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Protistology ■ 29
intact-looking to half-digested), but there are also some naked chloroplasts with three surrounding membranes in the ciliate cytoplasm. To identify the origin of these chloroplasts, we attempted to analyze PCR products obtained from isolated whole ciliate cells using various specific primers. The diatom-targeted primers yielded a probable nuclear SSU rDNA, and phylogenetic analyses based on Neighbor Joining showed that this sequence pertains to the centric diatom genus Discostella (Stephanodiscaceae). It was especially close to that of the diatom symbiont of the dinoflagellate Peridiniopsis penardii. We are now carrying out a detailed study of the ciliate and its symbiont from taxonomic and life history points of view.
EVOLUTION OF THE MITOCHONDRIAL PROTEOME, FROM THE LARGE-SCALE PATTERNS TO THE NITTY-GRITTY DETAILS Huynen M.A., Duarte I., Szklarczyk R., Elurbe D. CMBI, Radboud University Medical Centre, Nijme-gen, Netherlands huynen@cmbi.ru.nl
I will review what has been inferred about the changes at the level of the proteome that accompanied the evolution of the mitochondrion from an Alphaproteobacterium, quantifying the amount of gene loss, gene replacement and gene gain. Most of the loss and replacements that separate current day mitochondria from Alphaproteobacteria took place before the radiation ofthe eukaryotes. Detailed analyses of the evolution of the mitochondrial complexes like the ribosome and oxidative phos-phorylation show that also the acquisition of new proteins occurred mainly before the radiation of the eukaryotes, supporting an early acquisition of mitochondria in eukaryotic evolution. Secondly I will analyze the accumulation ofnew supernumerary subunits and assembly factors from pre-existing protein families for one protein complex in detail: complex I. There is a spectrum of protein function conservation between the complex I representatives and their non-complex I homologs. In general, the new complex I proteins appear to have been recruited from proteins that are active in mitochondria: proteins one expects to be expressed when and where complex I is active. Within complex I and its assembly there are many cases of neo-functionalization after gene duplication, one case of sub-functionalization, and one case in which a complex I protein itself appears to have been the source of the evolution of a new protein in another complex. Complex I and its assembly can therewith be regarded as a treasure trove for pathway evolution.
CONDITIONAL EXPRESSION SYSTEM IS NOT SUITABLE FOR DEVELOPMENTAL STUDIES IN LEISHMANIA Ishemgulova A.1, Kraeva N.1, Faktorova D.2, Podesvova L.1, Lukes J.234, Yurchenko V.125
1 - Life Science Research Centre, Faculty ofScience, University ofOstrava, 71000Ostrava, Czech Republic
2 - Czech Academy of Sciences, 370 05 Ceske Budejovice (Budweis), Czech Republic
3 - Faculty of Sciences, University ofSouth Bohemia, 370 05Cesku Budejovice (Budweis), Czech Republic
4 - Canadian Institute for Advanced Research, Toronto, ONM5G1Z8, Canada
5 - Department of Pathology, Albert Einstein College of Medicine, Bronx, NY, 10461, USA vyacheslav.yurchenko@osu.cz
The genus Leishmania unites parasitic protists of the family Trypanosomatidae causing leishmaniases, several closely related diseases that affect human and animal populations mainly in the tropical and subtropical regions. The clinical manifestations vary from spontaneously healing skin lesions to progressive and potentially fatal visceral infections. Leishmaniases represent a global health problem with over 350 million people at risk and an annual incidence rate of 2—10 million worldwide. Conventional and conditional systems allow for a controlled activation or repression of gene expression in time and space. Such systems are nowadays widely used to analyse a variety of cellular processes in numerous parasites including Leishmania. A T7-driven, tetracycline-inducible system for protein expression was established in a human pathogen Leishmania mexicana. The gene expression in this strain is tightly regulated and dose- and time dependent. We believe that it can be widely used by the parasitology community to analyse effects of genes of interest on biology, physiology and virulence of parasites causing cutaneous leishmaniases. This system was used to analyse gene expression profiles during L. mexicana differentiation (procyclics, metacyclics, and amastigotes). The transcription/translation of the gene of interest was severely decreased upon Leishmania differentiation into metacyclic and amastigotes. However, the same expression profile was documented for the T7 polymerase. The expression was demonstrated to be not locus-specific but dependent on untranslated regions flanking open reading frames of studied genes. We concluded that the previously established conventional gene expression systems might have certain limitations in their common applications.
