Biomedical subjects

Guntram Christiansen

Publications and source records attributed to Guntram Christiansen.

9 recordsLinked to original sources

PubMed2006

Transposons inactivate biosynthesis of the nonribosomal peptide microcystin in naturally occurring Planktothrix spp.

The filamentous cyanobacteria Planktothrix spp. occur in the temperate region of the Northern hemisphere. The red-pigmented Planktothrix rubescens bacteria occur in deep, physically stratified, and less eutrophic lakes. Planktothrix is a known producer of the toxic heptapeptide microcystin (MC), which is produced nonribosomally by a large enzyme complex consisting of peptide synthetases and polyketide synthases encoded by a total of nine genes (mcy genes). Planktothrix spp. differ in their cellular MC contents as well as the production of MC variants; however, the mechanisms favoring this diversity are not understood. Recently, the occurrence of Planktothrix strains containing all mcy genes but lacking MC has been reported. In this study, 29 such strains were analyzed to find out if mutations of the mcy genes lead to the inability to synthesize MC. Two deletions, spanning 400 bp (in mcyB; one strain) and 1,869 bp (in mcyHA; three strains), and three insertions (IS), spanning 1,429 bp (in mcyD; eight strains), 1,433 bp (in mcyEG; one strain), and 1,433 bp (in mcyA; one strain), were identified. Though found in different genes and different isolates and transcribed in opposite directions, IS were found to be identical and contained conserved domains assigned to transposable elements. Using mutation-specific primers, two insertions (in mcyD and mcyA) and one deletion (in mcyHA) were found regularly in populations of P. rubescens in different lakes. The results demonstrate for the first time that different mutations resulting in inactivation of MC synthesis do occur frequently and make up a stable proportion of the mcy gene pool in Planktothrix populations over several years.

Bacterial Proteins↗

PubMed2006

Natural products from cyanobacteria: Exploiting a new source for drug discovery.

In the 1990s, the pharmaceutical industry shifted its focus to a combinatorial chemistry approach to fill drug-discovery pipelines; however, more recently there has been renewed interest in natural products as sources of lead compounds. Cyanobacteria are prolific producers of natural products displaying enormous chemical diversity, yet, until recently, exploitation of the genera was hampered by a number of issues related to their handling. With most of these problems now resolved, cyanobacteria have the potential to expand the variety of natural products obtained from microorganisms. The relative disregard in the past of cyanobacteria compared with other microbial sources of natural products, as well as the huge chemical diversity and biological activities of their products, recommend them as an attractive source of novel drugs for use in diverse therapeutic areas.

Biological Products↗

PubMed2005

Genetic identification of microcystin ecotypes in toxic cyanobacteria of the genus Planktothrix.

Microcystins (MCs) are toxic heptapeptides which are synthesized by the filamentous cyanobacterium Planktothrix and other genera via non-ribosomal peptide synthesis. MCs share the common structure cyclo(-D-ala1-L-X2-D-erythro-beta-iso-aspartic acid3-L-Z4-adda5-D-Glu6-N-methyl-dehydroalanine7) [Adda; (2S, 3S, 8S, 9S)-3-amino-9-methoxy-2,6,8-trimethyl-10-phenyldeca-4,6-dienoic acid], in which numerous MC variants have been reported. In general, the variation in structure is due to different amino acid residues in positions 7, 2 and 4 within the MC molecule, which are thought to be activated by the adenylation domains mcyAAd1, mcyBAd1 and mcyCAd, respectively. It was the aim of the study (i) to identify MC ecotypes that differed in the production of specific MC variants and (ii) to correlate the genetic variation within adenylation domains with the observed MC variants among 17 Planktothrix strains. Comparison of the sequences of mcyAAd1 revealed two distinctive Ad-genotypes differing in base pair composition and the insertion of an N-methyl transferase (NMT) domain. The mcyAAd1 genotype with NMT (2854 bp) correlated with N-methyl-dehydroalanine and the mcyAAd1 genotype without NMT (1692 bp) correlated with dehydrobutyrine in position 7. Within mcyBAd1, a lower genetic variation (0-4 %) and an exclusive correlation between one Ad-genotype and homotyrosine as well as another Ad-genotype and arginine in position 2 was found. The sequences of mcyCAd were found to be highly similar (0-1 % dissimilarity) and all strains contained arginine in position 4. The results on adenylation domain polymorphism do provide insights into the evolutionary origin of adenylation domains in Planktothrix and may be combined with ecological research in order to provide clues about the abundance of genetically defined MC ecotypes in nature.

