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Ohne das Eingreifen des Menschen wäre Mitteleuropa fast ein reines Waldgebiet. Noch heute beheimaten die Wälder eine große Vielfalt an Pflanzen und Tieren, die für diese Region spezifisch sind. Regionale Besonderheiten gehen aber verloren, je mehr Menschen in die Ökosysteme eingreifen: So unterscheiden sich die Pflanzenarten auf der North Charles Street in Baltimore nur wenig von denjenigen der Mainzer Landstraße in Frankfurt. Gleichzeitig verdrängen zugewanderte und eingeschleppte Arten heimische Tiere und Pflanzen. Allerdings gibt es auch im Frankfurter Stadtgebiet echte Horte der Biodiversität.
We performed a bioinformatical analysis of protein export elements (PEXEL) in the putative proteome of the malaria parasite Plasmodium falciparum. A protein family-specific conservation of physicochemical residue profiles was found for PEXEL-flanking sequence regions. We demonstrate that the family members can be clustered based on the flanking regions only and display characteristic hydrophobicity patterns. This raises the possibility that the flanking regions may contain additional information for a family-specific role of PEXEL. We further show that signal peptide cleavage results in a positional alignment of PEXEL from both proteins with, and without, a signal peptide.
Ribosome biogenesis in eukaryotes requires the participation of a large number of ribosome assembly factors. The highly conserved eukaryotic nucleolar protein Nep1 has an essential but unknown function in 18S rRNA processing and ribosome biogenesis. In Saccharomyces cerevisiae the malfunction of a temperature-sensitive Nep1 protein (nep1-1ts) was suppressed by the addition of S-adenosylmethionine (SAM). This suggests the participation of Nep1 in a methyltransferase reaction during ribosome biogenesis. In addition, yeast Nep1 binds to a 6-nt RNA-binding motif also found in 18S rRNA and facilitates the incorporation of ribosomal protein Rps19 during the formation of pre-ribosomes. Here, we present the X-ray structure of the Nep1 homolog from the archaebacterium Methanocaldococcus jannaschii in its free form (2.2 Å resolution) and bound to the S-adenosylmethionine analog S-adenosylhomocysteine (SAH, 2.15 Å resolution) and the antibiotic and general methyltransferase inhibitor sinefungin (2.25 Å resolution). The structure reveals a fold which is very similar to the conserved core fold of the SPOUT-class methyltransferases but contains a novel extension of this common core fold. SAH and sinefungin bind to Nep1 at a preformed binding site that is topologically equivalent to the cofactor-binding site in other SPOUT-class methyltransferases. Therefore, our structures together with previous genetic data suggest that Nep1 is a genuine rRNA methyltransferase.
Background Drought is the major constraint to increase yield in chickpea (Cicer arietinum). Improving drought tolerance is therefore of outmost importance for breeding. However, the complexity of the trait allowed only marginal progress. A solution to the current stagnation is expected from innovative molecular tools such as transcriptome analyses providing insight into stress-related gene activity, which combined with molecular markers and expression (e)QTL mapping, may accelerate knowledge-based breeding. SuperSAGE, an improved version of the serial analysis of gene expression (SAGE) technique, generating genome-wide, high-quality transcription profiles from any eukaryote, has been employed in the present study. The method produces 26 bp long fragments (26 bp tags) from defined positions in cDNAs, providing sufficient sequence information to unambiguously characterize the mRNAs. Further, SuperSAGE tags may be immediately used to produce microarrays and probes for real-time-PCR, thereby overcoming the lack of genomic tools in non-model organisms. Results We applied SuperSAGE to the analysis of gene expression in chickpea roots in response to drought. To this end, we sequenced 80,238 26 bp tags representing 17,493 unique transcripts (UniTags) from drought-stressed and non-stressed control roots. A total of 7,532 (43%) UniTags were more than 2.7-fold differentially expressed, and 880 (5.0%) were regulated more than 8-fold upon stress. Their large size enabled the unambiguous annotation of 3,858 (22%) UniTags to genes or proteins in public data bases and thus to stress-response processes. We designed a microarray carrying 3,000 of these 26 bp tags. The chip data confirmed 79% of the tag-based results, whereas RT-PCR confirmed the SuperSAGE data in all cases. Conclusion This study represents the most comprehensive analysis of the drought-response transcriptome of chickpea available to date. It demonstrates that – inter alias – signal transduction, transcription regulation, osmolyte accumulation, and ROS scavenging undergo strong transcriptional remodelling in chickpea roots already 6 h after drought stress. Certain transcript isoforms characterizing these processes are potential targets for breeding for drought tolerance. We demonstrate that these can be easily accessed by micro-arrays and RT-PCR assays readily produced downstream of SuperSAGE. Our study proves that SuperSAGE owns potential for molecular breeding also in non-model crops.
