de novo, but also exogenous free fatty acids from the culture medium can be directly incorporated into lipids. Activation of exogenous fatty acids is likely required prior to their utilization. To identify the enzymatic activity responsible for activation we cloned candidate genes from Synechocystis sp. PCC 6803 and Synechococcus elongatus PCC 7942 and identified the encoded proteins as acyl-acyl carrier protein synthetases (Aas). The enzymes catalyze the ATP-dependent esterification of fatty acids to the thiol of acyl carrier protein. The two protein sequences are only distantly related to known prokaryotic Aas proteins but they display strong similarity to sequences that can be found in almost all organisms that perform oxygenic photosynthesis. To investigate the biological role of Aas activity in cyanobacteria, aas knockout mutants were generated in the background of Synechocystis sp. PCC 6803 and S. elongatus PCC 7942. The mutant strains showed two phenotypes characterized by the inability to utilize exogenous fatty acids and by the secretion of endogenous fatty acids into the culture medium. The analyses of extracellular and intracellular fatty acid profiles of aas mutant strains as well as labeling experiments indicated that the detected free fatty acids are released from membrane lipids. The data suggest a considerable turnover of lipid molecules and a role for Aas activity in recycling the released fatty acids. In this model, lipid degradation represents a third supply of fatty acids for lipid synthesis in cyanobacteria.Cyanobacteria present a diverse group of Gram-negative bacteria capable of oxygenic photosynthesis (Margulis, 1975). Their two photosystems, as well as other genetic and morphological similarities, identified them as putative predecessors of chloroplasts of eukaryotic plants (Wallace, 1982; Pakrasi, 1995). The structural similarities of cyanobacteria and chloroplasts are reflected in part by equivalence of biochemical pathways and their components. For instance, cyanobacterial fatty acid and glycerolipid compositions closely resemble those of the inner envelope and thylakoid membranes of chloroplasts (Roughan et al., 1980; Heinz and Roughan, 1983). In cyanobacteria, as well as in chloroplasts, fatty acids are synthesized by a type II fatty acid synthase (FAS) complex utilizing a freely dissociable acyl carrier protein (ACP; Froehlich et al., 1990). The products of FAS are released as acyl ACPs and may serve directly as substrates for acyltransferases, incorporating the fatty acids into membrane lipids (Frentzen et al., 1983). The substrate specificity of the acyltransferases establishes in cyanobacteria as well as in plastids the typical prokaryotic fatty acid pattern characterized by C16 fatty acids esterified to the sn-2 position. The correspondence of metabolic pathways between cyanobacteria and chloroplasts is reflected by the shared presence of closely related enzymes that catalyze key reactions. Besides the many similarities, however, there are also clear discrepancies that in part account for the fact that cyanobacteria are unicellular organisms, whereas chloroplasts are embedded in the metabolism of a eukaryotic cell. In terms of lipid metabolism, such differences become obvious if one considers the fact that the plastidial FAS also supplies the extraplastidic compartment with fatty acids (Browse et al., 1986). Fatty acid export from the chloroplast necessitates the release of synthesized acyl chains from ACP to allow transport across both envelope membranes. The release is achieved by the action of acyl-ACP thioesterases that hydrolyze the acyl-ACP thioester to liberate the fatty acid (Voelker et al., 1997). In cyanobacteria such export would obviously result in an unfavorable loss of fatty acids, and consequently homologous proteins to acyl-ACP thioesterases cannot be found here. Whereas cyanobacteria seem to be unable to release fatty acids enzymatically from their activated state, all cyanobacterial genomes available to date encode an activity most likely responsible for the activation of free fatty acids. The respective sequences are annotated as acyl-CoA synthetases. Conserved motifs in the amino acid sequence identify these proteins as members of the well-established superfamily of AMP-binding proteins. This protein family comprises several hundred amino acid sequences spreading across all organisms analyzed so far. The family members are annotated in the PROSITE database under entry number PS00455. Although these predicted fatty acid-activating enzymes of cyanobacteria are annotated as acyl-CoA synthetases due to their sequence similarity to proteins with such enzymatic activity, there is a much higher degree of similarity to certain AMP-binding proteins of plant origin with less-well-established function. These plant proteins are predicted to reside in chloroplasts and one member of this subgroup from Arabidopsis (Arabidopsis thaliana) designated as AAE15 was recently described as acyl-ACP synthetase. The conclusions were based on the comparison of enzymatic activity between plant extracts of wild-type and knockout mutant lines (Koo et al., 2005). Whereas the biological role of this activity remained largely elusive, it was shown that the capacity of plant extracts to elongate supplied medium fatty acids depended on AAE15 activity. Since the elongation of medium chain fatty acids in the plastid depends on the FAS requiring acyl ACPs, it was concluded that the fatty acids must have been activated by ACP. The elongated fatty acids ultimately appeared in membrane lipids. Together these findings suggested that AAE15 is an acyl-ACP synthetase.Besides encoding a protein homologous to AAE15 from Arabidopsis, cyanobacteria are also able to utilize exogenous fatty acids like it was shown for isolated chloroplasts. It is well established that feeding different cyanobacteria with free fatty acids results in the incorporation of these fatty acids into membrane lipids. For this process the activation of the fatty acids is believed to be essential. This causal relationship was clearly shown at least for other unicellular organisms like Escherichia coli and yeast (Saccharomyces cerevisiae) where the deletion of acyl-CoA synthetase activity resulted in the inability to utilize exogenous fatty acids (Overath et al., 1969; Knoll et al., 1995). It is not easy to assess how regularly cyanobacterial cells are exposed to exogenous free fatty acids in nature but at least for marine strains this is most likely a rather artificial situation. Therefore, it can be speculated that the capacity to activate free fatty acids might be of different relevance in the lipid metabolism of cyanobacteria in vivo.In this article, we investigated the fatty acid metabolism of cyanobacteria. We isolated candidate genes potentially encoding enzymes involved in fatty acid activation from the strains Synechocystis sp. PCC 6803 (hereafter Synechocystis) and Synechococcus elongatus PCC 7942 (hereafter Synechococcus) and performed heterologous expression in E. coli. The recombinant proteins were shown to possess acyl-ACP synthetase activity with broad substrate specificity. Knockout mutant strains deficient in acyl-ACP synthetase activity were characterized by secretion of endogenous free fatty acids into the culture medium. Combined with labeling experiments, the results suggest an essential role for acyl-ACP synthetase in fatty acid recycling in cyanobacteria.     

