**1. Introduction to the History and State of the Art**

Inheritance has always played a central part in the quest for elucidating the origin of nature, life and mankind. Beyond the epic mythical assumptions, it also has been obvious for millennia that the evolutionary transfer of information plays a key role during the manipulation of inheritance by mating and breeding. Already in antique times many a "theory" was devoted to the apparent, as well as especially to the obvious fact that nature seemed to be composed of small, similar, and consistent subcomponents—so called atoms. With the description of the tissue of plants (including its substructures of vesicles and bubbles) by Robert Hooke or in the case of the cell nucleus by Anton van Leeuwenhook, in the 17th century new momentum entered the field. Nevertheless, it took until 1830 when Robert Brown defined the cell nucleus as such and until 1939 when Theodor Schwann established the cell as the fundamental unit of all plant and animal tissues while linking to the assumed fundamental design principle of life as well as nature in general. Despite fast growing microscopic resolutions there were huge challenges: not only staining and visualization methods were lacking, but also huge preparatory issues were faced especially concerning the "notorious" hard to stain cell nucleus. With the development of the natural sciences many a discovery was made culminating in the structural description of the DNA double helix [1] and the discovery of the nucleosome [2–4] at the atomic level, full genome sequences and finally histone modifications defining epigenetic landscapes. It also became obvious that the structure and function of genomes co-evolved as an inseparable system allowing the physical storage, replication, and expression of genetic information [5–7].

interphase organization as in the models of Comings [30, 31] or Vogel and Schroeder [32]. In the radial-loop-scaffold model of Paulson and Laemmli [33] ~60 kbp-sized chromatin loops attached to a nuclear matrix/scaffold explained the condensation degree of metaphase chromosomes. According to Pienta and Coffey [34], these loops persisted in interphase and formed stacked rosettes in metaphase. Micro-irradiation studies by C. Cremer and T. Cremer [35, 36] and fluorescence *in situ* hybridization (FISH) by Lichter [37] as well as C. Cremer and T. Cremer [38] and publications thereafter [39, 40], confirmed a territorial organization of chromosomes, their arms, and stable sub-chromosomal domains during interphase, including their structural persistence during metaphase (de-)condensation. The assumption since then has been that the ~850 G, Q, R, and C ideogram bands [41, 42] split into and thus also consist actually of ~2500 subchromosomal interphase domains. Chromatin rosettes explaining a (sub-)territorial folding were first visualized using electron microscopy by Jekatrina Erenpreisa [43] and others [44] but remained unappreciated, until Belmont and Bruce proposed the EM-based helical hierarchy chromonema fibre (CF) model [45]. Spatial distance measurements between small FISH-labelled genetic regions, led to the Random-Walk/Giant-Loop (RW/GL) model with the first analytical looped polymer description by Sachs [46–48]. Here, 1 to 5 Mbp loops are attached to a non-protein backbone, following the line of Pienta and Coffey [34]. Later, a combination of distance measurements using structure-preserving FISH protocols, high-resolution microscopy, and huge parallel polymer simulations of chromosomes and entire cell nuclei, only were compatible with the rosette-like Multi-Loop-Subcompartment (MLS) model in which around 60 to 120 kbp loops form rosettes connected by similar sized linkers [7, 21–24, 49, 50]. Thereafter, the RW/ GL model has then been discussed in terms of methodological "demolition" of the architecture [21, 22, 51, 52]. This is also in agreement with studies on replication (see [39] and thereafter). Again *in vivo* FCS measurements of nucleosome concentration distributions and dynamic and functional properties such as the diffusion of macromolecules are only in agreement with a small multi-loop aggregate/rosette-like chromatin folding [18–20, 22, 53, 54]. The fine-structured multi-scaling long-range correlations of the DNA sequence once

