From observations to paradigms; the importance of theories and models. An interview with Hans Meinhardt by Richard Gordon and Lev Beloussov

Hans Meinhardt received his PhD in physics from the University of Cologne at 1966. For a postdoctoral fellowship, he went to the European High Energy Laboratory CERN in Geneva where he joined a group working on the leptonic decay of the Xi-minus particle. One of his duties was to perform computer simulations to optimize the complex experimental setup -- a skill which turned out to be helpful later on. In 1969 he switched to biology and joined the department of Alfred Gierer at the Max Planck Institute for Developmental Biology (formerly Virus Research) in Tubingen. His interest was focused on mechanisms of biological pattern formation. Using computer simulations as a tool, he developed models for essential steps in development. Most fascinating for him was the possibility to recapitulate and to reconstructusing the computer the genesis of structures where no structures were before and to see how these emerging structures become subsequently further refined. In addition to the interaction with Alfred Gierer and his group working on hydra development, the Max-Planck Institute as a whole provided a very stimulating environment. In the seventies, the work of Klaus Sander on gradients in early insect development was highly influential. Collaboration with Martin Klinger in the eighties revealed that the pigmentation patterns on tropical sea shells are convenient to study highly dynamic patterning processes. The variability and the asthetic beauty of these patterns turned out to result from the chaotic nature of the underlying reactions. Mechanisms deduced from shell patterns became a key to understand other developing systems such as orientation of chemotactic cells or phyllotaxis. Officially Hans Meinhardt retired at the end of 2003. At present he works on refinements and extensions of models which account for the different modes of embryonic axis formation in different phyla from an evolutionary point of view.     

Models for organizer and notochord formation

In the development of higher organisms, small groups of cells can play an important role by directing the fate of the surrounding cells. Models are discussed that account for the generation of such organizing regions. The generation of local high concentrations of signalling substances was proposed to depend on local self-enhancement combined with a long-range inhibition. The model accounts for pattern regulation, for instance, for the formation of multiple embryos after fragmentation of the early blastodisc in chickens or for head regeneration in the fresh water polyp Hydra. The model has found support from more recently discovered interactions involved in organizer formation. The mutual down-regulation of noggin/chordin and BMP-4 is proposed to function as an indirect self-enhancement, establishing in this way an essential prerequisite for primary pattern formation. Self-enhancement and long-range inhibition is also crucial for the generation of substructures such as bristles or tracheae. A poisoning of an organizing region by a second antagonistic reaction of a short range but a long time constant can lead to its displacement. Long extended structures can be formed as a trace behind the moving organizer. The notochord and the tracheae of insects are discussed as examples.     

An Archaeal Immune System Can Detect Multiple Protospacer Adjacent Motifs (PAMs) to Target Invader DNA

The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated (Cas) system provides adaptive and heritable immunity against foreign genetic elements in most archaea and many bacteria. Although this system is widespread and diverse with many subtypes, only a few species have been investigated to elucidate the precise mechanisms for the defense of viruses or plasmids. Approximately 90% of all sequenced archaea encode CRISPR/Cas systems, but their molecular details have so far only been examined in three archaeal species: Sulfolobus solfataricus, Sulfolobus islandicus, and Pyrococcus furiosus. Here, we analyzed the CRISPR/Cas system of Haloferax volcanii using a plasmid-based invader assay. Haloferax encodes a type I-B CRISPR/Cas system with eight Cas proteins and three CRISPR loci for which the identity of protospacer adjacent motifs (PAMs) was unknown until now. We identified six different PAM sequences that are required upstream of the protospacer to permit target DNA recognition. This is only the second archaeon for which PAM sequences have been determined, and the first CRISPR group with such a high number of PAM sequences. Cells could survive the plasmid challenge if their CRISPR/Cas system was altered or defective, e.g. by deletion of the cas gene cassette. Experimental PAM data were supplemented with bioinformatics data on Haloferax and Haloquadratum.     

Genetic analysis of the Bacillus licheniformis degSU operon and the impact of regulatory mutations on protease production

Disruption experiments targeted at the Bacillus licheniformis degSU operon and GFP-reporter analysis provided evidence for promoter activity immediately upstream of degU. pMutin mediated concomitant introduction of the degU32 allele--known to cause hypersecretion in Bacillus subtilis-- resulted in a marked increase in protease activity. Application of 5-fluorouracil based counterselection through establishment of a phosphoribosyltransferase deficient Δupp strain eventually facilitated the marker-free introduction of degU32 leading to further protease enhancement achieving levels as for hypersecreting wild strains in which degU was overexpressed. Surprisingly, deletion of rapG--known to interfere with DegU DNA-binding in B. subtilis--did not enhance protease production neither in the wild type nor in the degU32 strain. The combination of degU32 and Δupp counterselection in the type strain is not only equally effective as in hypersecreting wild strains with respect to protease production but furthermore facilitates genetic strain improvement aiming at biological containment and effectiveness of biotechnological processes.     

Haplotype reconstruction error as a classical misclassification problem: introducing sensitivity and specificity as error measures

Background

Statistically reconstructing haplotypes from single nucleotide polymorphism (SNP) genotypes, can lead to falsely classified haplotypes. This can be an issue when interpreting haplotype association results or when selecting subjects with certain haplotypes for subsequent functional studies. It was our aim to quantify haplotype reconstruction error and to provide tools for it.

Methods and Results

By numerous simulation scenarios, we systematically investigated several error measures, including discrepancy, error rate, and R², and introduced the sensitivity and specificity to this context. We exemplified several measures in the KORA study, a large population-based study from Southern Germany. We find that the specificity is slightly reduced only for common haplotypes, while the sensitivity was decreased for some, but not all rare haplotypes. The overall error rate was generally increasing with increasing number of loci, increasing minor allele frequency of SNPs, decreasing correlation between the alleles and increasing ambiguity.

Conclusions

We conclude that, with the analytical approach presented here, haplotype-specific error measures can be computed to gain insight into the haplotype uncertainty. This method provides the information, if a specific risk haplotype can be expected to be reconstructed with rather no or high misclassification and thus on the magnitude of expected bias in association estimates. We also illustrate that sensitivity and specificity separate two dimensions of the haplotype reconstruction error, which completely describe the misclassification matrix and thus provide the prerequisite for methods accounting for misclassification.     

Adaptive changes of rat liver cells induced by repeated intraperitoneal injections of d-galactosamine. III-Light- and electron-microscopic investigations of hepatocellular cytoplasmic changes.

In rats hepatocellular cytoplasmic changes after daily repeated D-galactosamine (GalN) intoxication--i.e. subacute GalN intoxication--were studied by light and electron microscopy. The number of GalN injections--and thus the days of survival--was between one and 30. The rats were killed six hours after the last GalN injection. Less degenerative changes were found after repeated GalN injections. An increased formation of atypical dense bodies (ADB), a temporary pronounced lipid accumulation and changes of the rough and smooth endoplasmic reticulum were prominent features of subacute GalN intoxication. The implications with respect to a modified GalN action in subacute GalN intoxication are discussed with special reference to biochemical data obtained in the same experimental model (Schuchhardt et al., 1977).