Adenosine Monophosphate↗

PubMed2005

The microcystin composition of the cyanobacterium Planktothrix agardhii changes toward a more toxic variant with increasing light intensity.

The cyanobacterium Planktothrix agardhii, which is dominant in many shallow eutrophic lakes, can produce hepatotoxic microcystins. Currently, more than 70 different microcystin variants have been described, which differ in toxicity. In this study, the effect of photon irradiance on the production of different microcystin variants by P. agardhii was investigated using light-limited turbidostats. Both the amount of the mRNA transcript of the mcyA gene and the total microcystin production rate increased with photon irradiance up to 60 micromol m(-2) s(-1), but they started to decrease with irradiance greater than 100 micromol m(-2) s(-1). The cellular content of total microcystin remained constant, independent of the irradiance. However, of the two main microcystin variants detected in P. agardhii, the microcystin-DeRR content decreased twofold with increased photon irradiance, whereas the microcystin-DeLR content increased threefold. Since microcystin-DeLR is considerably more toxic than microcystin-DeRR, this implies that P. agardhii becomes more toxic at high light intensities.

Bacterial Proteins↗

PubMed2004

Diversity of coexisting Planktothrix (cyanobacteria) chemotypes deduced by mass spectral analysis of microystins and other oligopeptides.

Cyanobacteria are reported to produce secondary metabolites of which toxic and bioactive peptides are of scientific and public interest. Many peptides are synthesized by the non-ribosomal peptide synthesis pathway and their presence is a stable feature of individual clones. We isolated 18 clonal strains of Planktothrix from a single water sample from lake Maxsee near Berlin and analyzed them by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, HPLC, and PCR for their production of peptides and the presence of microcystin synthetase genes. Microcystins could be detected in seven of the strains with considerable variability of contents and numbers of structural variants. Other known peptides like anabaenopeptins B and E/F, microviridin I, and prenylagaramide B and new variants of known peptide classes like aeruginosins and cyanopeptolins were detected in some strains while lacking in others. The 18 strains represented 15 chemotypes with respect to their peptide patterns. In contrast, all strains were morphologically very similar with respect to cell dimensions and pigmentation. Given the diversity of chemotypes among the randomly selected isolates, an immense diversity of chemotypes in the entire population can be assumed.

Bacterial Proteins↗

PubMed2004

Abundance of active and inactive microcystin genotypes in populations of the toxic cyanobacterium Planktothrix spp.

To investigate the abundance of active and inactive microcystin genotypes in populations of the filamentous cyanobacterium Planktothrix spp., individual filaments were grown as clonal strains in the laboratory and analysed for microcystin synthetase (mcy) genes and microcystin. Twenty-three green-pigmented strains of P. agardhii originating mostly from shallow water bodies fell into two groups, those possessing mcyA and those lacking mcyA. In contrast, all of the 49 strains that were assigned to the red-pigmented P. rubescens contained mcyA. One strain of P. agardhii and eight strains of P. rubescens contained the total microcystin synthetase gene cluster but were found inactive in microcystin synthesis. To investigate the natural abundance of inactive mcy genotypes in P. rubescens individual filaments sampled from Lake Irrsee and Lake Mondsee (Austria) were analysed directly for the presence of mcyA and microcystin by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry. All filaments assigned to P. rubescens contained mcyA. The proportion of inactive microcystin genotypes in populations with a low (Irrsee) or high density (Mondsee) of P. rubescens was 5% and 21%, each. The results of this study demonstrate that P. rubescens typically contain mcy genes whereas P. agardhii have a patchy distribution of mcy genes. In both species microcystin producers co-occur with non-microcystin producers due to the absence/inactivation of mcy genes.

Bacterial Proteins↗

PubMed2003

PCR-based identification of microcystin-producing genotypes of different cyanobacterial genera.

