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Ludwig Eichinger

Publications and source records attributed to Ludwig Eichinger.

17 recordsLinked to original sources
PubMed2006

Filamin-regulated F-actin assembly is essential for morphogenesis and controls phototaxis in Dictyostelium.

Dictyostelium strains lacking the F-actin cross-linking protein filamin (ddFLN) have a severe phototaxis defect at the multicellular slug stage. Filamins are rod-shaped homodimers that cross-link the actin cytoskeleton into highly viscous, orthogonal networks. Each monomer chain of filamin is comprised of an F-actin-binding domain and a rod domain. In rescue experiments only intact filamin re-established correct phototaxis in filamin minus mutants, whereas C-terminally truncated filamin proteins that had lost the dimerization domain and molecules lacking internal repeats but retaining the dimerization domain did not rescue the phototaxis defect. Deletion of individual rod repeats also changed their subcellular localization, and mutant filamins in general were less enriched at the cell cortex as compared with the full-length protein and were increasingly present in the cytoplasm. For correct phototaxis ddFLN is only required at the tip of the slug because expression under control of the cell type-specific extracellular-matrix protein A (ecmA) promoter and mixing experiments with wild type cells supported phototactic orientation. Likewise, in chimeric slugs wild type cells were primarily found at the tip of the slug, which acts as an organizer in Dictyostelium morphogenesis.

Actins↗
PubMed2006

Contrasting evolution of expression differences in the testis between species and subspecies of the house mouse.

Regulatory changes in genes involved in reproduction are thought to be prime targets for divergence during speciation, since they are expected to play an important role in sexual selection and sexual conflict. We used microarray analysis of RNA from different wild populations of house mouse subspecies (including Mus m. musculus, Mus m. domesticus, and Mus m. castaneus) and from the sister species Mus spretus to test this assumption. A comparison of expression divergence in brain, liver/kidney, and testis shows a major difference in the evolutionary dynamics of testis-related genes. While the comparison between species confirms an excess in divergence in testis genes, we find that all comparisons between subspecies yield only a very small number of genes with significantly different expression levels in the testis. These results suggest that the early phase of the speciation process may not be driven by regulatory changes in genes that are potential targets of sexual selection, and that the divergence in these genes is only established during a later phase of the speciation process.

Animals↗
PubMed2006

Characterization of the Ste20-like kinase Krs1 of Dictyostelium discoideum.

Ste20-like kinases constitute a ubiquitous and expanding group of serine/threonine kinases, homologous to Ste20 in Saccharomyces cerevisiae. The social amoeba Dictyostelium discoideum contains at least 17 members of this kinase family, 13 from the germinal center kinase (GCK) subgroup and 4 p21-activated kinases (PAK). Here, we describe the kinase Krs1 which is encoded by the gene krsA, and phylogenetic analysis groups it into subfamily GCK-II together with human MST2 and MST1 or Hippo from Drosophila melanogaster. Significant similarities are found especially in the catalytic domain and in a short regulatory region (SARAH) which is thought to be important for protein/protein interactions. Northern blot analysis showed a single krsA transcript throughout development with an upregulation at 12h after the onset of starvation. The protein levels as detected with anti-Krs1 polyclonal antibodies revealed a similar pattern. Gel filtration experiments suggested that AX2 wild-type cells harbored multimeric forms of Krs1. In vitro phosphorylation assays with recombinant protein showed that the kinase exhibits autophosphorylation and accepts myelin basic protein and D. discoideum severin as substrates. A series of C-terminal deletions of Krs1 indicated that the regulatory domain in the C-terminal half contains inhibitory elements, and highlighted the importance of two predicted alpha-helices following subdomain XI of the classical catalytic domain. GFP-Krs1-overexpressing wild-type cells showed an enrichment of the kinase in the cortex, and motility of these cells during aggregation was reduced. Krs1 knockout strains exhibited only subtle differences to wild-type cells which suggests a certain redundancy among Ste20-like kinases in D. discoideum.

Amino Acid Sequence↗
PubMed2006

The Dictyostelium repertoire of seven transmembrane domain receptors.

