The profound structural rearrangements that chromatin must undergo during the cell cycle underscores one aspect of the dynamic nature of chromatin. There is, in addition, compelling evidence of an important role for local structural changes in compaction and/or position of specific genetic loci likely to be of critical importance in their transcriptional regulation. In fact, most of the functionally important characteristics of chromatin show dynamic behavior in the sense of time-dependent changes. These include the mobility of many chromatin-binding components, including chromatin architectural proteins (Phair and Misteli, 2000
), and the status of posttranslational modifications, especially of the core histones. Many of these phenomena are likely to impact local and global chromatin conformation and thus modulate higher-order structure.
The study of chromatin dynamics in all of its manifestations has opened up exciting new perspectives—indeed, a literature search reveals over 60 reviews with titles containing the terms “chromatin” and “dynamics” (e.g., Huebner and Spector 2010
). Here, we focus on aspects of dynamics that are likely to impact higher-order structures. All involve changes in location and/or shape of specific genetic loci that are thought to have important functional implications.
As the field of chromatin dynamics has developed, it has become increasingly clear that a number of underlying physical and molecular phenomena are involved, and that to understand them fully, information at many different levels is required. For a given locus these may include the temporal and spatial scales of observed movements, their energy dependence, and their location, both within the nucleus and within their specific chromosome territory. One important goal of current research is to be able to relate these changes in location/shape to the underlying physical changes in chromatin higher-order structure. For movements that are correlated with changes in transcriptional activity, it is also important to determine whether a change in location is a prerequisite for, or a consequence of, the altered level of RNA synthesis.
Advances in light microscopy techniques over the past ~25 years have allowed chromatin dynamics to be examined with steadily improving temporal and spatial resolution, and has enabled three principal types of dynamic change to be recognized (Soutoglou and Misteli 2007
). These differ in the extent of motion, their energy dependence, and the time scales involved (). Early studies tracking changes in location of large segments of chromatin and whole chromosome territories indicated that, when corrected for nuclear rotation, the regions were essentially immobile over distances >0.4 µm (Cremer et al. 1982
, Diboni and Mintz 1986
, Shelby et al. 1996
). However, more recent work combining higher spatial resolution with in vivo labeling of smaller defined segments of chromatin, revealed a more complex scenario with different regions of chromatin showing very different mobilities. For example, a segment of budding yeast chromatin located near a centromere was found to show constrained random walk diffusive motion with a confinement radius of ~0.3 µm (Marshall et al. 1997
), that was independent of the metabolic state of the cell. The rate of mobility of this region of chromatin was calculated to be approximately three orders of magnitude lower than expected for free DNA of similar length (Marshall et al. 1997
), leading to the suggestion that the locus was tethered within the nucleus, perhaps to the nuclear envelope or some internal structure. A detailed study of chromatin mobility in Drosophila
spermatocytes using loci labeled with the lac repressor system recorded relatively large movements during short time intervals in early nuclei. Movements of this magnitude would be expected to result in displacements of over 4 µm in 1 h and cover the entire 11–12 µm diameter of a nucleus in 6–7 h (Vazquez et al. 2001
). However, when long time periods were examined, it was clear that loci were constrained within a volume approximately equivalent to a chromosome territory. Another important finding was that spermatocytes in late G2 stage and approaching meiosis showed greatly reduced chromatin mobility, suggesting that some form of additional tethering precedes chromosome condensation.
