Uwe-G Maier

Publications of Uwe-G Maier

• Chloroplast Omp85 proteins change orientation during evolution.

  Authors: Maik S Sommer, Bertram Daum, Lucia E Gross, Benjamin L M Weis, Oliver Mirus, Lars Abram, Uwe-G Maier, Werner Kühlbrandt, Enrico Schleiff

  Proceedings of the National Academy of Sciences of the United States of America. 08/2011; 108(33):13841-6.

  The majority of outer membrane proteins (OMPs) from gram-negative bacteria and many of mitochondria and chloroplasts are β-barrels. Insertion and assembly of these proteins are catalyzed by the Omp85The majority of outer membrane proteins (OMPs) from gram-negative bacteria and many of mitochondria and chloroplasts are β-barrels. Insertion and assembly of these proteins are catalyzed by the Omp85 protein family in a seemingly conserved process. All members of this family exhibit a characteristic N-terminal polypeptide-transport-associated (POTRA) and a C-terminal 16-stranded β-barrel domain. In plants, two phylogenetically distinct and essential Omp85's exist in the chloroplast outer membrane, namely Toc75-III and Toc75-V. Whereas Toc75-V, similar to the mitochondrial Sam50, is thought to possess the original bacterial function, its homolog, Toc75-III, evolved to the pore-forming unit of the TOC translocon for preprotein import. In all current models of OMP biogenesis and preprotein translocation, a topology of Omp85 with the POTRA domain in the periplasm or intermembrane space is assumed. Using self-assembly GFP-based in vivo experiments and in situ topology studies by electron cryotomography, we show that the POTRA domains of both Toc75-III and Toc75-V are exposed to the cytoplasm. This unexpected finding explains many experimental observations and requires a reevaluation of current models of OMP biogenesis and TOC complex function.

• Making new out of old: recycling and modification of an ancient protein translocation system during eukaryotic evolution. Mechanistic comparison and phylogenetic analysis of ERAD, SELMA and the peroxisomal importomer.

  Authors: Kathrin Bolte, Nicole Gruenheit, Gregor Felsner, Maik S Sommer, Uwe-G Maier, Franziska Hempel

  BioEssays : news and reviews in molecular, cellular and developmental biology. 03/2011; 33(5):368-76.

  At first glance the three eukaryotic protein translocation machineries--the ER-associated degradation (ERAD) transport apparatus of the endoplasmic reticulum, the peroxisomal importomer and SELMA,At first glance the three eukaryotic protein translocation machineries--the ER-associated degradation (ERAD) transport apparatus of the endoplasmic reticulum, the peroxisomal importomer and SELMA, the pre-protein translocator of complex plastids--appear quite different. However, mechanistic comparisons and phylogenetic analyses presented here suggest that all three translocation machineries share a common ancestral origin, which highlights the recycling of pre-existing components as an effective evolutionary driving force. Editor's suggested further reading in BioEssays ERAD ubiquitin ligases Abstract.

• Omp85 in eukaryotic systems: one protein family with distinct functions.

  Authors: Enrico Schleiff, Uwe G Maier, Thomas Becker

  Biological chemistry. 01/2011; 392(1-2):21-7.

  Omp85-like proteins are evolutionary ancient components of bacterial outer membranes and their evolutionary offspring. As a consequence, proteins of this family can be found in the outer membraneOmp85-like proteins are evolutionary ancient components of bacterial outer membranes and their evolutionary offspring. As a consequence, proteins of this family can be found in the outer membrane systems of Gram-negative bacteria and endosymbiotically derived organelles. In the different membranes, they perform distinct functions such as catalyzing protein insertion into or protein transport across the bilayer. Here, the knowledge on the Omp85-like proteins in the eukaryotic system with regard to structural properties and physiological behavior is summarized.

• In silico and in vivo investigations of proteins of a minimized eukaryotic cytoplasm.

  Authors: Daniel Moog, Simone Stork, Stefan Zauner, Uwe-G Maier

  Genome biology and evolution. 01/2011; 3:375-82.