The availability of fully sequenced genomes allows the in silico analysis of whole gene families in a given genome. A particularly large and interesting gene family is the G-protein-coupled receptor family. These receptors detect a variety of extracellular signals and transduce them, generally via heterotrimeric G-proteins, to effector proteins inside the cell and thus elicit a physiological response. G-protein-coupled receptors are found in all eukaryotes and constitute in vertebrates 3-5% of all genes. They are also very important drug targets and approximately 25 of the top 100 selling drugs are directed against these receptors. The Dictyostelium discoideum genome contains a surprisingly high number of 55 such receptors, approximately 0.5% of the encoded genes. Besides the four well-studied cAMP receptors the genome encodes eight additional cAMP receptor-like proteins and one of these is distinguished by a novel domain structure, one secretin-like receptor, 17 GABA(B)-like and 25 Frizzled-like receptors. The existence of the latter three types of receptors in D. discoideum was surprising because they had not been observed outside the animal kingdom before. Their presence suggests unprecedentedly complex and so far unknown signaling activities in this lower eukaryote.

Animals↗
PubMed2006

Dictyostelium transcriptional host cell response upon infection with Legionella.

Differential gene expression of Dictyostelium discoideum after infection with Legionella pneumophila was investigated using DNA microarrays. Investigation of a 48 h time course of infection revealed several clusters of co-regulated genes, an enrichment of preferentially up- or downregulated genes in distinct functional categories and also showed that most of the transcriptional changes occurred 24 h after infection. A detailed analysis of the 24 h time point post infection was performed in comparison to three controls, uninfected cells and co-incubation with Legionella hackeliae and L. pneumophilaDeltadotA. One hundred and thirty-one differentially expressed D. discoideum genes were identified as common to all three experiments and are thought to be involved in the pathogenic response. Functional annotation of the differentially regulated genes revealed that apart from triggering a stress response Legionella apparently not only interferes with intracellular vesicle fusion and destination but also profoundly influences and exploits the metabolism of its host. For some of the identified genes, e.g. rtoA involvement in the host response has been demonstrated in a recent study, for others such a role appears plausible. The results provide the basis for a better understanding of the complex host-pathogen interactions and for further studies on the Dictyostelium response to Legionella infection.

Animals↗
PubMed2006

Analysis of gene expression using cDNA microarrays.

A DNA microarray consists of an orderly arrangement of DNA, cDNA, or oligonucleotide probes that represent individual genes of an organism. Today, microarrays are the most important tools used to analyze gene transcription on a large scale. Investigations reach from finding key genes in whole genomes to looking at overall patterns of gene expression. This provides a better understanding of the architecture of genetic regulatory networks and helps to identify those genes in the genome that are differentially expressed, e.g., during cellular programs, in tumors or as the result of a treatment. The use of different fluorescent dyes allows mRNAs from two different sources (tissues, strains, or differently treated cells) to be labeled in different colors, mixed, and hybridized to the same array, which results in competitive binding of the target to the arrayed sequences. After hybridization and washing, the slide is scanned using two different wavelengths corresponding to the dyes used, and the intensity of the same spot in both channels is compared. This results in a measurement of the ratio of transcript levels for each gene represented on the array. In order to find significantly regulated genes, the resulting data are analyzed by different bioinformatic tools such as R and significance analysis of microarrays (SAM).

Animals↗
PubMed2005

Silencing of retrotransposons in Dictyostelium by DNA methylation and RNAi.

We have identified a DNA methyltransferase of the Dnmt2 family in Dictyostelium that was denominated DnmA. Expression of the dnmA gene is downregulated during the developmental cycle. Overall DNA methylation in Dictyostelium is approximately 0.2% of the cytosine residues, which indicates its restriction to a limited set of genomic loci. Bisulfite sequencing of specific sites revealed that DnmA is responsible for methylation of mostly asymmetric C-residues in the retrotransposons DIRS-1 and Skipper. Disruption of the gene resulted in a loss of methylation and in increased transcription and mobilization of Skipper. Skipper transcription was also upregulated in strains that had genes encoding components of the RNA interference pathway disrupted. In contrast, DIRS-1 expression was not affected by a loss of DnmA but was strongly increased in strains that had the RNA-directed RNA polymerase gene rrpC disrupted. A large number of siRNAs were found that corresponded to the DIRS-1 sequence, suggesting concerted regulation of DIRS-1 expression by RNAi and DNA modification. No siRNAs corresponding to the standard Skipper element were found. The data show that DNA methylation plays a crucial role in epigenetic gene silencing in Dictyostelium but that different, partially overlapping mechanisms control transposon silencing.

