Condensed mitotic chromatin is accessible to transcription factors and chromatin structural proteins

Abstract

During mitosis, chromosomes are highly condensed and transcription is silenced globally. One explanation for transcriptional repression is the reduced accessibility of transcription factors. To directly test this hypothesis and to investigate the dynamics of mitotic chromatin, we evaluate the exchange kinetics of several RNA polymerase I transcription factors and nucleosome components on mitotic chromatin in living cells. We demonstrate that these factors rapidly exchange on and off ribosomal DNA clusters and that the kinetics of exchange varies at different phases of mitosis. In addition, the nucleosome component H1c-GFP also shows phase-specific exchange rates with mitotic chromatin. Furthermore, core histone components exchange at detectable levels that are elevated during anaphase and telophase, temporally correlating with H3-K9 acetylation and recruitment of RNA polymerase II before the onset of bulk RNA synthesis at mitotic exit. Our findings indicate that mitotic chromosomes in general and ribosomal genes in particular, although highly condensed, are accessible to transcription factors and chromatin proteins. The phase-specific exchanges of nucleosome components during late mitotic phases are consistent with an emerging model of replication independent core histone replacement.

Translocation of histone H1 subtypes between chromatin and cytoplasm during mitosis in normal human fibroblasts

“…In contrast to our findings, H1.2-GFP was solely located on chromatin during the cell cycle in transfected HeLa cells (34) and NIH 3T3 cells (32). Possibly, there are differences between the binding and phosphorylation of H1.2-GFP and the native H1.2 but the most likely explanation for this difference is that the residence time for H1.2 bound to chromatin is too short to become captured by formaldehyde crosslinking during fixation in accordance with recent results (35).…”

“…Alterations in chromatin binding characteristics between the native protein and its GFP-labeled counterpart have recently been recognized in HMGN proteins (36). In addition, the rate of H1.2-GFP recovery after photobleaching was higher at metaphase compared with prophase, anaphase or telophase, indicating that the rate of dynamic exchange of H1.2-GFP increased during metaphase (32), in line with our present results.…”

“…Transcription factors and chromatin proteins have been shown to have access to mitotic chromosomes (32). Histone H1.2 has previously been shown to have the highest turnover rate compared with the other H1 subtypes.…”

Chromatin Remodeling Complexes Interact Dynamically with a Glucocorticoid Receptor–regulated Promoter

“…In fact, this modulation is one of the key mechanisms essential for the functional role of nuclear hormone receptors (Stenoien et al, 2001;Schaaf and Cidlowski, 2003;Stavreva et al, 2004;Elbi et al, 2004b;Farla et al, 2005;Rayasam et al, 2005). In vivo FRAP has been used to study the dynamic properties of chromatin proteins and that the FRAP recovery kinetics of chromatin proteins are directly related to their chromatin-binding properties (Lefebvre et al, 1991;Fragoso et al, 1998;Lever et al, 2000;Kimura and Cook, 2001;Kimura et al, 2002;Maruvada et al, 2003;Phair et al, 2004;Becker et al, 2005;Chen et al, 2005). Because BRG1 and BRM were selectively recruited to the MMTV array in response to hormone (Figures 2 and 3), we used FRAP to study the binding kinetics of chromatin-remodeling complexes at the amplified MMTV target in vivo.…”

Nuclear Reprogramming and Mitosis – how does mitosis enhance changes in gene expression?

“…This may be due to shifting regulatory molecules and exposing transcription factor binding sites on DNA. It has been demonstrated that although mitotic chromatin is condensed, transcription factors are still able to bind to their target sites 15 and this may provide a hitherto untested explanation for the enhancement of reprogramming associated with mitotic transition.…”