  Algae with secondary plastids such as diatoms maintain two different eukaryotic cytoplasms. One of them, the so-called periplastidal compartment (PPC), is the naturally minimized cytoplasm of aAlgae with secondary plastids such as diatoms maintain two different eukaryotic cytoplasms. One of them, the so-called periplastidal compartment (PPC), is the naturally minimized cytoplasm of a eukaryotic endosymbiont. In order to investigate the protein composition of the PPC of diatoms, we applied knowledge of the targeting signals of PPC-directed proteins in searches of the genome data for proteins acting in the PPC and proved their in vivo localization via expressing green fluorescent protein (GFP) fusions. Our investigation increased the knowledge of the protein content of the PPC approximately 3-fold and thereby indicated that this narrow compartment was functionally reduced to some important cellular functions with nearly no housekeeping biochemical pathways.

• Microalgae as bioreactors for bioplastic production.

  Authors: Franziska Hempel, Andrew S Bozarth, Nicole Lindenkamp, Andreas Klingl, Stefan Zauner, Uwe Linne, Alexander Steinbüchel, Uwe G Maier

  Microbial cell factories. 01/2011; 10:81.

  Poly-3-hydroxybutyrate (PHB) is a polyester with thermoplastic properties that is naturally occurring and produced by such bacteria as Ralstonia eutropha H16 and Bacillus megaterium. In contrast toPoly-3-hydroxybutyrate (PHB) is a polyester with thermoplastic properties that is naturally occurring and produced by such bacteria as Ralstonia eutropha H16 and Bacillus megaterium. In contrast to currently utilized plastics and most synthetic polymers, PHB is biodegradable, and its production is not dependent on fossil resources making this bioplastic interesting for various industrial applications. In this study, we report on introducing the bacterial PHB pathway of R. eutropha H16 into the diatom Phaeodactylum tricornutum, thereby demonstrating for the first time that PHB production is feasible in a microalgal system. Expression of the bacterial enzymes was sufficient to result in PHB levels of up to 10.6% of algal dry weight. The bioplastic accumulated in granule-like structures in the cytosol of the cells, as shown by light and electron microscopy. Our studies demonstrate the great potential of microalgae like the diatom P. tricornutum to serve as solar-powered expression factories and reveal great advantages compared to plant based production systems.

• A single peroxisomal targeting signal mediates matrix protein import in diatoms.

  Authors: Nicola H Gonzalez, Gregor Felsner, Frederic D Schramm, Andreas Klingl, Uwe-G Maier, Kathrin Bolte

  PloS one. 01/2011; 6(9):e25316.

  Peroxisomes are single membrane bound compartments. They are thought to be present in almost all eukaryotic cells, although the bulk of our knowledge about peroxisomes has been generated from only aPeroxisomes are single membrane bound compartments. They are thought to be present in almost all eukaryotic cells, although the bulk of our knowledge about peroxisomes has been generated from only a handful of model organisms. Peroxisomal matrix proteins are synthesized cytosolically and posttranslationally imported into the peroxisomal matrix. The import is generally thought to be mediated by two different targeting signals. These are respectively recognized by the two import receptor proteins Pex5 and Pex7, which facilitate transport across the peroxisomal membrane. Here, we show the first in vivo localization studies of peroxisomes in a representative organism of the ecologically relevant group of diatoms using fluorescence and transmission electron microscopy. By expression of various homologous and heterologous fusion proteins we demonstrate that targeting of Phaeodactylum tricornutum peroxisomal matrix proteins is mediated only by PTS1 targeting signals, also for proteins that are in other systems imported via a PTS2 mode of action. Additional in silico analyses suggest this surprising finding may also apply to further diatoms. Our data suggest that loss of the PTS2 peroxisomal import signal is not reserved to Caenorhabditis elegans as a single exception, but has also occurred in evolutionary divergent organisms. Obviously, targeting switching from PTS2 to PTS1 across different major eukaryotic groups might have occurred for different reasons. Thus, our findings question the widespread assumption that import of peroxisomal matrix proteins is generally mediated by two different targeting signals. Our results implicate that there apparently must have been an event causing the loss of one targeting signal even in the group of diatoms. Different possibilities are discussed that indicate multiple reasons for the detected targeting switching from PTS2 to PTS1.