Amino Acid Sequence↗
PubMed2005

The Dictyostelium genome encodes numerous RasGEFs with multiple biological roles.

BACKGROUND: Dictyostelium discoideum is a eukaryote with a simple lifestyle and a relatively small genome whose sequence has been fully determined. It is widely used for studies on cell signaling, movement and multicellular development. Ras guanine-nucleotide exchange factors (RasGEFs) are the proteins that activate Ras and thus lie near the top of many signaling pathways. They are particularly important for signaling in development and chemotaxis in many organisms, including Dictyostelium. RESULTS: We have searched the genome for sequences encoding RasGEFs. Despite its relative simplicity, we find that the Dictyostelium genome encodes at least 25 RasGEFs, with a few other genes encoding only parts of the RasGEF consensus domains. All appear to be expressed at some point in development. The 25 genes include a wide variety of domain structures, most of which have not been seen in other organisms. The LisH domain, which is associated with microtubule binding, is seen particularly frequently; other domains that confer interactions with the cytoskeleton are also common. Disruption of a sample of the novel genes reveals that many have clear phenotypes, including altered morphology and defects in chemotaxis, slug phototaxis and thermotaxis. CONCLUSION: These results suggest that the unexpectedly large number of RasGEF genes reflects an evolutionary expansion of the range of Ras signaling rather than functional redundancy or the presence of multiple pseudogenes.

Actins↗
PubMed2005

DictyMOLD-a Dictyostelium discoideum genome browser database.

UNLABELLED: With the Dictyostelium Genome Project nearing completion, we initiated the construction of a data repository for all Dictyostelium discoideum genomic data. Up to now this database, called DictyMOLD (Dicty Map Of Linked Data), incorporates the recently completed D.discoideum chromosomes 1 and 2 sequences together with related annotations. To visualise maps, sequences and annotations and to provide access for the scientific community a perl-based browser was developed. AVAILABILITY: The DictyMOLD database is freely accessible via http://genome.imb-jena.de/dictyostelium/ CONTACT: gernot@imb-jena.de.

Animals↗
PubMed2005

Comparative genomics of Dictyostelium discoideum and Entamoeba histolytica.

Amoebozoa represent one of the earliest branches from the last common ancestor of all eukaryotes and contain some of the most dangerous human pathogens. Two amoebozoan genomes -- from the model organism Dictyostelium discoideum and the human pathogen Entamoeba histolytica -- have been published this year. Owing to their high A+T content, both genomes were difficult to sequence. In addition to nine amoebozoan expressed sequence tag projects, efforts are underway for comparative sequencing of four additional Entamoeba species. The completed genome sequences of D. discoideum and E. histolytica revealed unusual telomere structures, a high percentage of repetitive elements and a remarkably high gene content that is close to the one of Drosophila melanogaster. Finally, both organisms are brilliant examples of the influence of the lifestyle of an organism on its genome.

Adaptation, Biological↗
PubMed2005

Manifestations of multicellularity: Dictyostelium reports in.

The recent release of the Dictyostelium genome sequence is important because Dictyostelium has become a much-favoured model system for cell and developmental biologists. The sequence has revealed a remarkably high total number of approximately 12 500 genes, only a thousand fewer than are encoded by Drosophila. Previous protein-sequence comparisons suggested that Dictyostelium is evolutionarily closer to animals and fungi than to plants, and the global protein sequence comparison, now made possible by the genome sequence, confirms this. This review focuses on several classes of proteins that are shared by Dictyostelium and animals: a highly sophisticated array of microfilament components, a large family of G-protein-coupled receptors and a diverse set of SH2 domain-containing proteins. The presence of these proteins strengthens the case for a relatively close relationship with animals and extends the range of problems that can be addressed using Dictyostelium as a model organism.