A Consistent Systems Mechanics Model of the 3D Architecture and Dynamics of Genomes

http://dx.doi.org/10.5772/intechopen.89836

69

To further distinguish between the different architecture proposals, proximity crosslinking techniques (developed and used already in the last century) were further developed into a family of interaction capture techniques such as 3C [56, 57], 3C-qPCR [58], 4C [59], 3C-seq/4Cseq [60], 5C [61], and Hi-C [62]. They once more confirmed the existence of looping and subchromosomal domains, now inconsistenly referred to as topologically associated domains (TAD; [63]) with a somewhat higher localization accuracy when compared to FISH. These approaches also led to a number of - although by the underlying (raw) data basically unsupported - conjectures (Imam et al., in preparation), e.g. the fractal globule model [62], the loop array architecture of mitotic chromosomes [64], and the highly dynamic loop formation based on single-cell experiments [65] or in a genome wide assay [66]. In contrast, with the introduction of targeted chromatin capture *T2C* [25, 67–69], we were able to show that the chromatin quasi-fibre forms small stable loops of ~30-100 kbp which form stable multi-loop aggregates/ rosettes connected by linkers of similar sizes as the loops [25, 26]. The development of our

again also predict this [22–24, 55].

novel *in vivo* FCS approach came to the same conclusion [27].

However, the immense size and structural complexity of genomes spanning many orders of magnitude has always imposed huge experimental challenges. Thus, the higher-order architecture has been and still is widely discussed with many interesting details yet to be described. Already how nucleosomes are spaced, positioned, remodelled, and whether and how nucleosome chains fold into fibres at physiological salt concentrations have been matters of continuing debate: e.g. Finch and Klug [8] proposed a relatively regular solenoid and *in vivo* neutron scattering experiments revealed a fibre diameter of 30 ± 5 nm as a dominant nuclear feature [9–12]. In contrast more recent work suggested no compaction at all (rev. [13, 14]), and highly polymorphic, nucleosome position- [15] and dynamic function-dependent structures [16, 17], which are essential to explain nucleosome concentration distributions [18–20], or dynamic and functional properties such as the nuclear diffusion of macromolecules. Moreover, the fine-structured multi-scaling long-range correlation behaviour of the DNA sequence also predicts a compacted chromatin fibre [21–24]. With a novel chromatin interaction technique—*T2C*—we were, however and indeed, able to show that nucleosomes form in general a quasi-fibre with a differential compaction of ~5 ± 1 nucleosomes/11 nm [25, 26], which is in agreement with a novel *in vivo* fluorescence correlation spectroscopy (FCS) approach measuring the dynamics of chromatin [27].

The higher-order chromatin architecture has been a matter of even greater debate: Pioneering light microscopy studies by Rabl [28] and Boveri [29] hinted towards a hierarchical self-similar, territorial organization. Electron microscopy suggested a more random interphase organization as in the models of Comings [30, 31] or Vogel and Schroeder [32]. In the radial-loop-scaffold model of Paulson and Laemmli [33] ~60 kbp-sized chromatin loops attached to a nuclear matrix/scaffold explained the condensation degree of metaphase chromosomes. According to Pienta and Coffey [34], these loops persisted in interphase and formed stacked rosettes in metaphase. Micro-irradiation studies by C. Cremer and T. Cremer [35, 36] and fluorescence *in situ* hybridization (FISH) by Lichter [37] as well as C. Cremer and T. Cremer [38] and publications thereafter [39, 40], confirmed a territorial organization of chromosomes, their arms, and stable sub-chromosomal domains during interphase, including their structural persistence during metaphase (de-)condensation. The assumption since then has been that the ~850 G, Q, R, and C ideogram bands [41, 42] split into and thus also consist actually of ~2500 subchromosomal interphase domains. Chromatin rosettes explaining a (sub-)territorial folding were first visualized using electron microscopy by Jekatrina Erenpreisa [43] and others [44] but remained unappreciated, until Belmont and Bruce proposed the EM-based helical hierarchy chromonema fibre (CF) model [45]. Spatial distance measurements between small FISH-labelled genetic regions, led to the Random-Walk/Giant-Loop (RW/GL) model with the first analytical looped polymer description by Sachs [46–48]. Here, 1 to 5 Mbp loops are attached to a non-protein backbone, following the line of Pienta and Coffey [34]. Later, a combination of distance measurements using structure-preserving FISH protocols, high-resolution microscopy, and huge parallel polymer simulations of chromosomes and entire cell nuclei, only were compatible with the rosette-like Multi-Loop-Subcompartment (MLS) model in which around 60 to 120 kbp loops form rosettes connected by similar sized linkers [7, 21–24, 49, 50]. Thereafter, the RW/ GL model has then been discussed in terms of methodological "demolition" of the architecture [21, 22, 51, 52]. This is also in agreement with studies on replication (see [39] and thereafter). Again *in vivo* FCS measurements of nucleosome concentration distributions and dynamic and functional properties such as the diffusion of macromolecules are only in agreement with a small multi-loop aggregate/rosette-like chromatin folding [18–20, 22, 53, 54]. The fine-structured multi-scaling long-range correlations of the DNA sequence once again also predict this [22–24, 55].

