Titolo della tesi: Nuclear re-localization of Argonaute2 during doxorubicin induced senescence and its possible roles.
ABSTRACT
The Argounate family (Ago) is a group of proteins highly conserved in Eukaryotes that play a key role in small-RNA guided gene silencing processes. These proteins have been found to be involved in chromatin remodeling, constitutive and alternative splicing and in double stranded DNA damage repair. Recent evidence involves Argonaute2 (Ago2) in senescent programs, during Oncogene-Induced Senescence (OIS) and Cytokine-Induced Senescence (CIS), in human cells. The goal of our research was to look into the probable involvement of Ago2 during doxorubicin-induced senescence in MCF7 breast cancer cells. To begin, we determine the reliability of doxorubicin treated MCF7 cells as senescent cellular system. We discovered the re-localization of Ago2 in the nucleus of MCF7 following doxorubicin treatment. In order to better understand its molecular role into the nucleus, we performed immunoprecipitation of endogenous Ago2 from the nuclei of doxorubicin-treated MCF7 cells and untreated MCF7 cells, followed by Orbitrap LS-MS proteomic analysis. The mass spectrometry analysis reveals that nuclear Ago2 preferentially immunoprecipitates with heterogeneous nuclear ribonucleoproteins (hnRNPs), a class of RNA binding proteins involved in mRNA processing and maturation. Many studies have found Ago2 to be involved in alternative splicing, and splicing has recently been identified as a hallmark of senescence. Here, we identify the hnRNPM as a novel Ago2 interactor in nuclei of senescent cells, and we can speculate that Ago2 is involved in alternative splicing during senescence. Furthermore, we found that both partial and complete Ago2 deficiency impairs cellular senescence to some extent. These findings, on the one hand, confirm Ago2's role in senescence and, on the other hand, raise questions about the possible redundancy role of other Ago proteins.