• Algae as protein factories: expression of a human antibody and the respective antigen in the diatom Phaeodactylum tricornutum.

  Authors: Franziska Hempel, Julia Lau, Andreas Klingl, Uwe G Maier

  PloS one. 01/2011; 6(12):e28424.

  Microalgae are thought to offer great potential as expression system for various industrial, therapeutic and diagnostic recombinant proteins as they combine high growth rates with all benefits ofMicroalgae are thought to offer great potential as expression system for various industrial, therapeutic and diagnostic recombinant proteins as they combine high growth rates with all benefits of eukaryotic expression systems. Moreover, microalgae exhibit a phototrophic lifestyle like land plants, hence protein expression is fuelled by photosynthesis, which is CO(2)-neutral and involves only low production costs. So far, however, research on algal bioreactors for recombinant protein expression is very rare calling for further investigations in this highly promising field. In this study, we present data on the expression of a monoclonal human IgG antibody against the Hepatitis B surface protein and the respective antigen in the diatom Phaeodactylum tricornutum. Antibodies are fully-assembled and functional and accumulate to 8.7% of total soluble protein, which complies with 21 mg antibody per gram algal dry weight. The Hepatitis B surface protein is functional as well and is recognized by algae-produced and commercial antibodies.

• ERAD components in organisms with complex red plastids suggest recruitment of a preexisting protein transport pathway for the periplastid membrane.

  Authors: Gregor Felsner, Maik S Sommer, Nicole Gruenheit, Franziska Hempel, Daniel Moog, Stefan Zauner, William Martin, Uwe G Maier

  Genome biology and evolution. 11/2010; 3:140-50.

  The plastids of cryptophytes, haptophytes, and heterokontophytes (stramenopiles) (together once known as chromists) are surrounded by four membranes, reflecting the origin of these plastids throughThe plastids of cryptophytes, haptophytes, and heterokontophytes (stramenopiles) (together once known as chromists) are surrounded by four membranes, reflecting the origin of these plastids through secondary endosymbiosis. They share this trait with apicomplexans, which are alveolates, the plastids of which have been suggested to stem from the same secondary symbiotic event and therefore form a phylogenetic clade, the chromalveolates. The chromists are quantitatively the most important eukaryotic contributors to primary production in marine ecosystems. The mechanisms of protein import across their four plastid membranes are still poorly understood. Components of an endoplasmic reticulum-associated degradation (ERAD) machinery in cryptophytes, partially encoded by the reduced genome of the secondary symbiont (the nucleomorph), are implicated in protein transport across the second outermost plastid membrane. Here, we show that the haptophyte Emiliania huxleyi, like cryptophytes, stramenopiles, and apicomplexans, possesses a nuclear-encoded symbiont-specific ERAD machinery (SELMA, symbiont-specific ERAD-like machinery) in addition to the host ERAD system, with targeting signals that are able to direct green fluorescent protein or yellow fluorescent protein to the predicted cellular localization in transformed cells of the stramenopile Phaeodactylum tricornutum. Phylogenies of the duplicated ERAD factors reveal that all SELMA components trace back to a red algal origin. In contrast, the host copies of cryptophytes and haptophytes associate with the green lineage to the exclusion of stramenopiles and alveolates. Although all chromalveolates with four membrane-bound plastids possess the SELMA system, this has apparently not arisen in a single endosymbiotic event. Thus, our data do not support the chromalveolate hypothesis.

• The physical and functional borders of transit peptide-like sequences in secondary endosymbionts.

  Authors: Gregor Felsner, Maik S Sommer, Uwe G Maier

  BMC plant biology. 10/2010; 10:223.