A comparison of the mobility of different chromosomal sites in yeast also revealed striking locus- and cell cycle-dependent differences in chromatin dynamics (Heun et al. 2001
). Some chromatin regions showed occasional large (~0.5 µm) movements over time periods as short as 10 seconds that were inhibited in ATP-depleted cells and thus dependent on the metabolic state of the cell. The fact that the magnitude of these large movements is similar for organisms with widely different nuclear volumes has important structural and functional implications. For example, in budding yeast, which has a nuclear diameter of ~2 µm, movement within a radius of ~0.5 µm would allow access to a large portion of the nuclear volume. In contrast, in a typical mammalian nucleus of ~10 µm in diameter this motion would explore only a thousandth of the nuclear volume. In yeast, the effectively large displacements of loci within nuclei may promote the observed high recombination frequency, which may require substantial intermingling among different regions of the genome (Gasser 2002
Observations of “free” ectopic regions of chromatin, exemplified by large (~15 kb) circular plasmids in yeast underscore the context dependence of chromatin motion. An early study using a centromere-containing plasmid reported very limited movements, similar to those observed for centromeric regions in intact chromosomes (Marshall et al. 1997
). However, more recently, Gartenberg et al. (2004)
using plasmids designed to be either transcriptionally silent or competent, found that the active plasmids showed unconstrained movement, whereas the silent ones were strongly constrained and preferentially located near the nuclear envelope. Also relevant to the factors controlling intranuclear mobility is the finding that whereas double-stranded DNA breaks tend to stay together through the repair process (Kruhlak et al. 2006
), the absence of a critical repair factor leads to long-range movements of the broken ends (Downs et al. 2004
A general pattern emerging from numerous studies using different organisms is that all chromatin loci show constrained Brownian motion with rather similar diffusion constants, but a wide range of confinement volumes. A strong relationship between the confinement volume, and distance from the nuclear envelope has also been established, with loci closer to the envelope being more constrained. The periphery of the nucleus proximal to the nuclear envelope is considered a transcriptionally repressive environment, establishing a correlation between transcriptional silencing and limited intranuclear mobility (Marshall 2002
). The underlying mechanism for this correlated behavior is not clear, but there is evidence that the constrained motion results from physical tethering to the nuclear lamina (Heun et al. 2001
; Chubb et al. 2002
; Gartenberg et al. 2004
The higher spatial resolution (~20 nm) of two-photon microscopy, together with a temporal resolution of ~30 msec has revealed new levels of complexity for constrained diffusive motion of chromatin (Levi et al. 2005
). In these experiments, GFP-labeled repeats inserted in CHO cells showed periods of rapid constrained diffusive motion alternating with occasional energy-dependent curvilinear leaps of ~150 nm that lasted only 0.3–2.0 second. The energy-dependence of these leaps suggested that they may need the activity of chromatin remodeling complexes or other ATP-requiring changes in chromatin organization that could lead to events such as the decondensation of compact 30-nm fibers resulting in the rapid changes in location. Another important outcome of this study pertinent to the extent of physical coupling between neighboring loci on the same chromosome was the finding that loci 1–2 µm apart in the nucleus moved independently (Levi et al. 2005
). A similar conclusion was reached by Hu et al. (2009)
who showed that for a ~2 Mb region of chromatin that appeared as a linear cluster of beads in the light microscope, the individual beads moved independently. These findings suggest that neighboring loci are not structurally tied together by, for example, being anchored to an underlying nonchromatin structure. As it becomes possible to study yet closer regions of chromatin, there will be a point at which movement of the neighbors is correlated, and defining this point will be very useful in being able to infer the effective chromatin compaction level. Thus, although the accumulating data clearly demonstrate that chromatin in the interphase nucleus undergoes constant dynamic reorganization through constrained diffusive motion, the level(s) of chromatin higher-order structure that participate in this phenomenon remain to be determined.
Some hints of the mechanism whereby loci undergo directed movement have emerged from work on an inserted transgene that typically resides in peripheral heterochromatin, but tends to relocate toward the center of the nucleus within 1–2 hour of transcriptional up-regulation (Chuang et al. 2006
). Importantly, this movement was blocked by the expression of a mutant nuclear myosin I and by a nonpolymerizable mutant of actin. This intriguing finding should be viewed in the context that several families of actin and actin-related proteins (ARPs) are found in the nucleus, many of which are components of chromatin remodeling and histone modification complexes (Chen and Shen 2007
). Also, it has been shown that the BAF remodeling complex binds to the ends and branch points of actin filaments in a PIP2-dependent manner (Rando et al. 2002
). Unlike their cytoplasmic counterparts, nuclear actins and ARPs tend to occur as monomers rather than the filamentous structures, and it is therefore curious that it was an actin polymerization mutant that was defective in long range movement (Chuang et al. 2006
). Determining the mechanism by which actin/myosin influences the mobility and directionality of loci is clearly an important future goal.