Chanhee Kang

The DNA damage response induces inflammation and senescence by inhibiting autophagy of GATA4

Transcriptional control of cell senescence Senescent cells that have stopped proliferating secrete molecules that influence the cells around them. Prevention of this senescence-activated secretory phenotype seems to slow organismal aging. Kang et al. explored the regulatory process behind cell senescence and found that DNA damage led to stabilization of the transcription factor GATA4 (see the Perspective by Cassidy and Narita). Increased activity of GATA4 in senescent cells stimulated genes encoding secreted factors. GATA4 also accumulates in the brains of aging mice or humans. Science, this issue 10.1126/science.aaa5612; see also p. 1448 The transcription factor GATA4 promotes cell senescence. [Also see Perspective by Cassidy and Narita] INTRODUCTION Cellular senescence is a program of arrested proliferation and altered gene expression triggered by many stresses. Although it is a potent tumor-suppressive mechanism, senescence has been implicated in several pathological processes including aging, age-associated diseases, and (counterintuitively) tumorigenesis. One potential mechanism through which senescent cells exert such pleiotropic effects is the secretion of proinflammatory cytokines, chemokines, growth factors, and proteases, termed the senescence-associated secretory phenotype (SASP), which affects senescent cells and their microenvironment. The mechanism by which the SASP is initiated and maintained is not well characterized beyond the classical regulators of inflammation, including the transcription factors NF-κB and C/EBPβ. RATIONALE In senescence growth arrest, two core senescence-regulating pathways, p53 and p16INK4a/Rb, play a critical role. By contrast, the SASP does not depend on either p53 or p16INK4a, which suggests the existence of an independent senescence regulatory network that controls the SASP. Having observed high levels of induction of microRNA miR-146a during induced senescence in human fibroblasts, we developed a green fluorescent protein–tagged senescence reporter based on a miR-146a promoter fragment. This reporter responded to senescence-inducing stimuli, including replicative exhaustion, DNA damage, and oncogenic RAS activation—all of which activate the SASP. This system allowed us to identify additional regulators of senescence and the SASP. RESULTS Through miR-146a promoter analysis, we mapped the critical region for senescence-induced activity and identified the transcriptional regulator responsible for this regulation, GATA4, previously known as a regulator of embryonic development. Ectopic expression of GATA4 induced senescence, whereas disruption of GATA4 suppressed it, thus establishing GATA4 as a senescence regulator. GATA4 protein abundance, but not mRNA, increased during sene1scence, primarily as a result of increased protein stability. Under normal conditions, GATA4 binds the p62 autophagy adaptor and is degraded by selective autophagy. Upon senescence induction, however, this selective autophagy was suppressed through decreased interaction between GATA4 and p62, thereby stabilizing GATA4. GATA4 in turn induced TRAF3IP2 (tumor necrosis factor receptor–associated factor interacting protein 2) and IL1A (interleukin 1A), which activate NF-κB to initiate and maintain the SASP, thus facilitating senescence. GATA4 pathway activation depends on the key DNA damage response (DDR) kinases ATM (ataxia telangiectasia mutated) and ATR (ataxia telangiectasia and Rad3–related), as does senescence-associated activation of p53 and p16INK4a. However, the GATA4 pathway is independent of p53 and p16INK4a. Finally, GATA4 protein accumulated in multiple tissues in mice treated with senescence-inducing stimuli and during normal mouse and human aging, including many cell types in the brain; these findings raise the possibility that the GATA4 pathway drives age-dependent inflammation. CONCLUSION Our results indicate that GATA4 connects autophagy and the DDR to senescence and inflammation through TRAF3IP2 and IL1A activation of NF-κB. These findings establish GATA4 as a key switch activated by the DDR to regulate senescence, independently of p53 and p16INK4a. Our in vivo data indicate a potential role of GATA4 during aging and its associated inflammation. Because accumulation of senescent cells is thought to promote aging and aging-associated diseases through the resulting inflammatory response, inhibiting the GATA4 pathway may provide an avenue for therapeutic intervention. GATA4 functions as a key switch in the senescence regulatory network to activate the SASP. The nonsenescent state is maintained by inhibitory barriers that prevent cell cycle arrest and inflammation. Upon senescence-inducing signals, ATM and ATR relieve inhibition of the p53 and p16INK4a pathways to induce growth arrest and also block p62-dependent autophagic degradation of GATA4, resulting in NF-κB activation and SASP induction. Cellular senescence is a terminal stress-activated program controlled by the p53 and p16INK4a tumor suppressor proteins. A striking feature of senescence is the senescence-associated secretory phenotype (SASP), a pro-inflammatory response linked to tumor promotion and aging. We have identified the transcription factor GATA4 as a senescence and SASP regulator. GATA4 is stabilized in cells undergoing senescence and is required for the SASP. Normally, GATA4 is degraded by p62-mediated selective autophagy, but this regulation is suppressed during senescence, thereby stabilizing GATA4. GATA4 in turn activates the transcription factor NF-κB to initiate the SASP and facilitate senescence. GATA4 activation depends on the DNA damage response regulators ATM and ATR, but not on p53 or p16INK4a. GATA4 accumulates in multiple tissues, including the aging brain, and could contribute to aging and its associated inflammation.

