Jörn Wolters

The troublesome parasites : molecular and morphological evidence that apicomplexa belong to the dinoflagellate-ciliate clade

Large insertions and deletions in the variable regions of eukaryotic 16S-like rRNA relative to the archaebacterial structure have been defined as a marker for rapidly evolving taxa. Deletions in the rRNA occur in the diplomonad Giardia and the microsporidian Vairimorpha, whereas insertions occur in Euglenozoa (Euglena and the kinetoplastids), Acanthamoeba, Naegleria, Physarum, Dictyostelium, the apicomplexan Plasmodium, the ciliate Euplotes, and some metazoa. Except Acanthamoeba and Euplotes, all of these protists were previously placed at the base of the eukaryote phylogeny. A re-analysis of the 16S-like rRNA and 5S rRNA data with the neighborliness method revealed a close relationship of Apicomplexa to the dinoflagellate-ciliate clade, most probably closer to the dinoflagellates. Morphological evidence that supports this grouping is the layer of sacs underneath the plasma membrane in all three taxa and the identical structure of trichocysts in the apicomplexan Spiromonas and dinoflagellates. The remaining rapidly evolving organisms might still be misplaced in the 16S-like rRNA trees.

Demonstration of nucleomorph-encoded eukaryotic small subunit ribosomal RNA in cryptomonads.

SummaryIn cryptomonads, unicellular phototrophic flagellates, the plastid(s) is (are) located in a special narrow compartment which is bordered by two membranes; it harbours neither mitochondria nor Golgi dictyosomes but comprises eukaryotic ribosomes and starch grains together with a small organelle called the nucleomorph. The nucleomorph contains DNA and is surrounded by a double membrane with pores. It is thought to be the vestigial nucleus of a phototrophic eukaryotic endosymbiont. Cryptomonads are therefore supposed to represent an intermediate state in the evolution of complex plastids from endosymbionts. We have succeeded in isolating pure nucleomorph fractions, and can thus provide, using pulsed field gel electrophoresis, polymerase chain reaction and sequence analysis, definitive proof for the eukaryotic nature of the symbiont and its phylogenetic origin.

Primary and secondary structure of the nuclear small subunit ribosomal RNA of the cryptomonadPyrenomonas salina as inferred from the gene sequence: Evolutionary implications

SummaryThe cryptomonadPyrenomonas salina presumably has arisen from a symbiotic event involving a flagellated phagotrophic host cell and a photosynthetic eukaryote as the symbiont. Correspondingly, in this unicellular alga there are four different genomes, e.g., the nuclear and the mitochondrial genomes of the host cell as well as the plastid genome and the genome contained in the vestigial nucleus of the endocytobiont (nucleomorph). To analyze the orgin of one of the symbiotic partners the small subunit rRNA gene sequence of the host cell nucleus was determined, and a secondary structure model has been constructed. This sequence is compared to those of 40 other eukaryotes. A phylogenetic tree constructed using the neighborliness method revealed a close relationship between the host cell ofP. salina and the chlorophytes, whereas the rhodophytes diverge more deeply in the tree.

Plastid DNA from Pyrenomonas salina (cryptophyceae) : physical map, genes,and evolutionary implications

SummaryCryptomonads are thought to have arisen from a symbiotic association between a eukaryotic flagellated host and a eukaryotic algal symbiont, presumably related to red algae. As organellar DNAs have proven to be useful tools in elucidating phylogenetic relationships, the plastid (pt) DNA of the cryptomonad alga Pyrenomonas salina has been characterized in some detail. A restriction map of the circular 127 kb ptDNA from Pyrenomonas salina was established. An inverted repeat (IR) region of about 5 kb separates two single-copy regions of 15 and 102 kb, respectively. It contains the genes for the small and large subunit of rRNA. Ten protein genes, coding for the large subunit of ribulose-1,5-bisphosphate carboxylase, the 47 kDa, 43 kDa and 32 kDa proteins of photosystem II, the ribosomal proteins L2, S7 and S11, the elongation factor Tu, as well as the α- and β-subunits of ATP synthase, have been localized on the restriction map either by hybridization of heterologous gene probes or by sequence homologies. The gene for the plastidal small subunit (SSUr) RNA has been sequenced and compared to homologous SSU regions from the cyanobacterium Anacystis nidulans and plastids from rhodophytes, chromophytes, euglenoids, chlorophytes, and land plants. A phylogenetic tree constructed with the neighborliness method and indicating a relationship of cryptomonad plastids with those of red algae is presented.

A comparison of leaf thionin sequences of barley cultivars and wild barley species.

SummaryLeaf thionins of several barley cultivars and wild barley species were analysed. We found large differences in the numbers of leaf thionin genes in different Hordeum species. While, for instance, cultivars of Hordeum vulgare (Section Hordeum) contain more than 50 copies of thionin genes per haploid genome, the numbers are much lower in Hordeum species belonging to the sections Critesion and Stenostachys. The apparent number of genes correlates with the concentration of leaf thionin and its mRNA, which differs more than 100-fold among various Hordeum species. Leaf thionins are synthesized as high molecular weight precursor proteins that contain a signal peptide domain, a thionin domain and an acidic polypeptide domain. Analysis of cDNA clones of leaf thionins revealed a family of related transcripts. When the predicted amino acid sequences of the precursor molecules of wild barley species were compared, differences in the sequence variability of the three domains became apparent. The frequency of amino acid exchanges is much higher within the thionin domain than in the signal peptide and acidic polypeptide domains. The amino acid exchanges within the thionin domain do not occur at random but are confined to variable regions that alternate with highly conserved areas. Conserved regions comprise mostly cysteine residues and adjacent amino acids and may be important for the correct formation of the specific disulphide configuration of thionins.