Nuclear pore density controls heterochromatin reorganization during senescence

 

Context

In non-transformed cells, excessive oncogene expression has long been known to cause oncogene-induced senescence (OIS). This cellular state is marked by several features, including permanent proliferative arrest, a displacement of the normally peripherally located heterochromatin to several distinct internal areas (Senescence-Associated Heterochromatic Foci, SAHF), and an elevated secretion of immune signaling molecules (SASP, the Senescence Associated Secretory Phenotype).

Recent findings suggest that the peripheral localization of mostly inactive heterochromatin is established by an active tethering mechanism of the chromatin to the nuclear lamina¹, thought to protect the nuclei from mechanical assault². On the contrary, internal heterochromatin is proposed to be the default state of its organization, as proposed from observations of chromatin architecture in nocturnal animal eye rod cells^{3, 4}. Indeed, it is hypothesized that the peripheral heterochromatin of most cells is maintained by a careful balance of repulsing and attracting forces from and to the nuclear envelope. Additionally, to maintain a stable flux of molecules between the nucleus and the cytoplasm, the nuclear envelope contains numerous nuclear pore complexes (NPCs). At these large multiprotein complexes, the heterochromatic fibers are excluded, largely through the repulsing action of the Translocated Promoter Region protein (TPR) component of the NPC⁵.

Although it has been proposed that the induction of SAHF is in part due to reduced interactions of heterochromatin with the nuclear periphery through, among others, lower levels of Lamin B1⁶, the role of NPCs, and potentially their repelling forces in establishing SAHF is unknown. Moreover, the order of phenotypic manifestations of OIS is not well understood, i.e. it is unclear if the SASP is required for SAHF formation and cell cycle arrest, or if cell cycle arrest is required for SAHF.

 

The preprint

In this preprint, the authors hypothesized that OIS would lead to an increase in NPCs, thus promoting the establishment of internal heterochromatic foci. During differentiation, normal quiescent cells downregulate their nucleoporin mRNA expression to stabilize the number of NPCs. In contrast, using an inducible constitutively active Ras overexpression system leading to OIS in non-transformed lung fibroblasts, the authors show that the mRNA levels of NPC components are stabilized, resulting in an increased number of NPCs. By further manipulating the levels of either an internally located component (POM121) or the nuclear interior-facing protein TPR, they find that the increase in NPCs is required for the establishment of the senescence associated heterochromatic foci. Consequently, the authors show that knockdown of the NPC not only prevents SAHF but also abrogates the senescence associated secretory phenotype, arguing that the correct order of events is cell cycle arrest, followed by SAHF and then SASP. The requirement of SAHF for SASP was further strengthened by knocking down a chromatin chaperone preventing SAHF formation (ASF1). Indeed, with this experiment they could effectively halt the secretory phenotype upon SAHF disruption, while the number of NPCs remained the same.

In conclusion, heterochromatic focus formation in oncogene induced senescence requires an increased number of nuclear pore complexes, leading to the sequential induction of two of its main phenotypic manifestations, SAHF and SASP. This process is proposed to function through an increased repulsing force of the chromatin by the higher numbers of NPCs.

 

My opinion

I like this preprint for several reasons. First and foremost, the study is straightforward, has a clear rationale and hypothesis, and confers simple but on-point experiments to address it. A second reason is that the findings provide a major advancement in the thinking of how, in addition to the lamina, the NPC helps the nucleus organize into mainly peripheral heterochromatin and central euchromatin. It allows us to ask what the evolutionary drives and functional advantages are for this organization. Furthermore, the study proposes an almost effortless mechanism by which this internal SAHF occur in the cell – especially if the hypothesis on repulsing and attracting forces turns out to be correct. Moreover, in line with these observations, recent work has suggested that heterochromatin forms a phase-separated compartment in the nucleus⁷. It would be interesting to further study in this context how the heterochromatin upon establishment of SAHF behaves on the physicochemical level, and to what extent NPCs influence these behaviors.

Considering the recent findings on heterochromatin organization in the rod cells of nocturnal animals, I also wonder what the role of the nuclear envelope and its interactions with the chromatin are in these cell types, and if the NPC may also contribute to the nuclear architecture in this specific context. Additionally, I am curious to learn more about the hypothesized forces that drive this organization both in senescent cells and in normal circumstances. One might ask, in light of the findings of this preprint, how NPCs contribute to general chromatin architecture during differentiation, proliferation, etc.

 

References

1 Lamina-Associated Domains: Links with Chromosome Architecture, Heterochromatin, and Gene Repression. Van Steensel B and Belmont AS, (2017) Cell 169, 780-791.

2 The tethering of chromatin to the nuclear envelope supports nuclear mechanics. Schreiner et al. (2015) Nature Communications (6), 7159

3 Nuclear architecture of rod photoreceptor cells adapts to vision in mammalian evolution. Solovei et al. (2009). Cell 137, 356–368.

4 Heterochromatin drives organization of conventional and inverted nuclei. Falk et al. (2018) bioRxiv, https://doi.org/10.1101/244038 – preLighted by Boyan Bonev here

5 Protein Tpr is required for establishing nuclear pore‐associated zones of heterochromatin exclusion. Krull et al. (2010) Embo J: 29 (10).

6 Redistribution of the Lamin B1 genomic binding profile affects rearrangement of heterochromatic domains and SAHF formation during senescence. Sadaie et al. Genes Dev. (2013) ;27(16):1800-8

7 The Role of Phase Separation in Heterochromatin Formation, Function, and Regulation. Larson et al. (2018) Biochemistry. 57, 2540−2548

 

doi: https://doi.org/10.1242/prelights.3946

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