How Does Stress Induce Cellular Adaptation: Can Epigenetic Alterations Influence Transcription and Translation?

By Stuart P. Atkinson

Observed changes in H3K4me3 deposition around gene TSS in normal versus hypoxic conditions. From figure four of Watt et al..

How Does Stress Induce Cellular Adaptation: Can Epigenetic Alterations Influence Transcription and Translation?

Among the range of stressors facing tumor cells, insufficient oxygen levels (hypoxia) can lead to enhanced tumor cell plasticity and cancer stem cell-like phenotypes through distinct types of metabolic restriction, which prompt an increase in cancer aggressiveness and therapeutic resistance. But what mechanisms link hypoxia to these outcomes?

Low oxygen levels induce significant transcriptomic and epigenetic alterations (Prickaerts et al., Chakraborty et al., and Batie et al.), including the accumulation of the permissive H3K4me3 histone modification at gene transcription start sites (TSSs). Stressors such as hypoxia also reprogram the mRNA translation machinery to selectively translate transcripts encoding regulators of stress responses (Wouters & Koritzinsky). Notably, these mRNAs possess 5′ untranslated regions (5′UTRs) that contain features that can facilitate their translation under hypoxia (Guan et al. and Jewer et al.), suggesting that changes in 5′UTR composition may drive changes in protein synthesis in response to stressors. Importantly, most genes have multiple TSSs, whose differential implementation linked to tissue-specificity and disease (Forrest et al., Haberle et al., and Qamra et al.).

To deeply explore the consequences of hypoxia in cancer cells, researchers from the labs of Ola Larsson (Karolinska Institutet) and Lynne-Marie Postovit quantified hypoxia-dependent transcriptomes (total RNA), epigenomes, and translatomes in breast cancer and human embryonic stem cells. As reported in Nature Cell Biology, the authors revealed that hypoxia reprograms gene expression by coordinating epigenetic, transcriptional, and translational programs. Overall, they describe H3K4me3-based "TSS switching" as a hypoxia-induced gene expression control mechanism that generates 5′UTR isoforms of mRNAs with distinct translation efficiencies, thereby establishing an adaptive translational landscape (Watt et al.).

Paired-Tag technology from Epigenome Technologies 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. Implementing this technology to more deeply study hypoxia-induced epigenetic mechanisms could reveal the interplay among multiple histone modifications in TSS switching and underscore the importance of the H3K4me3-based control of alternate TSS selection in specific cell subsets during cancer development as they adapt to stress, including treatment administration.

Hypoxic conditions alter transcription, translation, and TSS selection of mRNA species; TSS switching is related to translation changes: genes showing TSS switching are more likely to show changes in translation compared to normoxic conditions. From figure 2 of Watt et al..

H3K4me3 Coordinates Adaptive Translational Responses by Inducing TSS Switching and 5′UTR Remodeling

The authors first investigated hypoxia-associated 5′UTR isoform dynamics using nanoCAGE sequencing on total mRNA isolated from breast cancer and embryonic stem cells, revealing extensive 5′UTR remodeling via hypoxia-inducible factor 1 (HIF1 a transcription factor that responds to hypoxia)-independent hypoxia-induced TSS switching. The subsequent analysis of translational reprogramming revealed that hypoxia-induced TSS switching significantly altered the translatome by modulating a range of 5′UTR features that drive hypoxia-induced changes in translation efficiency. Even modest changes to 5′UTR length significantly altered translation under hypoxic conditions, demonstrating the impact of this regulatory mechanism. Furthermore, alterations in the hypoxia-induced 5′UTR landscape occurred alongside the reprogramming of the translational apparatus, thereby promoting a survival-enhancing adaptive translational program.

The authors next employed ChIP-seq and ATAC-seq to establish a connection between altered H3K4me3 levels and alterations in nucleosome context in response to hypoxia-induced TSS switching. They observed associations between directional shifts in H3K4me3 distribution and the selection of TSSs that result in longer/shorter 5′UTRs, and found evidence that pre-existing chromatin states play crucial roles in determining cell-type-specific stress-responsive TSS switching. Interestingly, they also discovered that inhibiting the H3K4 demethylase KDM5 (Howe et al.) (which increases H3K4me3 levels) induced TSS switching and altered the proteome without significant changes in mRNA abundance. As such, these findings demonstrate that TSS-switching-dependent 5′UTR remodeling can impact the proteome independently of changes in mRNA levels. Meanwhile, lowering H3K4me3 levels by inhibiting MLL-containing COMPASS methyltransferase activity blocked TSS switching and decreased cellular fitness under hypoxic conditions. Overall, these findings provide evidence that hypoxia-induced H3K4me3 alterations can drive TSS-switching events, identifying a previously unknown role for this histone modification in 5′UTR-determining TSS selection and the adaptation of cells to hypoxia. Notably, these changes coincide with shifts in nucleosome occupancy and positioning, suggesting H3K4me3 modifications may drive TSS selection concomitant with changes in nucleosome conformations.

Finally, the authors examined whether H3K4me3-mediated TSS switching regulated specific biological processes in response to hypoxia. Overall, an analysis of hypoxia-induced TSS-switching events revealed a link to metabolism-associated genes such as PDK1(a HIF1 target), which encodes a glycolytic enzyme required for the switch from oxidative phosphorylation to glycolysis under hypoxic conditions (Kim et al.). Hypoxia increased PDK1 protein and transcript levels, which correlated with the gain of a shorter, more efficiently translated 5′UTR mRNA PDK1 isoform . In this case, the extension of H3K4me3 around the TSS accompanied the expression of the shorter PDK1 5′UTR isoform, which prompted alterations to nucleosome conformations and, as such, altered TSS selection. Overall, the data from this fascinating study propose a model by which chromatin modification coordinates adaptive translational responses that drive cellular phenotypes through TSS switching and 5′UTR remodeling.

H3K4me3 deposition, nucleosome positioning, and TSS sites under normal (red), hypoxic (blue), DMSO (green) and C48 (purple) treated conditions. From figure 8 of Watt et al..

H3K4me3-mediated TSS Switching and 5′UTR Remodeling: An Unappreciated Translational Regulation Mechanism

Overall, the authors of this fascinating study reveal that hypoxia-induced alterations to H3K4me3 distribution alter TSS selection to establish an adaptive translatome through 5′UTR remodeling, suggesting that this mechanism adjusts the abundance of "strong" versus "weak" 5′UTR isoforms to remodel the proteome.

What other stressors are sensed at the chromatin level to direct downstream effects, and could the implementation of Paired-Tag technology from Epigenome Technologies help to answer this question? Epigenome Technologies Blog to find out!