  Plastids rely on protein supply by their host cells. In plastids surrounded by two membranes (primary plastids) targeting of these proteins is facilitated by an N-terminal targeting signal, thePlastids rely on protein supply by their host cells. In plastids surrounded by two membranes (primary plastids) targeting of these proteins is facilitated by an N-terminal targeting signal, the transit peptide. In secondary plastids (surrounded by three or four membranes), transit peptide-like regions are an essential part of a bipartite topogenic signal sequence (BTS), and generally found adjacent to a N-terminally located signal peptide of the plastid pre-proteins. As in primary plastids, for which no wealth of functional information about transit peptide features exists, the transit peptide-like regions used for import into secondary ones show some common features only, which are also poorly characterized. We modified the BTS (in the transit peptide-like region) of the plastid precursor fucoxanthin-chlorophyll a/c binding protein D (FcpD) fused to GFP as model substrate for the characterization of pre-protein import into the secondary plastids of diatoms. Thereby we show that (i) pre-protein import is highly charge dependent. Positive net charge is necessary for transport across the plastid envelope, but not across the periplastid membrane. Acidic net charge perturbs pre-protein import within the ER. Moreover, we show that (ii) the mature domain of the pre-protein can provide intrinsic transit peptide functions. Our results indicate important characteristics of targeting signals of proteins imported into secondary plastids surrounded by four membranes. In addition, we show a self-targeting mechanism, in which the mature protein domain contributes to the transit peptide function. Thus, this phenomenon lowers the demand for pre-sequences evolved during the course of endosymbiosis.

• New mechanistic insights into pre-protein transport across the second outermost plastid membrane of diatoms.

  Authors: Franziska Hempel, Gregor Felsner, Uwe G Maier

  Molecular microbiology. 03/2010; 76(3):793-801.

  Chromalveolates like the diatom Phaeodactylum tricornutum arose through the uptake of a red alga by a phagotrophic protist, a process termed secondary endosymbiosis. In consequence, the plastids areChromalveolates like the diatom Phaeodactylum tricornutum arose through the uptake of a red alga by a phagotrophic protist, a process termed secondary endosymbiosis. In consequence, the plastids are surrounded by two additional membranes compared with primary plastids. This plastid morphology poses additional barriers for plastid-destined proteins, which are mostly nucleus-encoded. Recent investigations have focused on the postulated translocon of the second outermost membrane (periplastidal membrane, PPM). These studies identified a symbiont-specific ERAD (endoplasmic reticulum-associated degradation)-like machinery (SELMA), which has been implicated in plastid pre-protein import. Despite this recent progress, key factors for protein transport via SELMA are still unknown. As SELMA substrates presumably undergo ubiquitination, a corresponding ubiquitin ligase and an enzyme for the subsequent removal of ubiquitin need to reside in the space between the second and third membrane (periplastidal compartment, PPC). Here we characterize two proteins fulfilling these criteria. We show that ptE3P (P.t ricornutumE3 enzyme of the PPC), the ubiquitin ligase, and ptDUP (P.t ricornutumde-ubiquitinating enzyme of the PPC), the de-ubiquitinase, localize to the PPM and PPC, respectively. In addition, we demonstrate their retained functionality by in vitro data.

• Filling the gap, evolutionarily conserved Omp85 in plastids of chromalveolates.

  Authors: Lars Bullmann, Raimund Haarmann, Oliver Mirus, Rolf Bredemeier, Franziska Hempel, Uwe G Maier, Enrico Schleiff

  The Journal of biological chemistry. 02/2010; 285(9):6848-56.

  Chromalveolates are a diverse group of protists that include many ecologically and medically relevant organisms such as diatoms and apicomplexan parasites. They possess plastids generally surroundedChromalveolates are a diverse group of protists that include many ecologically and medically relevant organisms such as diatoms and apicomplexan parasites. They possess plastids generally surrounded by four membranes, which evolved by engulfment of a red alga. Today, most plastid proteins must be imported, but many aspects of protein import into complex plastids are still cryptic. In particular, how proteins cross the third outermost membrane has remained unexplained. We identified a protein in the third outermost membrane of the diatom Phaeodactylum tricornutum with properties comparable to those of the Omp85 family. We demonstrate that the targeting route of P. tricornutum Omp85 parallels that of the translocation channel of the outer envelope membrane of chloroplasts, Toc75. In addition, the electrophysiological properties are similar to those of the Omp85 proteins involved in protein translocation. This supports the hypothesis that P. tricornutum Omp85 is involved in precursor protein translocation, which would close a gap in the fundamental understanding of the evolutionary origin and function of protein import in secondary plastids.

