Intracellular amphiregulin is critical for heterochromatin maintenance and genomic stability in response to replication stress in BRCA2 ^mut/+ mammary epithelial cells

The BRCA2 tumor suppressor protein has roles in homologous recombination DNA repair and replication fork protection and carriers of mutation in the BRCA2 gene ( BRCA2 ^mut/+ ) are at high risk for breast cancers as a consequence of genomic instability. Mammary epithelial cells (MECs) are subjected to significant hormone-driven replication. BRCA2 ^mut/+ MECs must therefore be able to mitigate RS-induced chronic DNA damage that could result in cell death or senescence. In response to acute RS in hTERT-MECs, we have found that the EGFR ligand, amphiregulin (AREG) rapidly undergoes retrograde trafficking to the nucleus and nuclear membrane (NM) in association with lamin A resulting in a transient increase in H3K9me3 heterochromatin most markedly in BRCA2 ^mut/+ hTERT-MECs. RS promoted NM incorporation of prelamin A that was dependent on endogenous but not exogenous AREG. Prelamin A is reported to stabilize SUV39h1 and expression of a C-terminal truncated AREG that translocates to the NM increased SUV39h1, HP1α and H3K9me3. AREG knockdown reduced both HP1α and SUV39h1 accompanied by massive decompaction and reduction in global H3K9me3 heterochromatin. Loss of heterochromatin upon AREG depletion increased DNA damage, reduced replication fork speed and resulted in a significant increase in firing of new and multiple origins. Knockdown of endogenous AREG also impacted the NM resulting in activation of a type-1 interferon (IFN)-like response promoting senescence. Chronic RS also stimulated nuclear AREG along with FOXM1 and ESR1 mRNA and protein. Consistent with this, nuclear AREG, FOXM1 and ERα were detected in hyperplastic regions of human tissue. Overall, our work reveals a critical and unanticipated nuclear role for AREG in maintenance and transient induction of heterochromatin in response to RS preventing excessive DNA damage and senescence in BRCA2 ^mut/+ MECs. Since nuclear AREG also promotes ERα expression, this may link RS with aberrant expansion of estrogen receptor-expressing MECs found in BRCA2 ^mut/+ mammary tissue.