Background One of the central issues in ecology is the question what allows sympatric occurrence of closely related species in the same general area? The non-biting midges Chironomus riparius and C. piger, interbreeding in the laboratory, have been shown to coexist frequently despite of their close relatedness, similar ecology and high morphological similarity. Methodology/Principal Findings In order to investigate factors shaping niche partitioning of these cryptic sister species, we explored the actual degree of reproductive isolation in the field. Congruent results from nuclear microsatellite and mitochondrial haplotype analyses indicated complete absence of interspecific gene-flow. Autocorrelation analysis showed a non-random spatial distribution of the two species. Though not dispersal limited at the scale of the study area, the sister species occurred less often than expected at the same site, indicating past or present competition. Correlation and multiple regression analyses suggested the repartition of the available habitat along water chemistry gradients (nitrite, conductivity, CaCO3), ultimately governed by differences in summer precipitation regime. Conclusions We show that these morphologically cryptic sister species partition their niches due to a certain degree of ecological distinctness and total reproductive isolation in the field. The coexistence of these species provides a suitable model system for the investigation of factors shaping the distribution of closely related, cryptic species.
The moderate halophile Halobacillus halophilus is the paradigm for chloride dependent growth in prokaryotes. Recent experiments shed light on the molecular basis of the chloride dependence that is reviewed here. In the presence of moderate salinities Halobacillus halophilus mainly accumulates glutamine and glutamate to adjust turgor. The transcription of glnA2 (encoding a glutamine synthetase) as well as the glutamine synthetase activity were identified as chloride dependent steps. Halobacillus halophilus switches its osmolyte strategy and produces proline as the main compatible solute at high salinities. Furthermore, Halobacillus halophilus also shifts its osmolyte strategy at the transition from the exponential to the stationary phase where proline is exchanged by ectoine. Glutamate was found as a second messenger" essential for proline production. This observation leads to a new model of sensing salinity by sensing the physico-chemical properties of different anions.
Quantitative analysis of snoRNA association with pre-ribosomes and release of snR30 by Rok1 helicase
(2008)
In yeast, three small nucleolar RNAs (snoRNAs) are essential for the processing of pre-ribosomal RNA—U3, U14 and snR30—whereas 72 non-essential snoRNAs direct site-specific modification of pre-rRNA. We applied a quantitative screen for alterations in the pre-ribosome association to all 75 yeast snoRNAs in strains depleted of eight putative helicases implicated in 40S subunit synthesis. For the modification-guide snoRNAs, we found no clear evidence for the involvement of these helicases in the association or dissociation of pre-ribosomes. However, the DEAD box helicase Rok1 was required specifically for the release of snR30. Point mutations in motif I, but not in motif III, of the helicase domain of Rok1 impaired the release of snR30, but this was less marked than in strains depleted of Rok1, and resulted in a dominant-negative growth phenotype. Dissociation of U3 and U14 from pre-ribosomes is also dependent on helicases, suggesting that release of the essential snoRNAs might differ mechanistically from release of the modification-guide snoRNAs. Keywords: ribosome biogenesis; RNA helicase; snoRNA
Exported proteases of Helicobacter pylori (H. pylori) are potentially involved in pathogen-associated disorders leading to gastric inflammation and neoplasia. By comprehensive sequence screening of the H. pylori proteome for predicted secreted proteases, we retrieved several candidate genes. We detected caseinolytic activities of several such proteases, which are released independently from the H. pylori type IV secretion system encoded by the cag pathogenicity island (cagPAI). Among these, we found the predicted serine protease HtrA (Hp1019), which was previously identified in the bacterial secretome of H. pylori. Importantly, we further found that the H. pylori genes hp1018 and hp1019 represent a single gene likely coding for an exported protein. Here, we directly verified proteolytic activity of HtrA in vitro and identified the HtrA protease in zymograms by mass spectrometry. Overexpressed and purified HtrA exhibited pronounced proteolytic activity, which is inactivated after mutation of Ser205 to alanine in the predicted active center of HtrA. These data demonstrate that H. pylori secretes HtrA as an active protease, which might represent a novel candidate target for therapeutic intervention strategies.
Chloroplast function depends on the translocation of cytosolically synthesized precursor proteins into the organelle. The recognition and transfer of most precursor proteins across the outer membrane depend on a membrane inserted complex. Two receptor components of this complex, Toc34 and Toc159, are GTPases, which can be phosphorylated by kinases present in the hosting membrane. However, the physiological function of phosphorylation is not yet understood in detail. It is demonstrated that both receptors are phosphorylated within their G-domains. In vitro, the phosphorylation of Toc34 disrupts both homo- and heterodimerization of the G-domains as determined using a phospho-mimicking mutant. In endogenous membranes this mutation or phosphorylation of the wild-type receptor disturbs the association of Toc34, but not of Toc159 with the translocation pore. Therefore, phosphorylation serves as an inhibitor for the association of Toc34 with other components of the complex and phosphorylation can now be discussed as a mechanism to exchange different isoforms of Toc34 within this ensemble.