Secretory-senescence associated Enhancer Chromatin Remodeling

By Stuart P. Atkinson

Pioneer Transcription Factors Regulate the Accessibility of SASP-associated Gene Enhancers in Senescent Cells: Researchers describe how the pioneer transcription factors Foxf1/2 regulate chromatin accessibility at SASP-associated gene enhancers in senescent cells.

The Importance of the Epigenetic Regulation of Enhancer Elements in Senescent Cells

The pro-inflammatory senescence-associated secretory phenotype or "SASP" represents a hallmark of irreversibly cell-cycle-arrested senescent cells. While the SASP does play a beneficial role during tissue homeostasis and regeneration (Demaria et al., Reyes et al., and Grosse et al.), the pro-inflammatory SASP also exacerbates age-related disorders through chronic inflammation (Gorgoulis et al. and D'Ambrosio & Gil). At the epigenetic level, the deposition of the permissive H3K27ac histone modification at gene enhancers - regulatory genomic elements that activate the transcription of target genes and drive cell-type-specific gene expression (Nott et al. and Calo & Wysocka) - represents a critical regulatory mechanism controlling the gene regulatory network underlying the pro-inflammatory SASP (Etoh et al. and Tasdemir et al.). Could additional epigenetic insights into SASP-specific molecular mechanisms deepen our understanding of senescence and even have therapeutic value regarding the improved clinical management of an increasingly aged society?

The DNA-binding activity of Forkhead box (FOX) proteins - a superfamily of transcription factors that play key roles in development and adult tissue homeostasis (Lam et al.) - promotes chromatin accessibility (Obsil & Obsilova); furthermore, FOX proteins function as pioneer transcription factors - initiating the establishment of de novo enhancers (Iwafuchi-Doi & Zaret, Wolf et al., Adam et al., and Sinha et al.) and initiating chromatin opening - to determine cell fate (Lee et al., Wang et al., and Matsui et al.). Could FOX family transcription factors play integral roles in defining senescence-specific enhancer landscapes?

CUT&Run fragment heatmaps, ATAC accessibility volcano, and TF motif enrichment
p300/CBP knockout oblates histone acetylation in senescent cells, and results in an overall closure of chromatin - predominanty at FOX-family binding sites. From Etoh et al.

In their new Cell Stem Cell study, researchers led by Kan Etoh and Mitsuyoshi Nakao (Kumamoto University) now reveal that the FOXF1/2 transcription factors help define the senescence-specific enhancer landscape that drives the pro-inflammatory SASP by regulating chromatin accessibility (Etoh et al.). In brief, FOXF1/2 function as pioneer transcription factors that recruit the p300/CBP H3K27ac histone acetyltransferases to shape the enhancer landscape in senescent cells, thereby regulating the transcriptional activation of SASP genes, in combination with the AP-1 transcription factor member c-JUN. The authors hope that these exciting findings may pave the way towards the development of therapeutics that may promote healthy aging and/or combat aging- and chronic inflammation-associated conditions.

Paired-Tag technology from Epigenome Technologies generates joint epigenetic and transcriptomic profiles at single-cell resolution and detects histone modifications and RNA transcripts in nuclei with efficiency comparable to single-nucleus RNA-seq/ChIP-seq assays. Could the in-depth simultaneous analysis of histone modification and transcriptomic profiles in single senescent cells via the integration of Paired-Tag technology help to reveal more regarding the enhancer landscapes in senescent cells and perhaps identify additional therapeutic targets?

FOXF1/2 and p300/CBP Bind and Activate Pro-inflammatory SASP-associated Gene Enhancers

CUT&Run volcano plot, scatter plot of RNA vs C&R logFC, and peak enrichment plots
FOXF1 binding is significantly increased in senescent vs non-sensecent cells, and changes in FOXF1 occupancy strongly correlate with changes in accessibility, H3K27ac deposition, and expression. From Etoh et al.

To tease apart the contrbution of chromatin regulatory state to gene expression, the authors defined a quantitative measure - Regulatory Potential - that aggregates the influence of multiple nearby regulatory elements into a single expression influence score. This approach confirms that changes to the local enhancer landscape correlated with gene expression. Notably, H3K27ac-based regulatory potential was far more effective at predicting expression differences than basing regulatory potential on open chromatin, highlighting the importance of chromatin state (not merely openness) on gene regulation.

Expression barplot, boxplot for several genes, cartoon of RP calculation, and rank-probability curves.
Senescent cells express a number of proinflammatory factors and lose expression of proliferative genes. FOXF1 is highly expressed in these cells, and knockout reverses the inflammatory senescence signature gene expression. Regulatory Potential can identify altered changes in expression particularly well using H3K27ac deposition at local enhancers. From From Etoh et al.

The study further revealed that FOXF1/2 interacted with and recruited p300/CBP to de novo enhancers associated with pro-inflammatory SASP-associated genes following senescence induction, thereby increasing H3K27Ac levels and, consequently, chromatin accessibility. In addition, the authors underscored the importance of recruiting AP-1 c-JUN to these regions alongside FOXF1/2. Interestingly, FOXF1/2 depletion in senescent cells significantly suppressed the pro-inflammatory SASP (independent of growth arrest) and diminished the paracrine effects known to affect surrounding cells; furthermore, FOXF1/2 loss prompted c-JUN redistribution to regulatory elements (predominantly lacking FOXF1/2 DNA-binding motifs) associated with senescence-associated genes that normally undergo transcriptional downregulation. Overall, these exciting results indicated that FOXF1/2 and c-JUN function cooperatively to establish enhancer landscapes in senescent cells.

Chromatin Remodeling at Enhancers: A Therapeutic Target to Support Healthy Aging?

This exciting study highlights the general importance of the FOXF1/2 pioneer transcription factors in inducing the pro-inflammatory SASP in senescent cells via the recruitment of histone acetyltransferases to relevant gene enhancers and mediating improved chromatin accessibility; therefore, the authors suggest that FOXF1/2-mediated enhancer remodeling may represents a target for therapeutics (such as senostatics and senomorphics) that may reduce the detrimental impact of the pro-inflammatory SASP while maintaining senescent cells to promote healthy aging and help combat diseases associated with aging and chronic inflammation.

The study does detail additional research aims that may strengthen their claims further, which include: i) the use of more complex models that capture the full complexity of in vivo senescence observed during pathophysiological aging; ii) the exploration of the mechanism by which senescence induction activates FOXF1/2; iii) the further dissection of the pioneering activity of FOXF1/2; and iv) the identification of the precise mechanism underlying the redistribution of c-JUN to the regulatory elements of senescence-associated downregulated genes after FOXF1/2 depletion.

The implementation of Paired-Tag technology from Epigenome Technologies, which generates joint epigenetic and transcriptomic profiles at single-cell resolution and detects histone modifications and RNA transcripts in individual nuclei with efficiency comparable to single-nucleus RNA-seq/ChIP-seq assays, has the potential to provide deeper insight into such research aims. What more could the simultaneous single-cell analysis of histone modification and transcriptomic profiles tell us about the enhancer landscapes of senescent cells?