Bulk chromatin motion has not been analyzed at high resolution. We present HiD , a method to quantitatively map dynamics of chromatin and abundant nuclear proteins for every pixel simultaneously over the entire nucleus from fluorescence image series. HiD combines reconstruction of chromatin motion and classification of local diffusion processes by Bayesian inference. We show that DNA dynamics in the nuclear interior are spatially partitioned into 0.3-3-μm domains in a mosaic-like pattern, uncoupled from chromatin compaction. This pattern was remodeled in response to transcriptional activity. HiD can be applied to any dense and bulk structures opening new perspectives towards understanding motion of nuclear molecules.

The histone variant macroH2A1.1 plays a role in cancer development and metastasis. To determine the underlying molecular mechanisms, we mapped the genome-wide localization of endogenous macroH2A1.1 in the human breast cancer cell line MDA-MB 231. We demonstrate that macroH2A1.1 specifically binds to active promoters and enhancers in addition to facultative heterochromatin. Selective knock-down of macroH2A1.1 deregulates expression of hundreds of highly active genes. Depending on the chromatin landscape, macroH2A1.1 acts through two distinct molecular mechanisms. The first mitigates excessive transcription by binding over domains including the promoter and the gene body. The second stimulates expression of RNA Polymerase II (Pol II) paused genes, including genes regulating mammary tumor cell migration. In contrast to the first one, macroH2A1.1 specifically associates with the TSS of Pol II paused genes. These processes occur in a predefined local 3D genome landscape but do not require rewiring of enhancer-promoter contacts. We thus propose that macroH2A1.1 serves as a transcriptional modulator with a potential role in assisting the conversion of promoter-locked into a productive and elongating Pol II.