Animals↗
PubMed2003

Revamp a model-status and prospects of the Dictyostelium genome project.

International efforts are underway that aim at determining the complete genome sequence of the social amoeba Dictyostelium discoideum. As strategy, a whole chromosome shotgun (WCS) approach was chosen and each of the six Dictyostelium chromosomes was assigned to project partners. The project is well advanced, chromosome 2 was recently published, and it is expected that the sequences of chromosomes 1 and 6 and a gene catalogue for the complete genome will be available at the end of this year. The genome sequence, together with powerful molecular genetic tools, will undoubtedly further accelerate Dictyostelium research into a number of fundamental biological processes that are common to a wide range of eukaryotes. Furthermore, it will constitute the basis for genome-wide functional analyses. The integration of results from these studies should ultimately lead to a better understanding of the complex networks that govern cellular behavior and development.

Animals↗
PubMed2003

Crawling into a new era-the Dictyostelium genome project.

The social amoeba Dictyostelium discoideum is a well-established model organism for the study of basic aspects of differentiation, signal transduction, phagocytosis, cytokinesis and cell motility. Its genome is being sequenced by an international consortium using a whole chromosome shotgun approach. The pacemaker of the D.discoideum genome project has been chromosome 2, the largest chromosome, which at 8 Mb represents approximately 25% of the genome and whose sequence and analysis have been published recently. Chromosomes 1 and 6 are close to being finished. To accelerate completion of the genome sequence, the next step in the project will be a whole-genome assembly followed by the analysis of the complete gene content. The completed genome sequence and its analysis provide the basis for genome-wide functional studies. It will position Dictyostelium at the same level as other model organisms and further enhance its experimental attractiveness.

Animals↗
PubMed2003

The Nucleo-cytoplasmic actin-binding protein CapG lacks a nuclear export sequence present in structurally related proteins.

Despite thorough structure-function analyses, it remains unclear how CapG, a ubiquitous F-actin barbed end capping protein that controls actin microfilament turnover in cells, is able to reside in the nucleus and cytoplasm, whereas structurally related actin-binding proteins are predominantly cytoplasmic. Here we report the molecular basis for the different subcellular localization of CapG, severin, and fragminP. Green fluorescent protein-tagged fragminP and severin accumulate in the nucleus upon treatment of transfected cells with the CRM1 inhibitor leptomycin B. We identified a nuclear export sequence in severin and fragminP, which is absent in CapG. Deletion of amino acids Met(1)-Leu(27) resulted in nuclear accumulation of severin and fragminP. Tagging this sequence to CapG triggered nuclear export, whereas mutation of single leucine residues (Leu(17), Leu(21), and Leu(27)) in the export sequence inhibited nuclear export. Based on these findings, a nuclear export signal was identified in myopodin, a muscle-specific actin-binding protein, and the Bloom syndrome protein, a RecQ-like helicase. Deletion of the myopodin nuclear export sequence blocked invasion into collagen type I of C2C12 cells transiently overexpressing myopodin. Our findings explain regulated subcellular targeting of distinct classes of actin-binding proteins.

Actins↗
PubMed2002

Sequence and analysis of chromosome 2 of Dictyostelium discoideum.

The genome of the lower eukaryote Dictyostelium discoideum comprises six chromosomes. Here we report the sequence of the largest, chromosome 2, which at 8 megabases (Mb) represents about 25% of the genome. Despite an A + T content of nearly 80%, the chromosome codes for 2,799 predicted protein coding genes and 73 transfer RNA genes. This gene density, about 1 gene per 2.6 kilobases (kb), is surpassed only by Saccharomyces cerevisiae (one per 2 kb) and is similar to that of Schizosaccharomyces pombe (one per 2.5 kb). If we assume that the other chromosomes have a similar gene density, we can expect around 11,000 genes in the D. discoideum genome. A significant number of the genes show higher similarities to genes of vertebrates than to those of other fully sequenced eukaryotes. This analysis strengthens the view that the evolutionary position of D. discoideum is located before the branching of metazoa and fungi but after the divergence of the plant kingdom, placing it close to the base of metazoan evolution.

Animals↗