RESUMO
Several members of the Poly(ADP-ribose) polymerase (PARP) family are essential regulators of genome integrity, actively prospected as drug targets for cancer therapy. Among them, PARP3 is well characterized for its functions in double-strand break repair and mitotis. Here we report that PARP3 also plays an integral role in TGFß and reactive oxygen species (ROS) dependent epithelial-to-mesenchymal transition (EMT) and stem-like cell properties in human mammary epithelial and breast cancer cells. PARP3 expression is higher in breast cancer cells of the mesenchymal phenotype and correlates with the expression of the mesenchymal marker Vimentin while being in inverse correlation with the epithelial marker E-cadherin. Furthermore, PARP3 expression is significantly upregulated during TGFß-induced EMT in various human epithelial cells. In line with this observation, PARP3 depletion alters TGFß-dependent EMT of mammary epithelial cells by preventing the induction of the Snail-E-cadherin axis, the dissolution of cell junctions, the acquisition of cell motility and chemoresistance. PARP3 responds to TGFß-induced ROS to promote a TG2-Snail-E-cadherin axis during EMT. Considering the link between EMT and cancer stem cells, we show that PARP3 promotes stem-like cell properties in mammary epithelial and breast cancer cells by inducing the expression of the stem cell markers SOX2 and OCT4, by increasing the proportion of tumor initiating CD44high/CD24low population and the formation of tumor spheroid bodies, and by promoting stem cell self-renewal. These findings point to a novel role of PARP3 in the control of TGFß-induced EMT and acquisition of stem-like cell features and further motivate efforts to identify PARP3 specific inhibitors.
Assuntos
Neoplasias da Mama/enzimologia , Caderinas/metabolismo , Proteínas de Ciclo Celular/metabolismo , Transição Epitelial-Mesenquimal , Proteínas de Ligação ao GTP/metabolismo , Glândulas Mamárias Humanas/enzimologia , Células-Tronco Neoplásicas/enzimologia , Poli(ADP-Ribose) Polimerases/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Fatores de Transcrição da Família Snail/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Transglutaminases/metabolismo , Células A549 , Antígenos CD , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Antígeno CD24/metabolismo , Caderinas/genética , Proteínas de Ciclo Celular/genética , Movimento Celular , Autorrenovação Celular , Resistencia a Medicamentos Antineoplásicos , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Etoposídeo/farmacologia , Feminino , Proteínas de Ligação ao GTP/genética , Regulação Neoplásica da Expressão Gênica , Células Hep G2 , Humanos , Receptores de Hialuronatos/metabolismo , Glândulas Mamárias Humanas/patologia , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Fator 3 de Transcrição de Octâmero/metabolismo , Fenótipo , Poli(ADP-Ribose) Polimerases/genética , Proteína 2 Glutamina gama-Glutamiltransferase , Interferência de RNA , Fatores de Transcrição SOXB1/metabolismo , Transdução de Sinais , Fatores de Transcrição da Família Snail/genética , Esferoides Celulares , Fatores de Tempo , Inibidores da Topoisomerase II/farmacologia , Transfecção , Transglutaminases/genéticaRESUMO
The repair of toxic double-strand breaks (DSB) is critical for the maintenance of genome integrity. The major mechanisms that cope with DSB are: homologous recombination (HR) and classical or alternative nonhomologous end joining (C-NHEJ versus A-EJ). Because these pathways compete for the repair of DSB, the choice of the appropriate repair pathway is pivotal. Among the mechanisms that influence this choice, deoxyribonucleic acid (DNA) end resection plays a critical role by driving cells to HR, while accurate C-NHEJ is suppressed. Furthermore, end resection promotes error-prone A-EJ. Increasing evidence define Poly(ADP-ribose) polymerase 3 (PARP3, also known as ARTD3) as an important player in cellular response to DSB. In this work, we reveal a specific feature of PARP3 that together with Ku80 limits DNA end resection and thereby helps in making the choice between HR and NHEJ pathways. PARP3 interacts with and PARylates Ku70/Ku80. The depletion of PARP3 impairs the recruitment of YFP-Ku80 to laser-induced DNA damage sites and induces an imbalance between BRCA1 and 53BP1. Both events result in compromised accurate C-NHEJ and a concomitant increase in DNA end resection. Nevertheless, HR is significantly reduced upon PARP3 silencing while the enhanced end resection causes mutagenic deletions during A-EJ. As a result, the absence of PARP3 confers hypersensitivity to anti-tumoral drugs generating DSB.