Coupling chromatin structure and dynamics by live super-resolution imaging

Abstract

<jats:p>Chromatin conformation regulates gene expression and thus, constant remodeling of chromatin structure is essential to guarantee proper cell function. To gain insight into the spatiotemporal organization of the genome, we use high-density photoactivated localization microscopy and deep learning to obtain temporally resolved super-resolution images of chromatin in living cells. In combination with high-resolution dense motion reconstruction, we find elongated ~45- to 90-nm-wide chromatin “blobs.” A computational chromatin model suggests that these blobs are dynamically associating chromatin fragments in close physical and genomic proximity and adopt topologically associated domain–like interactions in the time-average limit. Experimentally, we found that chromatin exhibits a spatiotemporal correlation over ~4 μm in space and tens of seconds in time, while chromatin dynamics are correlated over ~6 μm and last 40 s. Notably, chromatin structure and dynamics are closely related, which may constitute a mechanism to grant access to regions with high local chromatin concentration.</jats:p>

Dynamic nuclear structure emerges from chromatin crosslinks and motors

“…More specifically, Hi-C allows one to quantify the local chromatin interaction domains at the megabase scale [4]. Such domains are stable across different eukaryotic cell types and species [5]. To quantify such interactions in the simulations, one determines the number of monomers in the vicinity of the ith chain monomer.…”

“…This material is ∼ 1 meter of DNA with proteins, forming chromatin, and it is packaged across multiple spatial scales to fit inside a ∼ 10 µm nucleus [1]. Chromatin is highly dynamic; for instance, correlated motion of micron-scale genomic regions over timescales of tens of seconds has been observed in mammalian cell nuclei [2][3][4][5][6]. This correlated motion diminishes both in the absence of ATP and by inhibition of the transcription motor RNA polymerase II, suggesting that motor activity plays a key role [2,3].…”

Dynamics as a cause for the nanoscale organization of the genome

“…Given the observed relationships between structural reorganization of the genome [22,23], nuclear functions and chromatin dynamics, a lasting question in genome biology remains if, and if yes how, chromatin dynamics has an effect on genome organization in nuclear space. To tackle this question, we recently introduced Deep-PALM, a live-cell superresolution approach able to achieve sub-diffraction spatial resolution and 360 ms temporal resolution to image chromatin in vivo [24]. Using Deep-PALM, individual nucleosomes go unseen, and only clustered nucleosomes are eventually detected over the background of super-resolution images, in the form of nanometer-sized 'blobs' (see Figure 1(a)).…”

“…Each blob likely contains a limited number of nucleosomes (<30, ref. [24,25]) associating transiently within the timescale of about 1 second. The functional implications of blobs remain to be explored.…”

“…(a) Human osteosarcoma U2OS expressing H2B-PATagRFP cells were imaged. Deep-PALM combines the predictions of a deep learning algorithm from a subset of 12 images to reconstruct a super-resolved image of chromatin with a temporal resolution of 360 ms at 63 nm spatial resolution [24]. Segmentation of chromatin blobs and Optical Flow analysis allows ascribing a nearest neighbor distance (NND), blob area and flow magnitude to each blob in each frame individually.…”