Jörg Galle

Transcriptional regulation by histone modifications: towards a theory of chromatin re-organization during stem cell differentiation

Chromatin-related mechanisms, as e.g. histone modifications, are known to be involved in regulatory switches within the transcriptome. Only recently, mathematical models of these mechanisms have been established. So far they have not been applied to genome-wide data. We here introduce a mathematical model of transcriptional regulation by histone modifications and apply it to data of trimethylation of histone 3 at lysine 4 (H3K4me3) and 27 (H3K27me3) in mouse pluripotent and lineage-committed cells. The model describes binding of protein complexes to chromatin which are capable of reading and writing histone marks. Molecular interactions of the complexes with DNA and modified histones create a regulatory switch of transcriptional activity. The regulatory states of the switch depend on the activity of histone (de-) methylases, the strength of complex-DNA-binding and the number of nucleosomes capable of cooperatively contributing to complex-binding. Our model explains experimentally measured length distributions of modified chromatin regions. It suggests (i) that high CpG-density facilitates recruitment of the modifying complexes in embryonic stem cells and (ii) that re-organization of extended chromatin regions during lineage specification into neuronal progenitor cells requires targeted de-modification. Our approach represents a basic step towards multi-scale models of transcriptional control during development and lineage specification.

Modeling the dynamic epigenome: from histone modifications towards self-organizing chromatin

Epigenetic mechanisms play an important role in regulating and stabilizing functional states of living cells. However, in spite of an increasing amount of experimental data, models of transcriptional regulation by epigenetic processes, in particular by histone modifications, are rather rare. In this article, we focus on epigenetic modes of transcriptional regulation based on histone modifications and their potential dynamical interplay with DNA methylation and higher-order chromatin structure. The main purpose of this article is to review recent formal modeling approaches to the dynamics and propagation of histone modifications and to relate them to available experimental data. We evaluate their assumptions with respect to recruitment of relevant modifiers, establishment and processing of modifications, and compare the emerging stability properties and memory effects. Theoretical predictions that await experimental validation are highlighted and potential extensions of these models towards multiscale models of self-organizing chromatin are discussed.

A Global Genome Segmentation Method for Exploration of Epigenetic Patterns

Current genome-wide ChIP-seq experiments on different epigenetic marks aim at unraveling the interplay between their regulation mechanisms. Published evaluation tools, however, allow testing for predefined hypotheses only. Here, we present a novel method for annotation-independent exploration of epigenetic data and their inter-correlation with other genome-wide features. Our method is based on a combinatorial genome segmentation solely using information on combinations of epigenetic marks. It does not require prior knowledge about the data (e.g. gene positions), but allows integrating the data in a straightforward manner. Thereby, it combines compression, clustering and visualization of the data in a single tool. Our method provides intuitive maps of epigenetic patterns across multiple levels of organization, e.g. of the co-occurrence of different epigenetic marks in different cell types. Thus, it facilitates the formulation of new hypotheses on the principles of epigenetic regulation. We apply our method to histone modification data on trimethylation of histone H3 at lysine 4, 9 and 27 in multi-potent and lineage-primed mouse cells, analyzing their combinatorial modification pattern as well as differentiation-related changes of single modifications. We demonstrate that our method is capable of reproducing recent findings of gene centered approaches, e.g. correlations between CpG-density and the analyzed histone modifications. Moreover, combining the clustered epigenetic data with information on the expression status of associated genes we classify differences in epigenetic status of e.g. house-keeping genes versus differentiation-related genes. Visualizing the distribution of modification states on the chromosomes, we discover strong patterns for chromosome X. For example, exclusively H3K9me3 marked segments are enriched, while poised and active states are rare. Hence, our method also provides new insights into chromosome-specific epigenetic patterns, opening up new questions how “epigenetic computation” is distributed over the genome in space and time.

