Compromised nuclear envelope integrity drives tumor cell invasion

Abstract

While mutations leading to a fragile envelope of the cell nucleus are well known to cause diseases such as muscular dystrophies or accelerated aging, the pathophysiological consequences of the recently discovered mechanically induced nuclear envelope ruptures in cells harboring no mutation are less known. Here we show that repeated loss of nuclear envelope integrity in nuclei experiencing mechanical constraints promotes senescence in nontransformed cells, and induces an invasive phenotype including increased collagen degradation in human breast cancer cells, both in vitro and in a mouse xenograft model of breast cancer progression. We show that these phenotypic changes are due to the presence of chronic DNA damage and activation of the ATM kinase. In addition, we found that depletion of the cytoplasmic exonuclease TREX1 is sufficient to abolish the DNA damage in mechanically challenged nuclei and to suppress the phenotypes associated with the loss of nuclear envelope integrity. Our results also show that TREX1-dependent DNA damage induced by physical confinement of tumor cells inside the mammary duct drives the progression of in situ breast carcinoma to the invasive stage. We propose that DNA damage in mechanically challenged nuclei could affect the pathophysiology of crowded tissues by modulating proliferation and extracellular matrix degradation of normal and transformed cells. Main text:( 16,21-23 ), and corresponds to cells squeezing each other as they move. These deformed nuclei were also more often positive for γH2AX than less deformed ones, and in general displayed more intense γH2AX staining than nuclei in the bulk of the tumor ( Fig. 1A-C). This suggests that, at the in situ stage of human breast carcinoma, strands of motile cells at the periphery of the tumor exhibit pronounced nuclear deformation, associated with elevated DNA damage.We then investigated a xenograft mouse model based on the intraductal (intra-nipple) injection of transformed human breast MCF10DCIS.com (DCIS) cells, which are derived from the nontransformed MCF10A cell line, and that recapitulates the transition from in situ to invasive stages of breast cancer progression ( 17,24 ). Tumors at various stages were analyzed. In situ tumors (further divided in early and advanced stages based on the continuity of the myoepithelial layer) are characterized by a myoepithelial layer (stained with smooth muscle actin) surrounding human xenograft cells that were identified by the human-specific KU70 antibody. In order to verify whether human tumor cells similarly experience DNA damage in the tumor xenografts, tumor sections were stained with γH2AX ( Fig. 1E-F, Fig. S1A). Similar to human tumor specimens, xenograft tumors displayed numerous deformed nuclei, mostly found at the periphery of the tumor mass, close to the myoepithelium, especially in ducts inflated with large tumors (Fig. 1E-F), and enriched in regions showing micro-invasive foci. Strikingly, these regions were also enriched in γH2AX-positive nuclei (Fig. 1F). Since γH2AX is also known...

Nuclear Deformation Causes DNA Damage by Increasing Replication Stress

“…Consistent with this idea, p53 has been reported to mediate replication stress and restart DNA replication at stalled forks, thereby preventing replication fork collapse and associated DNA damage. 60 At the same time, differences in the susceptibility of cells to NE rupture-induced DNA damage could be due to differences in the expression of cytoplasmic nucleases 21,23 or other factors.…”

“…Taken together, these findings indicate that NE rupture-induced DNA damage occurs independent of cell cycle stage, consistent with it resulting from the influx of cytoplasmic nucleases, which would attack DNA irrespective of cell cycle stage ( Figure 6). [21][22][23] In contrast, deformation-induced DNA damage occurs primarily in the S/G2 phase of the cell cycle, suggesting that it is linked to DNA replication.…”

“…[9][10][11][12][13][14][15][16] Transient loss of NE integrity allows uncontrolled exchange between the nucleoplasm and cytoplasm; exposes the genomic DNA to cytoplasmic components, such as nucleases; and leads to DNA damage. [9][10][11]13,15,[17][18][19][20][21][22][23] Although cells rapidly repair their NE and continue to survive and migrate, 10,11,13 the acquired DNA damage can increase genomic instability in these cancer cells, 9,14,15,19,23,24 which could further enhance their metastatic potential and resistance to therapies.…”

“…Recent reports have implicated loss of DNA repair factors during NE rupture or local exclusion of repair factors due to nuclear deformation as possible mechanisms. [15][16][17][18][19] Exposure to cytoplasmic DNases, such as TREX1, following NE rupture [21][22][23] and mislocalization of organelles like mitochondria post-NE rupture 25 have also been suggested as cause of DNA damage. Contributing to the uncertainty about the molecular mechanism responsible for the confined migration-induced DNA damage is that previous studies have provided at times conflicting results on the association between DNA damage and NE rupture.…”

APOBEC3-dependent kataegis and TREX1-driven chromothripsis during telomere crisis

“…Data Fig. 1d ) 15 . Additionally, DNA bridges were not found to elicit significant cGAS/STING signaling in an analogous model of telomere crisis in cGAS-positive MCF10A cells 16 .…”