**1. Introduction to the History and State of the Art**

(FCS) approach measuring the dynamics of chromatin [27].

information [5–7].

68 Chromatin and Epigenetics

Inheritance has always played a central part in the quest for elucidating the origin of nature, life and mankind. Beyond the epic mythical assumptions, it also has been obvious for millennia that the evolutionary transfer of information plays a key role during the manipulation of inheritance by mating and breeding. Already in antique times many a "theory" was devoted to the apparent, as well as especially to the obvious fact that nature seemed to be composed of small, similar, and consistent subcomponents—so called atoms. With the description of the tissue of plants (including its substructures of vesicles and bubbles) by Robert Hooke or in the case of the cell nucleus by Anton van Leeuwenhook, in the 17th century new momentum entered the field. Nevertheless, it took until 1830 when Robert Brown defined the cell nucleus as such and until 1939 when Theodor Schwann established the cell as the fundamental unit of all plant and animal tissues while linking to the assumed fundamental design principle of life as well as nature in general. Despite fast growing microscopic resolutions there were huge challenges: not only staining and visualization methods were lacking, but also huge preparatory issues were faced especially concerning the "notorious" hard to stain cell nucleus. With the development of the natural sciences many a discovery was made culminating in the structural description of the DNA double helix [1] and the discovery of the nucleosome [2–4] at the atomic level, full genome sequences and finally histone modifications defining epigenetic landscapes. It also became obvious that the structure and function of genomes co-evolved as an inseparable system allowing the physical storage, replication, and expression of genetic

However, the immense size and structural complexity of genomes spanning many orders of magnitude has always imposed huge experimental challenges. Thus, the higher-order architecture has been and still is widely discussed with many interesting details yet to be described. Already how nucleosomes are spaced, positioned, remodelled, and whether and how nucleosome chains fold into fibres at physiological salt concentrations have been matters of continuing debate: e.g. Finch and Klug [8] proposed a relatively regular solenoid and *in vivo* neutron scattering experiments revealed a fibre diameter of 30 ± 5 nm as a dominant nuclear feature [9–12]. In contrast more recent work suggested no compaction at all (rev. [13, 14]), and highly polymorphic, nucleosome position- [15] and dynamic function-dependent structures [16, 17], which are essential to explain nucleosome concentration distributions [18–20], or dynamic and functional properties such as the nuclear diffusion of macromolecules. Moreover, the fine-structured multi-scaling long-range correlation behaviour of the DNA sequence also predicts a compacted chromatin fibre [21–24]. With a novel chromatin interaction technique—*T2C*—we were, however and indeed, able to show that nucleosomes form in general a quasi-fibre with a differential compaction of ~5 ± 1 nucleosomes/11 nm [25, 26], which is in agreement with a novel *in vivo* fluorescence correlation spectroscopy

The higher-order chromatin architecture has been a matter of even greater debate: Pioneering light microscopy studies by Rabl [28] and Boveri [29] hinted towards a hierarchical self-similar, territorial organization. Electron microscopy suggested a more random To further distinguish between the different architecture proposals, proximity crosslinking techniques (developed and used already in the last century) were further developed into a family of interaction capture techniques such as 3C [56, 57], 3C-qPCR [58], 4C [59], 3C-seq/4Cseq [60], 5C [61], and Hi-C [62]. They once more confirmed the existence of looping and subchromosomal domains, now inconsistenly referred to as topologically associated domains (TAD; [63]) with a somewhat higher localization accuracy when compared to FISH. These approaches also led to a number of - although by the underlying (raw) data basically unsupported - conjectures (Imam et al., in preparation), e.g. the fractal globule model [62], the loop array architecture of mitotic chromosomes [64], and the highly dynamic loop formation based on single-cell experiments [65] or in a genome wide assay [66]. In contrast, with the introduction of targeted chromatin capture *T2C* [25, 67–69], we were able to show that the chromatin quasi-fibre forms small stable loops of ~30-100 kbp which form stable multi-loop aggregates/ rosettes connected by linkers of similar sizes as the loops [25, 26]. The development of our novel *in vivo* FCS approach came to the same conclusion [27].
