The coordination of terminal differentiation and cell cycle exit is mediated through the regulation of chromatin accessibility

 

Background

The development of complex animal tissues requires the finely tuned expression of thousands of regulatory genes. Changes to underlying chromatin landscape are required to mediate the changes in gene expression necessary for fully differentiated tissues to arise. Consequently, chromatin landscapes are dynamic during development. Whilst it is commonly accepted that accessible or “open” chromatin is associated with regulatory elements that control gene expression, the full relationship between chromatin accessibility and development is incompletely understood.

Much work has been done on understanding the developmental programs that drive differentiation, however the question of how development is brought to a halt at the chromatin level (i.e. terminal differentiation and cell cycle exit) is less well understood. In this preprint, Ma et al. investigate the chromatin changes that accompany wing differentiation in Drosophila and uncover some surprising findings regarding the open chromatin at cell-cycle gene loci.

 

Key findings

Chromatin accessibility undergoes extensive changes concurrent with altered gene expression during wing development

Ma et al. initially set out to characterise the gene expression and chromatin accessibility profiles of Drosophila wing tissue at various timepoints during development. To achieve this, they conducted RNA-seq and FAIRE-seq at 6 developmental timepoints in the larval and pupal stages (with the final stage comprising post-mitotic differentiated cells). Unsurprisingly, the authors discovered both highly dynamic gene expression and chromatin accessibility landscapes during wing differentiation. Gene expression was correlated to chromatin accessibility and the majority of accessibility changes were observed to be associated with enhancers becoming activated. Importantly, cell-cycle genes were seen to have dynamic regions of open chromatin that correlated with known cell-cycle changes in the developing wing. The critical cell-cycle regulators CycE, stg and e2f1 all exhibited regions in which loss of chromatin accessibility was observed at cell-cycle exit.

 

Disruption of cell cycle has little impact on chromatin accessibility of cell cycle genes

Having confirmed that chromatin accessibility is correlated with cell-cycle gene expression at cell-cycle exit, the authors sought to understand whether the observed closing of chromatin was a function of cell-cycle exit, or a precursor to it. To answer this question cell cycles were artificially extended through the overexpression of E2F alone or E2F with cyclinD, resulting in either one extra cell division or several rounds of mitoses, respectively. Intriguingly, when wing tissue was profiled by RNA-seq and FAIRE-seq under these conditions, surprisingly few changes to chromatin accessibility were observed, despite extensive changes to gene expression. When several critical cell cycle regulator gene loci were examined in more detail in ectopically cycling tissue (e.g. CycE, stg), chromatin accessibility at putative enhancers was almost indistinguishable from postmitotic cells at the same timepoint (i.e. enhancers continue to be rendered inaccessible despite continued expression of the gene – see figure). This unusual finding suggests that the closing of chromatin is regulated by developmental cues that are not directly linked to the cell-cycle.

 

 

Cell cycle disruption impacts accessibility of chromatin at terminal differentiation genes

Whilst changes to chromatin accessibility were not observed at cell cycle genes, the authors went on to show that a subset of genes involved in terminal differentiation displayed a loss of open chromatin when cell cycle exit was delayed compared to controls. These regions largely represented enhancers that were seen to open following cell-cycle exit rather than open regions that became closed as a result of continued cell-cycling. In contrast to the cell cycle genes, this correlated with changes in expression and ultimately resulted in cuticle formation phenotypes.

 

Why I like this preprint

I found this preprint appealing due to the comprehensive approach the authors have taken to characterise gene expression and chromatin accessibility, resulting in a window into wing development with impressive temporal resolution.

The finding that chromatin accessibility and cell-cycle can be decoupled is surprising and seems to fly in the face of conventional thinking. This new perspective may force us to rethink our ideas about how terminal differentiation is achieved.

 

Questions for the authors

If chromatin accessibility changes occur independently of cell cycles and are not sufficient to disrupt expression (i.e. CycE is expressed highly in bypassed G0 despite loss of chromatin accessibility – Fig4D), then what is the purpose of the chromatin closing?

Is it possible that the key enhancers for the expression of cell-cycle genes are located distally and act at long range, so are difficult to detect? (i.e. the proximal open chromatin regions observed to be unaffected are redundant with long range enhancers?)

Do you expect that there are changes occurring to histone modifications at cell-cycle loci at which chromatin accessibility appears unaffected by continued cell-cycles?

Do you have any ideas about how the developmental closing of cell cycle genes (i.e. CycE, stg) chromatin is programmed if not directly linked to cell cycle, and do you expect that this will be a conserved mechanism across cell types?

 

Tags: chromatin, development, drosophila

Posted on: 7 January 2019

doi: https://doi.org/10.1242/prelights.6812

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