• An unusual ERAD-like complex is targeted to the apicoplast of Plasmodium falciparum.

  Authors: Simone Spork, Jan A Hiss, Katharina Mandel, Maik Sommer, Taco W A Kooij, Trang Chu, Gisbert Schneider, Uwe-G Maier, Jude M Przyborski

  Eukaryotic cell. 07/2009;

  Many apicomplexan parasites, including Plasmodium falciparum, harbour a so-called apicoplast; a complex plastid of red algal origin which was gained by a secondary endosymbiotic event. The exactMany apicomplexan parasites, including Plasmodium falciparum, harbour a so-called apicoplast; a complex plastid of red algal origin which was gained by a secondary endosymbiotic event. The exact molecular mechanisms directing the transport of nuclear encoded proteins to the apicoplast of P. falciparum are not well understood. Recently, in silico analyses revealed a second copy of proteins homologuous to components of the ER-associated protein degradation (ERAD) system in organisms with secondary plastids, including the malaria parasite P. falciparum. These proteins are predicted to be endowed with an apicoplast targeting signal, and are suggested to play a role in the transport of nuclear encoded proteins to the apicoplast. Here we have studied components of this ERAD-derived putative pre-protein translocon complex in malaria parasites. Using transfection technology coupled with fluorescence imaging techniques we can demonstrate that the n-terminus of several ERAD-derived components targets GFP to the apicoplast. Furthermore, we confirm that full-length PfsDer1-1 and PfsUba1 (homologues of yeast ERAD components) localise to the apicoplast, where PfsDer1-1 tightly associates with membranes. Constantly, PfhDer1-1 localises to the endoplasmic reticulum. Our data suggests that ERAD components have been "re-wired" to provide a conduit for protein transport to the apicoplast. Our results are discussed in relation to the nature of the apicoplast protein transport machinery.

• Genomic footprints of a cryptic plastid endosymbiosis in diatoms.

  Authors: Ahmed Moustafa, Bánk Beszteri, Uwe-G Maier, Chris Bowler, Klaus Valentin, Debashish Bhattacharya

  Science (New York, N.Y.). 06/2009; 324(5935):1724-6.

  Diatoms and other chromalveolates are among the dominant phytoplankters in the world's oceans. Endosymbiosis was essential to the success of chromalveolates, and it appears that the ancestral plastidDiatoms and other chromalveolates are among the dominant phytoplankters in the world's oceans. Endosymbiosis was essential to the success of chromalveolates, and it appears that the ancestral plastid in this group had a red algal origin via an ancient secondary endosymbiosis. However, recent analyses have turned up a handful of nuclear genes in chromalveolates that are of green algal derivation. Using a genome-wide approach to estimate the "green" contribution to diatoms, we identified >1700 green gene transfers, constituting 16% of the diatom nuclear coding potential. These genes were probably introduced into diatoms and other chromalveolates from a cryptic endosymbiont related to prasinophyte-like green algae. Chromalveolates appear to have recruited genes from the two major existing algal groups to forge a highly successful, species-rich protist lineage.

• ERAD-derived Pre-Protein Transport across the 2nd Outermost Plastid Membrane of Diatoms.