Genomic and transcriptomic heterogeneity of colorectal tumors arising in Lynch Syndrome

Colorectal cancer (CRC) arising in Lynch syndrome (LS) comprises tumours with constitutional mutations in DNA mismatch repair genes. There is still a lack of whole‐genome and transcriptome studies of LS‐CRC to address questions about similarities and differences in mutation and gene expression characteristics between LS‐CRC and sporadic CRC, about the molecular heterogeneity of LS‐CRC, and about specific mechanisms of LS‐CRC genesis linked to dysfunctional mismatch repair in LS colonic mucosa and the possible role of immune editing. Here, we provide a first molecular characterization of LS tumours and of matched tumour‐distant reference colonic mucosa based on whole‐genome DNA‐sequencing and RNA‐sequencing analyses. Our data support two subgroups of LS‐CRCs, G1 and G2, whereby G1 tumours show a higher number of somatic mutations, a higher amount of microsatellite slippage, and a different mutation spectrum. The gene expression phenotypes support this difference. Reference mucosa of G1 shows a strong immune response associated with the expression of HLA and immune checkpoint genes and the invasion of CD4+ T cells. Such an immune response is not observed in LS tumours, G2 reference and normal (non‐Lynch) mucosa, and sporadic CRC. We hypothesize that G1 tumours are edited for escape from a highly immunogenic microenvironment via loss of HLA presentation and T‐cell exhaustion. In contrast, G2 tumours seem to develop in a less immunogenic microenvironment where tumour‐promoting inflammation parallels tumourigenesis. Larger studies on non‐neoplastic mucosa tissue of mutation carriers are required to better understand the early phases of emerging tumours. Copyright

Mice overexpressing CD97 in intestinal epithelial cells provide a unique model for mammalian postnatal intestinal cylindrical growth

Transgenic mice overexpressing CD97 in intestinal epithelial cells develop an upper megaintestine with normal microscopic morphology after birth and before weaning. Intestinal enlargement by CD97 depends on signaling but does not require binding of its ligand, CD55.

Monte Carlo simulation of primitive water in slit-like pores: networks and clusters

The structure of water in smooth slit-like pores is studied by means of a primitive molecular model of associating fluids due to Nezbeda and Smith [W.R. Smith, I. Nezbeda, J. Chem. Phys. 81 (1984) 3694]. MC simulations in an NVT ensemble are performed for different slit widths L ranging from one to about three diameters of the water molecule. Detailed information about cluster and network formation of the associating fluid confined in the pores is obtained. The change of the aggregate topology from linear chains in case of quasi two-dimensional narrow slits into tetrahedral bulk-like structures in wider slits is studied by the increase of L. The research provides basic knowledge for a theoretical understanding of hydration phenomena at interfaces at a molecular level.

Mesenchymal Stem Cell Heterogeneity and Ageing In Vitro: A Model Approach

Mesenchymal Stem Cell (MSC)-based therapies have been suggested as a particular promising strategy in tissue regeneration. These cells can easily be obtained from the patient and are able to produce a large number of progeny that can be induced to form connective tissue. However, rapid amplification of the isolated cells is required for their therapeutic application. While already the isolated populations are heterogeneous regarding various functional and molecular aspects, this heterogeneity further evolves during amplification. Understanding the origin and development of MSC heterogeneity will help to improve MSC culture conditions and thus facilitate their clinical use. We here review recent results on MSC heterogeneity and introduce a mathematical framework that approaches MSC heterogeneity on the single cell level. This approach bases on the concept of noise-driven MSC differentiation and allows describing MSC heterogeneity with respect to their differentiation state and age. It is capable of describing the impact of MSC heterogeneity on in vitro expansion and differentiation. We present new results on the formation of an age structure in MSC populations in vitro and the age-dependent differentiation structure of MSC populations. Moreover, we discuss open questions regarding MSC adaptation to changing environments and the cell intrinsic control of state fluctuations.