Copine A,a calcium-dependent membrane-binding protein,transiently localizes to the plasma membrane and intracellular vacuoles in <Emphasis Type="Italic">Dictyostelium</Emphasis>

Background  

Copines are soluble, calcium-dependent membrane binding proteins found in a variety of organisms. Copines are characterized as having two C2 domains at the N-terminal region followed by an "A domain" at the C-terminal region. The "A domain" is similar in sequence to the von Willebrand A (VWA) domain found in integrins. The presence of C2 domains suggests that copines may be involved in cell signaling and/or membrane trafficking pathways.     

Transfer of fixed-N from N2-fixing cyanobacteria associated with the moss Sphagnum riparium results in enhanced growth of the moss

Background

Despite the general assumption that nitrogen fixed by associated cyanobacteria will be readily utilised for growth by the Sphagnum, no empirical evidence is available in the literature. Therefore the effects of nitrogen transfer from cyanobacteria associated with S. riparium were investigated.

Methods

Cultivation of S. riparium with and without cyanobacteria was performed under laboratory conditions for 57 days.

Results

We show that nitrogen fixation by cyanobacteria associated with Sphagnum mosses, influences moss growth by transfer of fixed nitrogen to the moss. More than 35 % of the nitrogen fixed by cyanobacteria was transferred to the newly formed moss biomass and resulted in an increase in the growth of Sphagnum biomass compared to the controls. The variation in the increase of nitrogen content explained 76 % of the biomass increment.

Conclusion

Hence, nitrogen fixation will have immediate effect on the carbon fixation by Sphagnum. This shows that factors regulating nitrogen fixation will have a direct effect on the role of Sphagnum dominated ecosystems with respect to carbon cycling.     

Alkane-induced expression,substrate binding profile,and immunolocalization of a cytochrome P450 encoded on the <Emphasis Type="Italic">nifD</Emphasis> excision element of <Emphasis Type="Italic">Anabaena</Emphasis> 7120

Background  

Alkanes have been hypothesized to act as universal inducers of bacterial cytochrome P450 gene expression. We tested this hypothesis on an unusual P450 gene (cyp110) found on a conserved 11 kilobase episomal DNA element of unknown function found in filamentous cyanobacteria. We also monitored the binding of potential substrates to the P450 protein and explored the distribution of P450 protein in vegetative cells and nitrogen-fixing heterocysts using immuno-electron microscopy.     

TonB-dependent transporters and their occurrence in cyanobacteria

Background  

Different iron transport systems evolved in Gram-negative bacteria during evolution. Most of the transport systems depend on outer membrane localized TonB-dependent transporters (TBDTs), a periplasma-facing TonB protein and a plasma membrane localized machinery (ExbBD). So far, iron chelators (siderophores), oligosaccharides and polypeptides have been identified as substrates of TBDTs. For iron transport, three uptake systems are defined: the lactoferrin/transferrin binding proteins, the porphyrin-dependent transporters and the siderophore-dependent transporters. However, for cyanobacteria almost nothing is known about possible TonB-dependent uptake systems for iron or other substrates.     

Evolution of mitochondrial TAT translocases illustrates the loss of bacterial protein transport machines in mitochondria

Background

Bacteria and mitochondria contain translocases that function to transport proteins across or insert proteins into their inner and outer membranes. Extant mitochondria retain some bacterial-derived translocases but have lost others. While BamA and YidC were integrated into general mitochondrial protein transport pathways (as Sam50 and Oxa1), the inner membrane TAT translocase, which uniquely transports folded proteins across the membrane, was retained sporadically across the eukaryote tree.

Results

We have identified mitochondrial TAT machinery in diverse eukaryotic lineages and define three different types of eukaryote-encoded TatABC-derived machineries (TatAC, TatBC and TatC-only). Here, we investigate TatAC and TatC-only machineries, which have not been studied previously. We show that mitochondria-encoded TatAC of the jakobid Andalucia godoyi represent the minimal functional pathway capable of substituting for the Escherichia coli TatABC complex and can transport at least one substrate. However, selected TatC-only machineries, from multiple eukaryotic lineages, were not capable of supporting the translocation of this substrate across the bacterial membrane. Despite the multiple losses of the TatC gene from the mitochondrial genome, the gene was never transferred to the cell nucleus. Although the major constraint preventing nuclear transfer of mitochondrial TatC is likely its high hydrophobicity, we show that in chloroplasts, such transfer of TatC was made possible due to modifications of the first transmembrane domain.

Conclusions

At its origin, mitochondria inherited three inner membrane translocases Sec, TAT and Oxa1 (YidC) from its bacterial ancestor. Our work shows for the first time that mitochondrial TAT has likely retained its unique function of transporting folded proteins at least in those few eukaryotes with TatA and TatC subunits encoded in the mitochondrial genome. However, mitochondria, in contrast to chloroplasts, abandoned the machinery multiple times in evolution. The overall lower hydrophobicity of the Oxa1 protein was likely the main reason why this translocase was nearly universally retained in mitochondrial biogenesis pathways.