Published at Senescent Cell Research Group
ABSTRACT
Cellular senescence is a stress response program characterized by a robust cell cycle arrest and the induction of a proinflammatory senescence-associated secretory phenotype (SASP) that is triggered through an unknown mechanism. Here, we show that, during oncogene-induced senescence (OIS), the Toll-like receptor 2 (TLR2) and its partner TLR10 are key mediators of senescence in vitro and in murine models. TLR2 promotes cell cycle arrest by regulating the tumor suppressors p53-p21CIP1, p16INK4a, and p15INK4b and regulates the SASP through the induction of the acute-phase serum amyloids A1 and A2 (A-SAAs) that, in turn, function as the damage-associated molecular patterns (DAMPs) signaling through TLR2 in OIS. Last, we found evidence that the cGAS-STING cytosolic DNA sensing pathway primes TLR2 and A-SAAs expression in OIS. In summary, we report that innate immune sensing of senescence-associated DAMPs by TLR2 controls the SASP and reinforces the cell cycle arrest program in OIS.
INTRODUCTION
Cellular senescence is a cell cycle arrest program induced by various stresses that renders cells insensitive to mitogenic signals and impairs the proliferation and expansion of damaged cells (1). Activation of oncogenes such as RAS in somatic cells induces a senescence program termed oncogene-induced senescence (OIS) (2). OIS is a cell-intrinsic tumor suppressor mechanism that impairs tumor progression (2–7). Senescence is also characterized by the activation of the senescence-associated secretory phenotype (SASP). The SASP is a cocktail of proinflammatory cytokines, chemokines, growth factors, and matrix-remodeling proteins with diverse functions and roles (8–10). Through the SASP, senescent cells elicit multiple paracrine effects to promote the normal processes associated with inflammation, wound healing, tissue remodeling, and cell plasticity (11–14) or it can disrupt tissue homeostasis, promoting aging and other pathophysiological conditions such as fibrosis and cancer (12). The SASP reinforces the cell cycle arrest by activating p53 and the cell cycle inhibitors p21CIP1 and p15INK4b (8, 10, 15). The SASP also promotes paracrine senescence (15) and stimulates immune surveillance that results in clearance of the senescent cells (16–18), which collectively contribute to the tumor suppressor program elicited by OIS. Therefore, the immune system can regulate senescence and determine senescent cell fate (19–21).
The activation of the SASP is primarily regulated by the transcription factors nuclear factor κB (NF-κB) and CCAAT/enhancer-binding protein β (C/EBPβ) (8, 10, 22). SASP activation responds to a hierarchical model where interleukin-1α (IL-1α) and IL-1β are the apical cytokines signaling through IL-1 receptor (IL-1R), triggering a cascade of additional amplifying feedback loops by other cytokines and chemokines of the SASP. As a consequence, NF-κB and C/EBPβ activation are sustained, and the expression of the SASP is maintained. Interfering with these feedback loops, such as the one set up by IL-1, IL-6, or IL-8, collapses the network, impairing the induction of the SASP and the activation of the cell cycle arrest (8, 10, 15, 23).
We have previously shown that the inflammasome regulates the activation of the SASP (15). The inflammasome is a multiprotein platform that induces the proteolytic activity of the inflammatory cysteine-aspartic protease Caspase-1. It is a key regulatory component of the innate immune response and the first line of defense against pathogens and damage (24). Activation of the inflammasome leads to cleavage and activation of the proinflammatory cytokine IL-1β. Inflammasomes are controlled by a family of receptors called pattern recognition receptors (PRRs). PRRs are receptors of the innate immune system that are activated by interaction with pathogen-associated molecular patterns (PAMPs) or with damage-associated molecular patterns (DAMPs) that are generated endogenously in cells under certain conditions of stress and damage (25). There are three major PRR families: Toll-like receptors (TLRs), RIG-I–like receptors, and NOD-like receptors (25). Upon activation, PRRs induce distinct signal transduction pathways that activate an immune transcriptional program mostly regulated by NF-κB and interferon regulatory factors (IRFs).
Although the steady-state signaling of the SASP is relatively well understood, the molecular mechanism(s) that initiates the SASP in OIS and how the inflammasome is primed during OIS remain ill defined. Here, we describe the mechanism that underlies the priming of the inflammasome in OIS by PRRs, identifying the senescence-associated DAMP that initiates the SASP and reinforces OIS.
Fig. 1TLR2 expression is induced during OIS.
(A) Schematic showing IMR90 ER:RAS cells treated with 4OHT undergo OIS. IMR90 ER:STOP cells serve as a control and retain proliferative capacity with 4OHT. (B) Quantitative reverse transcription polymerase chain reaction (qRT-PCR) analysis of TLR family member expression in IMR90 ER:RAS and ER:STOP cells treated with 4OHT for 5 and 8 days. (C) Western blot of TLR2 expression in IMR90 ER:RAS and ER:STOP (ER:S) cells with up to 10 days of 4OHT treatment (top). The 5-bromo-2′-deoxyuridine (BrdU) incorporation in IMR90 ER:RAS cells treated with 4OHT for up to 8 days, as indicated. (D) RNA was extracted from snap-frozen liver samples from wild-type (WT) mice 6 days following hydrodynamic delivery of NrasG12V/D38A (n = 5) and NrasG12V (n = 4) transposons. tlr2 mRNA expression was measured using qRT-PCR. Scatter plots represents value per animal, with the horizontal line representing group means ± SEM. Statistical significance was calculated using Students two-tailed t test. **P < 0.01. (E) Immunohistochemical staining for Nras and Tlr2 in consecutive liver sections from corresponding mice in (D) showing that oncogenic NrasG12V expressing, but not NrasG12V/D38A expressing, hepatocytes express Tlr2. Scale bars, 50 μm.
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F U L L T E X T : ScienceAdvances
Edited by Engadin, 08 June 2019 - 01:16 AM.