To be or not to be, the level of autophagy is the question: dual roles of autophagy in the survival response to starvation.

Autophagy is an evolutionally conserved lysosomal pathway used to degrade and turn over long-lived proteins and cytoplasmic organelles. Since autophagy was discovered, it has been thought to act as a pro-survival response to several stresses, especially starvation, at the cell and organism levels by providing recycled metabolic substrates to maintain energy homeostasis. However, several recent studies suggest that autophagy also plays a pro-death role through an autophagic cell death pathway mostly at the cellular level. The mechanism by which autophagy could perform these seemingly opposite roles as a pro-survival and a pro-death mechanism remained elusive until recently. Using C. elegans as a model system, we found that physiological levels of autophagy promote optimal survival of C. elegans during starvation, but either insufficient or excessive levels of autophagy render C. elegans starvation-hypersensitive. Furthermore, we found that muscarinic acetylcholine receptor signaling is important in modulating the level of autophagy during starvation, perhaps through DAP kinase and RGS-2. Our recent study provides in vivo evidence that levels of autophagy are critical in deciding its promotion of either survival or death: Physiological levels of autophagy are pro-survival, whereas insufficient or excessive levels of autophagy are pro-death. Addendum to: Kang C, You YJ, Avery L. Dual roles of autophagy in the survival of Caenorhabditis elegans during starvation. Genes Dev 2007; 21:2161-71.

Systemic regulation of starvation response in Caenorhabditis elegans

When the supply of environmental nutrients is limited, multicellular animals can make both physiological and behavioral changes so as to cope with nutrient starvation. Although physiological and behavioral effects of starvation are well known, the mechanisms by which animals sense starvation systemically remain elusive. Furthermore, what constituent of food is sensed and how it modulates starvation response is still poorly understood. In this study, we use a starvation-hypersensitive mutant to identify molecules and mechanisms that modulate starvation signaling. We found that specific amino acids could suppress the starvation-induced death of gpb-2 mutants, and that MGL-1 and MGL-2, Caenorhabditis elegans homologs of metabotropic glutamate receptors, were involved. MGL-1 and MGL-2 acted in AIY and AIB neurons, respectively. Treatment with leucine suppressed starvation-induced stress resistance and life span extension in wild-type worms, and mutation of mgl-1 and mgl-2 abolished these effects of leucine. Taken together, our results suggest that metabotropic glutamate receptor homologs in AIY and AIB neuron may modulate a systemic starvation response, and that C. elegans senses specific amino acids as an anti-hunger signal.

Death‐associated protein kinase (DAPK) and signal transduction: fine‐tuning of autophagy in Caenorhabditis elegans homeostasis

Autophagy is an evolutionarily conserved lysosomal pathway used to degrade and recycle long‐lived proteins and cytoplasmic organelles. This homeostatic ability makes autophagy an important pro‐survival mechanism in response to several stresses, such as nutrient starvation, hypoxia, damaged mitochondria, protein aggregation and pathogens. However, several recent studies have highlighted that autophagy also acts as a pro‐death mechanism. What on the surface seem like conflicting roles of autophagy may be explained by the fact that the decision between pro‐survival and pro‐death is determined by the level of activation. A better understanding of autophagy signaling pathways will be helpful to elucidate how the level of autophagy is precisely regulated under different conditions and eventually how the final outcome is decided. In this review, we briefly discuss the pro‐survival and pro‐death roles of autophagy, and then discuss the mechanism by which autophagy is regulated, mainly focusing on death‐associated protein kinase in the nematode Caenorhabditis elegans.