  Authors: Franziska Hempel, Lars Bullmann, Julia Lau, Stefan Zauner, Uwe-G Maier

  Molecular biology and evolution. 05/2009;

  The diatom Phaeodactylum tricornutum harbours a plastid that is surrounded by four membranes and evolved by way of secondary endosymbiosis. Like land plants, most of its plastid proteins are encodedThe diatom Phaeodactylum tricornutum harbours a plastid that is surrounded by four membranes and evolved by way of secondary endosymbiosis. Like land plants, most of its plastid proteins are encoded as pre-proteins on the nuclear genome of the host cell and are resultantly redirected into the organelle. Because two more membranes are present in diatoms than the one pair surrounding primary plastids, the targeting situation is obviously different and more complex. In this work, we focus on pre-protein transport across the second outermost plastid membrane - an issue that was experimentally inaccessible until now. We provide first indications that our hypothesis of an ERAD (ER-associated degradation)-derived pre-protein transport system might be correct. Our data demonstrate that the symbiont-specific Der1 proteins, sDer1-1 and sDer1-2, form an oligomeric complex within the second outermost membrane of the complex plastid. Moreover, we present first evidence that the complex interacts with transit peptides of pre-proteins being transported across this membrane into the periplastidal compartment, but not with transit peptides of stromal-targeted proteins. Thus, the sDer1-complex might have an additional role in discriminating pre-proteins that are transported across the two outermost membranes from pre-proteins directed across all four membranes of the complex plastid. Altogether, our studies of the symbiont-specific ERAD-like machinery of diatoms suggest that a pre-existing cellular machinery was recycled to fulfill a novel function during the transition of a former free-living eukaryote into a secondary endosymbiont.

• Diatoms in biotechnology: modern tools and applications.

  Authors: Andrew Bozarth, Uwe-G Maier, Stefan Zauner

  Applied microbiology and biotechnology. 01/2009;

  Diatoms have played a decisive role in the ecosystem for millions of years as one of the foremost set of oxygen synthesizers on earth and as one of the most important sources of biomass in oceans.Diatoms have played a decisive role in the ecosystem for millions of years as one of the foremost set of oxygen synthesizers on earth and as one of the most important sources of biomass in oceans. Previously, diatoms have been almost exclusively limited to academic research with little consideration of their practical uses beyond the most rudimentary of applications. Efforts have been made to establish them as decisively useful in such commercial and industrial applications as the carbon neutral synthesis of fuels, pharmaceuticals, health foods, biomolecules, materials relevant to nanotechnology, and bioremediators of contaminated water. Progress in the technologies of diatom molecular biology such as genome projects from model organisms, as well as culturing conditions and photobioreactor efficiency, may be able to be combined in the near future to make diatoms a lucrative source of novel substances with widespread relevance.

• Ancient Recruitment by Chromists of Green Algal Genes Encoding Enzymes for Carotenoid Biosynthesis.

  Authors: Ruth Frommolt, Sonja Werner, Harald Paulsen, Reimund Goss, Christian Wilhelm, Stefan Zauner, Uwe-G Maier, Arthur R Grossman, Debashish Bhattacharya, Martin Lohr

  Molecular biology and evolution. 10/2008;

  Chromist algae (stramenopiles, cryptophytes, and haptophytes) are major contributors to marine primary productivity. These eukaryotes acquired their plastid via secondary endosymbiosis, whereby anChromist algae (stramenopiles, cryptophytes, and haptophytes) are major contributors to marine primary productivity. These eukaryotes acquired their plastid via secondary endosymbiosis, whereby an early-diverging red alga was engulfed by a protist and the plastid was retained and its associated nuclear-encoded genes were transferred to the host genome. Current data suggest, however, that chromists are paraphyletic; therefore it remains unclear whether their plastids trace back to a single secondary endosymbiosis or alternatively, this organelle has resulted from multiple independent events in the different chromist lineages. Both scenarios, however, predict that plastid-targeted, nucleus-encoded chromist proteins should be most closely related to their red algal homologs. Here we analyzed the biosynthetic pathway of carotenoids that are essential components of all photosynthetic eukaryotes and find a mosaic evolutionary origin of these enzymes in chromists. Surprisingly, about one-third (5/16) of the proteins are most closely related to green algal homologs with three branching within or sister to the early diverging Prasinophyceae. This phylogenetic association is corroborated by shared diagnostic indels and the syntenic arrangement of a specific gene pair involved in the photoprotective xanthophyll cycle. The combined data suggest that the prasinophyte genes may have been acquired before the ancient split of stramenopiles, haptophytes, cryptophytes, and putatively also dinoflagellates. The latter point is supported by the observed monophyly of alveolates and stramenopiles in most molecular trees. One possible explanation for our results is that the green genes are remnants of a cryptic endosymbiosis that occurred early in chromalveolate evolution; i.e., prior to the postulated split of stramenopiles, alveolates, haptophytes, and cryptophytes. The subsequent red algal capture would have led to the loss or replacement of most green genes via intracellular gene transfer from the new endosymbiont. We argue that the prasinophyte genes were retained because they enhance photosynthetic performance in chromalveolates, thus extending the niches available to these organisms. The alternate explanation of green gene origin via serial endosymbiotic or horizontal gene transfers is also plausible, but the latter would require the independent origins of the same five genes in some or all of the different chromalveolate lineages.

