Julio Mateos-Langerak

Spatially confined folding of chromatin in the interphase nucleus

Genome function in higher eukaryotes involves major changes in the spatial organization of the chromatin fiber. Nevertheless, our understanding of chromatin folding is remarkably limited. Polymer models have been used to describe chromatin folding. However, none of the proposed models gives a satisfactory explanation of experimental data. In particularly, they ignore that each chromosome occupies a confined space, i.e., the chromosome territory. Here, we present a polymer model that is able to describe key properties of chromatin over length scales ranging from 0.5 to 75 Mb. This random loop (RL) model assumes a self-avoiding random walk folding of the polymer backbone and defines a probability P for 2 monomers to interact, creating loops of a broad size range. Model predictions are compared with systematic measurements of chromatin folding of the q-arms of chromosomes 1 and 11. The RL model can explain our observed data and suggests that on the tens-of-megabases length scale P is small, i.e., 10–30 loops per 100 Mb. This is sufficient to enforce folding inside the confined space of a chromosome territory. On the 0.5- to 3-Mb length scale chromatin compaction differs in different subchromosomal domains. This aspect of chromatin structure is incorporated in the RL model by introducing heterogeneity along the fiber contour length due to different local looping probabilities. The RL model creates a quantitative and predictive framework for the identification of nuclear components that are responsible for chromatin–chromatin interactions and determine the 3-dimensional organization of the chromatin fiber.

The Three-Dimensional Structure of Human Interphase Chromosomes Is Related to the Transcriptome Map

ABSTRACT The three-dimensional (3D) organization of the chromosomal fiber in the human interphase nucleus is an important but poorly understood aspect of gene regulation. Here we quantitatively analyze and compare the 3D structures of two types of genomic domains as defined by the human transcriptome map. While ridges are gene dense and show high expression levels, antiridges, on the other hand, are gene poor and carry genes that are expressed at low levels. We show that ridges are in general less condensed, more irregularly shaped, and located more closely to the nuclear center than antiridges. Six human cell lines that display different gene expression patterns and karyotypes share these structural parameters of chromatin. This shows that the chromatin structures of these two types of genomic domains are largely independent of tissue-specific variations in gene expression and differentiation state. Moreover, we show that there is remarkably little intermingling of chromatin from different parts of the same chromosome in a chromosome territory, neither from adjacent nor from distant parts. This suggests that the chromosomal fiber has a compact structure that sterically suppresses intermingling. Together, our results reveal novel general aspects of 3D chromosome architecture that are related to genome structure and function.

Nuclear architecture: Is it important for genome function and can we prove it?

Gene regulation in higher eukaryotes has been shown to involve regulatory sites, such as promoters and enhancers which act at the level of individual genes, and mechanisms which control the functional state of gene clusters. A fundamental question is whether additional levels of genome control exist. Nuclear organization and large‐scale chromatin structure may constitute such a level and play an important role in the cell‐type specific orchestration of the expression of thousands of genes in eukaryotic cells. Numerous observations indicate a tight correlation between genome activity and nuclear and large‐scale chromatin structure. However, causal relationships are rare. Here we explore how these might be uncovered. J. Cell. Biochem. 102: 1067–1075, 2007.

Shape normalization of 3D cell nuclei using elastic spherical mapping

Topological analysis of cells and subcellular structures on the basis of image data, is one of the major trends in modern quantitative biology. However, due to the dynamic nature of cell biology, the optical appearance of different cells or even time‐series of the same cell is undergoing substantial variations in shape and texture, which makes a comparison of shapes and distances across different cells a nontrivial task. In the absence of canonical invariances, a natural approach to the normalization of cells consists of spherical mapping, enabling the analysis of targeted regions in terms of canonical spherical coordinates, that is, radial distances and angles. In this work, we present a physically‐based approach to spherical mapping, which has been applied for topological analysis of multichannel confocal laser scanning microscopy images of human fibroblast nuclei. Our experimental results demonstrate that spherical mapping of entire nuclear domains can automatically be obtained by inverting affine and elastic transformations, performed on a spherical finite element template mesh.

Truncated HP1 lacking a functional chromodomain induces heterochromatinization upon in vivo targeting

Packaging of the eukaryotic genome into higher order chromatin structures is tightly related to gene expression. Pericentromeric heterochromatin is typified by accumulations of heterochromatin protein 1 (HP1), methylation of histone H3 at lysine 9 (MeH3K9) and global histone deacetylation. HP1 interacts with chromatin by binding to MeH3K9 through the chromodomain (CD). HP1 dimerizes with itself and binds a variety of proteins through its chromoshadow domain. We have analyzed at the single cell level whether HP1 lacking its functional CD is able to induce heterochromatinization in vivo. We used a lac-operator array-based system in mammalian cells to target EGFP-lac repressor tagged truncated HP1α and HP1β to a lac operator containing gene-amplified chromosome region in living cells. After targeting truncated HP1α or HP1β we observe enhanced tri-MeH3K9 and recruitment of endogenous HP1α and HP1β to the chromosome region. We show that CD-less HP1α can induce chromatin condensation, whereas the effect of truncated HP1β is less pronounced. Our results demonstrate that after lac repressor-mediated targeting, HP1α and HP1β without a functional CD are able to induce heterochromatinization.