How autophagy both activates and inhibits cellular senescence

ABSTRACT Autophagy and cellular senescence are stress responses essential for homeostasis. While recent studies indicate a genetic relationship between autophagy and senescence, whether autophagy acts positively or negatively on senescence is still subject to debate. Although autophagy was originally recognized as a nonspecific lysosomal degradation pathway (general autophagy), increasing evidence supports a selective form of autophagy that mediates the degradation of specific targets (selective autophagy). Our recent study revealed distinctive roles of selective autophagy and general autophagy in the regulation of senescence, at least in part resolving apparently contradictory reports regarding the relationship between these 2 important homeostatic stress responses.

A gain-of-function senescence bypass screen identifies the homeobox transcription factor DLX2 as a regulator of ATM–p53 signaling

Senescence stimuli activate multiple tumor suppressor pathways to initiate cycle arrest and a differentiation program characteristic of senescent cells. We performed a two-stage, gain-of-function screen to select for the genes whose enhanced expression can bypass replicative senescence. We uncovered multiple genes known to be involved in p53 and Rb regulation and ATM regulation, two components of the CST (CTC1-STN1-TEN1) complex involved in preventing telomere erosion, and genes such as REST and FOXO4 that have been implicated in aging. Among the new genes now implicated in senescence, we identified DLX2, a homeobox transcription factor that has been shown to be required for tumor growth and metastasis and is associated with poor cancer prognosis. Growth analysis showed that DLX2 expression led to increased cellular replicative life span. Our data suggest that DLX2 expression reduces the protein components of the TTI1/TTI2/TEL2 complex, a key complex required for the proper folding and stabilization of ATM and other members of the PIKK (phosphatidylinositol 3-kinase-related kinase) family kinase, leading to reduced ATM-p53 signaling and senescence bypass. We also found that the overexpression of DLX2 exhibited a mutually exclusive relationship with p53 alterations in cancer patients. Our functional screen identified novel players that may promote tumorigenesis by regulating the ATM-p53 pathway and senescence.

Systemic regulation of autophagy in Caenorhabditis elegans.

When no supply of environmental nutrients is available, cells induce autophagy, thereby generating a source of emergency metabolic substrates and energy to maintain the basal cellular activity needed for survival. This autophagy response to starvation has been well characterized in various multicellular organisms, including worms, flies, and mice. Although prosurvival effects of autophagy in response to starvation are well known in animals, the mechanisms by which animals regulate and coordinate autophagy systemically remain elusive. Using C. elegans as a model system, we found that specific amino acids could regulate starvation-induced autophagy, and that MGL-1 and MGL-2, Caenorhabditis elegans homologs of metabotropic glutamate receptors, were involved. MGL-1 and MGL-2 specifically acted in AIY and AIB neurons, respectively, to modulate the autophagy response in other tissues such as pharyngeal muscle. Our recent study suggests that the autophagy response to starvation, previously thought to be cell-autonomous, can be systemically regulated, and that there is a specific sensor for monitoring systemic amino acids levels in Caenorhabditis elegans.

Genetic interrogation of replicative senescence uncovers a dual role for USP28 in coordinating the p53 and GATA4 branches of the senescence program

Senescence is a terminal differentiation program that halts the growth of damaged cells and must be circumvented for cancer to arise. Here we describe a panel of genetic screens to identify genes required for replicative senescence. We uncover a role in senescence for the potent tumor suppressor and ATM substrate USP28. USP28 controls activation of both the TP53 branch and the GATA4/NFkB branch that controls the senescence-associated secretory phenotype (SASP). These results suggest a role for ubiquitination in senescence and imply a common node downstream from ATM that links the TP53 and GATA4 branches of the senescence response.