• The cyanobacterial endosymbiont of the unicellular algae Rhopalodia gibba shows reductive genome evolution.

  Authors: Christoph Kneip, Christine Voss, Peter J Lockhart, Uwe-G Maier

  BMC evolutionary biology. 02/2008; 8:30.

  BACKGROUND: Bacteria occur in facultative association and intracellular symbiosis with a diversity of eukaryotic hosts. Recently, we have helped to characterise an intracellular nitrogen fixingBACKGROUND: Bacteria occur in facultative association and intracellular symbiosis with a diversity of eukaryotic hosts. Recently, we have helped to characterise an intracellular nitrogen fixing bacterium, the so-called spheroid body, located within the diatom Rhopalodia gibba. Spheroid bodies are of cyanobacterial origin and exhibit features that suggest physiological adaptation to their intracellular life style. To investigate the genome modifications that have accompanied the process of endosymbiosis, here we compare gene structure, content and organisation in spheroid body and cyanobacterial genomes. RESULTS: Comparison of the spheroid body's genome sequence with corresponding regions of near free-living relatives indicates that multiple modifications have occurred in the endosymbiont's genome. These include localised changes that have led to elimination of some genes. This gene loss has been accompanied either by deletion of the respective DNA region or replacement with non-coding DNA that is AT rich in composition. In addition, genome modifications have led to the fusion and truncation of genes. We also report that in the spheroid body's genome there is an accumulation of deleterious mutations in genes for cell wall biosynthesis and processes controlled by transposases. Interestingly, the formation of pseudogenes in the spheroid body has occurred in the presence of intact, and presumably functional, recA and recF genes. This is in contrast to the situation in most investigated obligate intracellular bacterium-eukaryote symbioses, where at least either recA or recF has been eliminated. CONCLUSION: Our analyses suggest highly specific targeting/loss of individual genes during the process of genome reduction and establishment of a cyanobacterial endosymbiont inside a eukaryotic cell. Our findings confirm, at the genome level, earlier speculation on the obligate intracellular status of the spheroid body in Rhopalodia gibba. This association is the first example of an obligate cyanobacterial symbiosis involving nitrogen fixation for which genomic data are available. It represents a new model system to study molecular adaptations of genome evolution that accompany a switch from free-living to intracellular existence.

• Complementation of a phycocyanin-bilin lyase from Synechocystis sp. PCC 6803 with a nucleomorph-encoded open reading frame from the cryptophyte Guillardia theta.

  Authors: Kathrin Bolte, Oliver Kawach, Julia Prechtl, Nicole Gruenheit, Julius Nyalwidhe, Uwe-G Maier

  BMC plant biology. 02/2008; 8:56.