Microcystins are harmful hepatotoxins produced by many, but not all strains of the cyanobacterial genera Anabaena, Microcystis, Anabaena, Planktothrix, and Nostoc. Waterbodies have to be monitored for the mass development of toxic cyanobacteria; however, because of the close genetic relationship of microcystin-producing and non-producing strains within a genus, identification of microcystin-producers by morphological criteria is not possible. The genomes of microcystin-producing cells contain mcy genes coding for the microcystin synthetase complex. Based on the sequence information of mcy genes from Microcystis and Planktothrix, a primer pair for PCR amplification of a mcyA gene fragment was designed. PCR with this primer pair is a powerful means to identify microcystin-producing strains of the genera Anabaena, Microcystis, and Planktothrix. Moreover, subsequent RFLP analysis of the PCR products generated genus-specific fragments and allowed the genus of the toxin producer to be identified. The assay can be used with DNA from field samples.

Anabaena↗

PubMed2003

The abundance of microcystin-producing genotypes correlates positively with colony size in Microcystis sp. and determines its microcystin net production in Lake Wannsee.

The working hypotheses tested on a natural population of Microcystis sp. in Lake Wannsee (Berlin, Germany) were that (i) the varying abundance of microcystin-producing genotypes versus non-microcystin-producing genotypes is a key factor for microcystin net production and (ii) the occurrence of a gene for microcystin net production is related to colony morphology, particularly colony size. To test these hypotheses, samples were fractionated by colony size with a sieving procedure during the summer of 2000. Each colony size class was analyzed for cell numbers, the proportion of microcystin-producing genotypes, and microcystin concentrations. The smallest size class of Microcystis colonies (<50 microm) showed the lowest proportion of microcystin-producing genotypes, the highest proportion of non-microcystin-producing cells, and the lowest microcystin cell quotas (sum of microcystins RR, YR, LR, and WR). In contrast, the larger size classes of Microcystis colonies (>100 microm) showed the highest proportion of microcystin-producing genotypes, the lowest proportion of non-microcystin-producing cells, and the highest microcystin cell quotas. The microcystin net production rate was nearly one to one positively related to the population growth rate for the larger colony size classes (>100 microm); however, no relationship could be found for the smaller size classes. It was concluded that the variations found in microcystin net production between colony size classes are chiefly due to differences in genotype composition and that the microcystin net production in the lake is mainly influenced by the abundance of the larger (>100- microm) microcystin-producing colonies.

Bacterial Proteins↗

PubMed2003

Microcystin biosynthesis in planktothrix: genes, evolution, and manipulation.

Microcystins represent an extraordinarily large family of cyclic heptapeptide toxins that are nonribosomally synthesized by various cyanobacteria. Microcystins specifically inhibit the eukaryotic protein phosphatases 1 and 2A. Their outstanding variability makes them particularly useful for studies on the evolution of structure-function relationships in peptide synthetases and their genes. Analyses of microcystin synthetase genes provide valuable clues for the potential and limits of combinatorial biosynthesis. We have sequenced and analyzed 55.6 kb of the potential microcystin synthetase gene (mcy) cluster from the filamentous cyanobacterium Planktothrix agardhii CYA 126. The cluster contains genes for peptide synthetases (mcyABC), polyketide synthases (PKSs; mcyD), chimeric enzymes composed of peptide synthetase and PKS modules (mcyEG), a putative thioesterase (mcyT), a putative ABC transporter (mcyH), and a putative peptide-modifying enzyme (mcyJ). The gene content and arrangement and the sequence of specific domains in the gene products differ from those of the mcy cluster in Microcystis, a unicellular cyanobacterium. The data suggest an evolution of mcy clusters from, rather than to, genes for nodularin (a related pentapeptide) biosynthesis. Our data do not support the idea of horizontal gene transfer of complete mcy gene clusters between the genera. We have established a protocol for stable genetic transformation of Planktothrix, a genus that is characterized by multicellular filaments exhibiting continuous motility. Targeted mutation of mcyJ revealed its function as a gene coding for a O-methyltransferase. The mutant cells produce a novel microcystin variant exhibiting reduced inhibitory activity toward protein phosphatases.

Amino Acid Sequence↗