Monte Carlo simulation of hard sphere fluids in planar slits with molecularly rough surfaces

Abstract The structural and thermodynamic behaviour of hard-sphere fluids confined in planar slit-like pores with hard rough walls is investigated by canonical Monte Carlo simulations. The surfaces of the pores are modelled by means of parallel impenetrable walls. Additionally a molecular “roughness” of the pore surfaces is introduced using several models of randomly distributed hard spheres located (fixed or restricted in motion) at the walls. The density distribution of the hard spheres in the pore and the pressure normal to the walls are estimated and compared with the behaviour of fluids in slit-like pores with smooth hard walls. The results are discussed in the context of experimental studies of fluids confined in the polar interfaces of bilayers.

On the impact of single cell biomechanics on the spatio-temporal organization of regenerative tissue

Collective phenomena in multi-cellular assemblies can be modeled on different levels of complexity. So-called agent-based or individual-based models (IBMs) consider the dynamics of each individual cell. As a special feature, these models allow to account for intracellular decision processes which are triggered by biomechanical cell-cell or cell-matrix interactions. In simulation studies of lattice-free IBMs we analyzed the spatio-temporally organization of intestinal (ISC) and mesenchymal (MSC) stem cell systems. We discuss the impact of cell contact formation on the growth and homeostasis of these complex systems. We demonstrate that cell biomechanics can contribute in regulating homeostasis of the intestinal epithelium by affecting both tissue shape and function. Moreover, cell contact formation is suggested to generate an age structure in expanding MSC populations in vitro increasing their population heterogeneity. Our results implicate that a biomechanical characterization of single cells depending on their degree of differentiation and age is essential for a comprehensive understanding of regenerative tissue.

Der Zelloberflächenrezeptor CD97 stimuliert die Migration von Tumorzellen durch Interaktion mit dem Aktin-Zytoskelett

Introduction: The presence of scattered tumor cells at the invasion front of several carcinomas has clinical significance. These cells differ in their protein expression from cells in central tumor regions as shown by us for the EGF- seven transmembrane (TM7) receptor CD97 [1]. We recently clarified the mechanism by which CD97 in scattered tumor cells facilitates tumor invasion and showed that the short CD97 isoform (EGF 1,2,5/TM7) increased random intrinsic migration of tumor cells [2]. Here, we elucidated the molecular mechanisms involved in CD97 stimulated tumor cell migration. Methods: Cells stable overexpressing the whole CD97 (EGF 1,2,5/TM7) molecule, C-terminally truncated CD97 (TM5, TM3, and TM1) as well as CD97 missing the extracellular EGF-domains (delta EGF/TM7) were generated. The cells were applied to migration tracking experiments and laser scanning microscopy to investigate the surface cell distribution of CD97. Moreover, microarray analysis and pulldown-assays were used to define intracellular signaling during migration. Results: In nonconfluent 2D cultured CD97 overexpressing cells, the molecule was concentrated in membrane ruffles at the leading edge of migrating cells, co-localized with the raft marker ganglioside GM1, actin filaments and α-actinin. Treatment that disrupts actin filaments but not microtubules caused loss of CD97 from the ruffles, suggesting an interaction with the actin cytoskeleton in the localization and function of CD97. Eliminating the N-terminal EGF-domains of CD97 diminished migration but did not interrupt it. However, step-wise truncation of CD97 at the C-terminus to TM5 and TM3 decreased random migration. CD97 with only one transmembrane domain (TM1) did not migrate indicating intracellular signaling through the TM7 part during migration. Pull-down assays, microarray analysis and real-time RT-PCR revealed the involvement of the small Rho and Rac GTPases and LIM-kinase 1 in coupling CD97 to actin cytoskeleton formation and thus migration. Conclusion: Malignant cells utilize their intrinsic migratory ability to invade and to metastasize. CD97, upregulated in scattered invasive tumor cells, links migratory signals to the actin cytoskeleton.