  BACKGROUND: Cryptophytes are highly compartmentalized organisms, expressing a secondary minimized eukaryotic genome in the nucleomorph and its surrounding remnant cytoplasm, in addition to the cellBACKGROUND: Cryptophytes are highly compartmentalized organisms, expressing a secondary minimized eukaryotic genome in the nucleomorph and its surrounding remnant cytoplasm, in addition to the cell nucleus, the mitochondrion and the plastid. Because the members of the nucleomorph-encoded proteome may contribute to essential cellular pathways, elucidating nucleomorph-encoded functions is of utmost interest. Unfortunately, cryptophytes are inaccessible for genetic transformations thus far. Therefore the functions of nucleomorph-encoded proteins must be elucidated indirectly by application of methods in genetically accessible organisms. RESULTS: Orf222, one of the uncharacterized nucleomorph-specific open reading frames of the cryptophyte Guillardia theta, shows homology to slr1649 of Synechocystis sp. PCC 6803. Recently a further homolog from Synechococcus sp. PCC 7002 was characterized to encode a phycocyanin-beta155-bilin lyase. Here we show by insertion mutagenesis that the Synechocystis sp. PCC 6803 slr1649-encoded protein also acts as a bilin lyase, and additionally contributes to linker attachment and/or stability of phycobilisomes. Finally, our results indicate that the phycocyanin-beta155-bilin lyase of Synechocystis sp. PCC 6803 can be complemented in vivo by the nucleomorph-encoded open reading frame orf222. CONCLUSION: Our data show that the loss of phycocyanin-lyase function causes pleiotropic effects in Synechocystis sp. PCC 6803 and indicate that after separating from a common ancestor protein, the phycoerythrin lyase from Guillardia theta has retained its capacity to couple a bilin group to other phycobiliproteins. This is a further, unexpected example of the universality of phycobiliprotein lyases.

• Phospho-mimicry mutant of atToc33 affects early development of Arabidopsis thaliana.

  Authors: Mislav Oreb, Mikael Zoryan, Aleksandar Vojta, Uwe-G Maier, Lutz A Eichacker, Enrico Schleiff

  FEBS letters. 01/2008; 581(30):5945-51.

  The precursor protein receptor at the chloroplast outer membrane atToc33 is a GTPase, which can be inactivated by phosphorylation in vitro, being arrested in the GDP loaded state. To assess theThe precursor protein receptor at the chloroplast outer membrane atToc33 is a GTPase, which can be inactivated by phosphorylation in vitro, being arrested in the GDP loaded state. To assess the physiological function of phosphorylation, attoc33 knock out mutants were complemented with a mutated construct mimicking the constitutively phosphorylated state. Our data suggest that the reduced functionality of the mutant protein can be compensated by its upregulation. Chloroplast biogenesis and photosynthetic activity are impaired in the mutants during the early developmental stage, which is consistent with the requirement of atToc33 in young photosynthetic tissues.

• Translocation of a phycoerythrin alpha subunit across five biological membranes.

  Authors: Sven B Gould, Enguo Fan, Franziska Hempel, Uwe-G Maier, Ralf Bernd Klösgen

  The Journal of biological chemistry. 11/2007; 282(41):30295-302.

  Cryptophytes, unicellular algae, evolved by secondary endosymbiosis and contain plastids surrounded by four membranes. In contrast to cyanobacteria and red algae, their phycobiliproteins do notCryptophytes, unicellular algae, evolved by secondary endosymbiosis and contain plastids surrounded by four membranes. In contrast to cyanobacteria and red algae, their phycobiliproteins do not assemble into phycobilisomes and are located within the thylakoid lumen instead of the stroma. We identified two gene families encoding phycoerythrin alpha and light-harvesting complex proteins from an expressed sequence tag library of the cryptophyte Guillardia theta. The proteins bear a bipartite topogenic signal responsible for the transport of nuclear encoded proteins via the ER into the plastid. Analysis of the phycoerythrin alpha sequences revealed that more than half of them carry an additional, third topogenic signal comprising a twin arginine motif, which is indicative of Tat (twin arginine transport)-specific targeting signals. We performed import studies with several derivatives of one member using a diatom transformation system, as well as intact chloroplasts and thylakoid vesicles isolated from pea. We demonstrated the different targeting properties of each individual part of the tripartite leader and show that phycoerythrin alpha is transported across the thylakoid membrane into the thylakoid lumen and protease-protected. Furthermore, we showed that thylakoid transport of phycoerythrin alpha takes place by the Tat pathway even if the 36 amino acid long bipartite topogenic signal precedes the actual twin arginine signal. This is the first experimental evidence of a protein being targeted across five biological membranes.

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