Nuclear Translocation of Argonaute 2 in Cytokine-Induced Senescence

Abstract

Background/Aims: Cellular senescence, or permanent growth arrest, is known as an effective tumor suppressor mechanism that can be induced by different stressors, such as oncogenes, chemotherapeutics or cytokine cocktails. Previous studies demonstrated that the growth-repressing state of oncogene-induced senescent cells depends on argonaute protein 2 (Ago2)-mediated transcriptional gene silencing and Ago2/Rb corepression of E2F-dependent cell cycle genes. Cytokine-induced senescence (CIS) likewise depends on activation of the p16^Ink4a/Rb pathway, and consecutive inactivation of the E2F family of transcription factors. In the present study, we therefore analyzed the role of Ago2 in CIS. Methods: Human cancer cell lines were treated with interferon-gamma (IFN-γ) and tumor necrosis factor (TNF) to induce senescence. Senescence was determined by growth assays and measurement of senescence-associated β-galactosidase (SA-β-gal) activity, Ago2 translocation by Ago2/ Ki67 immunofluorescence staining and western blot analysis, and gene transcription by quantitative polymerase chain reaction (qPCR). Results: IFN-γ and TNF permanently stopped cell proliferation and time-dependently increased SA-β-gal activity. After 24 – 48 h of cytokine treatment, Ago2 translocated from the cytoplasm into the nucleus of Ki67-negative cells, an effect which was shown to be reversible. Importantly, the proinflammatory cytokine cocktail suppressed Ago2-regulated cell cycle control genes, and siRNA-mediated depletion of Ago2 interfered with cytokine-induced growth inhibition. Conclusion: IFN-γ and TNF induce a stable cell cycle arrest of cancer cells that is accompanied by a fast nuclear Ago2 translocation and repression of Ago2-regulated cell cycle control genes. As Ago2 downregulation impairs cytokine-induced growth regulation, Ago2 may contribute to tissue homeostasis in human cancers.

A nuclear function for an oncogenic microRNA as a modulator of snRNA and splicing

“…5 a) and [ 12 , 56 – 58 ], and second, the co-IP of the spliceosomal SART3 and PRPF8 complexes with Ago2, the core endonucleolytic components of RISC that efficiently pools-down the entire miR-10b population from glioma cells. Our data showing the presence of nuclear Ago2 and its interactions with splicing factors is in agreement with prior reports [ 59 – 61 ]. These data support the scenario of aberrant nuclear miRNA maturation in cancer cells and links nuclear miRNA with the regulation of alternative splicing.…”

FAM172A controls the nuclear import and alternative splicing function of AGO2

“…Interestingly, our data suggesting a regulatory role for CK2 in FAM172A-mediated AGO2 nuclear import (Fig.4) provide a potential mechanistic link to explain why and how different cellular stresses can increase AGO2 translocation toward the nucleus (Castanotto et al, 2018, Rentschler et al, 2018). Indeed, CK2 activity is known to be enhanced by multiple cellular stressors (Ampofo, Sokolowsky et al, 2013, Sayed, Kim et al, 2000, Watabe & Nakaki, 2011).…”

“…The relative subcellular distribution of AGO2 is also known to vary as a function of cell types and culture conditions, being notably generally increased in the nucleus of primary cell cultures as opposed to most immortalized cell lines (Sarshad et al, 2018; Sharma et al, 2016). Other conditions that increase AGO2 nuclear localization include cellular stress (Castanotto, Zhang et al, 2018, Rentschler, Chen et al, 2018), senescence (Benhamed et al, 2012; Rentschler et al, 2018) and differentiation (Sarshad et al, 2018). What is less clear is how AGO2 shuttles between both compartments.…”

Cytokine-Induced Senescence in the Tumor Microenvironment and Its Effects on Anti-Tumor Immune Responses

“…In addition to the functional evidence for immune-mediated senescence induction that was derived from complex in vivo studies or patients, other analyses focused on the mechanistic details in cell culture-based works. Different reports-including our own workcould show that the direct application of cytokines has a similar senescence-inducing effect compared with the action of certain immune cells that release these factors [25,107,111]. Although most reports focus on the effects mediated by IFN-γ and TNF, either alone or in combination, other cytokines have also been associated with the induction of senescence.…”