Variability analysis of the large-scale structure of interphase chromatin fiber based on statistical shape theory

The relationship between geometric folding of the chromatin fiber and genome function is a key issue in cell biology. We propose different approaches based on statistical shape theory to investigate the geometric variability of chromatin folding in nuclei of interphase human fibroblasts. Our main purpose is to assess the degree of variability of folding of the chromatin fiber, measured by fluorescent in situ hybridization, using BAC probes in combination with 3D confocal microscopy. We employ point-based registration, the complex Bingham distribution, generalized Procrustes method, and the Kendall spherical coordinate system. The approaches have been applied using 337 3D multi-channel microscopy images. We have analyzed the geometric structure formed by gene-rich highly expressed genomic regions and areas that are gene-poor and have a low transcriptional activity. It turned out that the structure formed by these genomic regions exhibit high shape variation, however, most of them can be characterized by a non-uniform shape distribution.

Geometrical probability approach for analysis of 3D chromatin structure in interphase cell nuclei

Investigation of 3D chromatin structure in interphase cell nuclei is important for the understanding of genome function. For a reconstruction of the 3D architecture of the human genome, systematic fluorescent in situ hybridization in combination with 3D confocal laser scanning microscopy is applied. The position of two or three genomic loci plus the overall nuclear shape were simultaneously recorded, resulting in statistical series of pair and triple loci combinations probed along the human chromosome 1 q-arm. For interpretation of statistical distributions of geometrical features (e.g. distances, angles, etc.) resulting from finite point sampling experiments, a Monte-Carlo-based approach to numerical computation of geometrical probability density functions (PDFs) for arbitrarily-shaped confined spatial domains is developed. Simulated PDFs are used as bench marks for evaluation of experimental PDFs and quantitative analysis of dimension and shape of probed 3D chromatin regions. Preliminary results of our numerical simulations show that the proposed numerical model is capable to reproduce experimental observations, and support the assumption of confined random folding of 3D chromatin fiber in interphase cell nuclei

Analyzing the variability of the 3D structure of chromatin fiber using statistical shape theory

The relationship between geometric folding of the chromatin fiber and genome function is a key issue in cell biology. We propose different approaches based on statistical shape theory to investigate the geometric variability of chromatin folding in nuclei of interphase human fibroblasts. Our main purpose is to assess the degree of variability of folding of the chromatin fiber, measured by fluorescent in situ hybridization, using BAC probes in combination with 3D confocal microscopy. We employ point-based registration, the complex Bingham distribution, generalized Procrustes analysis, and the Kendall spherical coordinate system. The approaches have been applied using 337 3D multi-channel microscopy images. We have analyzed the geometric structure formed by gene-rich highly expressed genomic regions and areas that are gene-poor and have a low transcriptional activity. It turned out that the structure formed by these genomic regions exhibit high shape variation, however, most of them can be characterized by a non-uniform shape distribution.

Stochastical analysis of finite point sampling of 3D chromatin fiber in interphase cell nuclei

Investigation of 3D chromatin structure in interphase cell nuclei is important for the understanding of genome function. For a reconstruction of the 3D architecture of the human genome, systematic fluorescent in situ hybridization in combination with 3D confocal laser scanning microscopy is applied. The position of two or three genomic loci plus the overall nuclear shape were simultaneously recorded, resulting in statistical series of pair and triple loci combinations probed along the human chromosome 1 q-arm. For interpretation of statistical distributions of geometrical features (e.g. distances, angles, etc.) resulting from finite point sampling experiments, a Monte-Carlo-based approach to numerical computation of geometrical probability density functions (PDFs) for arbitrarily-shaped confined spatial domains is developed. Simulated PDFs are used as bench marks for evaluation of experimental PDFs and quantitative analysis of dimension and shape of probed 3D chromatin regions. Preliminary results of our numerical simulations show that the proposed numerical model is capable to reproduce experimental observations, and support the assumption of confined random folding of 3D chromatin fiber in interphase cell nuclei.

Topological analysis of 3D cell nuclei using finite element template-based spherical mapping

Topological analysis of cells and subcellular structures on the basis of image data is one of the major trends in modern quantitative biology. However, due to the dynamic nature of cell biology, the optical appearance of different cells or even time series of the same cell is undergoing substantial variations in shape and texture which makes the analysis of image data a non-trivial task. In the absence of canonical invariances, a natural approach to the normalization of cell images consists in dimension reduction of the 3D problem by means of spherical mapping which enables the analysis of targeted regions in terms of radial distances. In this work, we present a finite element template-based approach for physically-base spherical mapping which has been applied for topological analysis of confocal laser scanning microscopy images of cell nuclei.