KLHL20 links the ubiquitin-proteasome system to autophagy termination

Autophagy is a dynamic and self-limiting process. The amplitude and duration of this process need to be properly controlled to maintain cell homeostasis, and excessive or insufficient autophagy activity could each lead to disease states. Compared to our understanding of the molecular mechanisms of autophagy induction, little is known about how the autophagy process is turned off after its activation. We recently identified KLHL20 as a key regulator of autophagy termination. By functioning as a substrate-binding subunit of CUL3 ubiquitin ligase, KLHL20 targets the activated ULK1 and phagophore-residing PIK3C3/VPS34 and BECN1 for ubiquitination and proteasomal degradation, which in turn triggers a destabilization of their complex components ATG13 and ATG14. These hierarchical degradation events cause the exhaustion of the autophagic pool of ULK1 and PIK3C3/VPS34 complexes, thereby preventing persistent and excessive autophagy activity. Impairment of KLHL20-dependent feedback regulation of autophagy enhances cell death under prolonged starvation and aggravates muscle atrophy in diabetic mice, which highlights the pathophysiological significance of this autophagy termination mechanism in cell survival and tissue homeostasis. Modulation of this autophagy termination pathway may be effective for treating diseases associated with deregulation of autophagy activity.     

Role of the proteolytic hierarchy between cathepsin L, cathepsin D and caspase-3 in regulation of cellular susceptibility to apoptosis and autophagy

The present investigation was undertaken to measure the relative abilities of pro-death versus pro-survival proteases in degrading each other and to determine how this might influence cellular susceptibility to death. For this, we first carried out in vitro experiments in which recombinant pro-death proteases (caspase-3 or cathepsin D) were incubated with the pro-survival protease (cathepsin L) in their respective optimal conditions and determined the effects of these reactions on enzyme integrity and activity. The results indicated that cathepsin L was able to degrade cathepsin D, which in turn cleaves caspase-3, however the later enzyme was unable to degrade any of the cathepsins. The consequences of this proteolytic sequence on cellular ability to undergo apoptosis or other types of cell death were studied in cells subjected to treatment with a specific inhibitor of cathepsin L or the corresponding siRNA. Both treatments resulted in suppression of cellular proliferation and the induction of a cell death with no detectable caspase-3 activation or DNA fragmentation, however, it was associated with increased accumulation of cathepsin D, cellular vaculolization, expression of the mannose-6-phosphate receptor, and the autophagy marker LC3-II, all of which are believed to be associated with autophagy. Genetic manipulations leading either to the gain or loss of cathepsin D expression implicated this enzyme as a key player in the switch from apoptosis to autophagy. Overall, these findings suggest that a hierarchy between pro-survival and pro-death proteases may have important consequences on cell fate.     

Crosstalk between apoptosis,necrosis and autophagy

Apoptosis and necrosis are the two major modes of cell death, the molecular mechanisms of which have been extensively studied. Although initially thought to constitute mutually exclusive cellular states, recent findings reveal cellular contexts that require a balanced interplay between these two modes of cellular demise. Several death initiator and effector molecules, signaling pathways and subcellular sites have been identified as key mediators in both processes, either by constituting common modules or alternatively by functioning as a switch allowing cells to decide which route to take, depending on the specific situation. Importantly, autophagy, which is a predominantly cytoprotective process, has been linked to both types of cell death, serving either a pro-survival or pro-death function. Here we review the recent literature that highlights the intricate interplay between apoptosis, necrosis and autophagy, focusing on the relevance and impact of this crosstalk in normal development and in pathology. This article is part of a Special Section entitled: Cell Death Pathways. Guest Editors: Frank Madeo and Slaven Stekovic.     

Regulation of cell growth by autophagy

Cell growth-the primary determinant of cell size-has an intimate relationship with proliferation; cells divide only after they reach a critical size. Despite its developmental and medical significance, little is known about cellular pathways that mediate the growth of cells. Accumulating evidence demonstrates a role for autophagy-a mechanism of eukaryotic cells to digest their own constituents during development or starvation-in cell size control. Increasing autophagic activity by prolonged starvation, rapamycin treatment inhibiting TOR (target of rapamycin) signaling, or genetic intervention, causes cellular atrophy in worms, flies and mammalian cell cultures. In contrast, we have shown that in the nematode Caenorhabditis elegans mutational inactivation of two autophagy genes, unc-51/Atg1 and bec-1/Atg6, confers reduced cell size. We argue that physiological levels of autophagy are required for normal cell size, whereas both insufficient and excessive levels of autophagy lead to retarded cell growth. Furthermore, we discuss data suggesting that the insulin/IGF-1 (insulin-like growth factor receptor-1) and TGF-beta (transforming growth factor-beta) signaling systems acting as major growth regulatory pathways converge on autophagy genes to control cell size. Thus, autophagy may act as a central regulatory mechanism of cell growth.     

Differential Dependence on Beclin 1 for the Regulation of Pro-Survival Autophagy by Bcl-2 and Bcl-xL in HCT116 Colorectal Cancer Cells

Autophagy is described to be involved in homeostasis, development and disease, both as a survival and a death process. Its involvement in cell death proceeds from interrelationships with the apoptotic pathway. We focused on survival autophagy and investigated its interplays with the apoptotic machinery. We found that while Mcl-1 remained ineffective, Bcl-2 and Bcl-xL were required for starved cells to display a fully functional autophagic pathway as shown by proteolysis activity and detection of autophagic vesicles. Such pro-autophagic functions of Bcl-2 and Bcl-xL were independent of Bax. However they appeared to operate through non redundant mechanisms as Bcl-xL wielded a tighter control than Bcl-2 over the regulation of autophagy: unlike Bcl-2, Bcl-xL and Atg7 manipulation yielded identical phenotypes suggesting they could be components of the same signalling pathway; Bcl-xL subcellular localisation was modified upon starvation, and importantly Bcl-xL acted independently of Beclin 1. Still an intact BH3-binding site was required for Bcl-xL to stimulate a fully functional autophagic pathway. This study highlights that, in addition to their well-established anti-death function during apoptosis, Bcl-2 and Bcl-xL have a broader role in cell survival. Should Bcl-2 and Bcl-xL stand at the cross-roads between pro-survival and pro-death autophagy, this study introduces the new concept that the regulation of autophagy by Bcl-2 and Bcl-xL is adjusted according to its survival or death outcome.     

Autophagy promotes survival of retinal ganglion cells after optic nerve axotomy in mice

Autophagy is an essential recycling pathway implicated in neurodegeneration either as a pro-survival or a pro-death mechanism. Its role after axonal injury is still uncertain. Axotomy of the optic nerve is a classical model of neurodegeneration. It induces retinal ganglion cell death, a process also occurring in glaucoma and other optic neuropathies. We analyzed autophagy induction and cell survival following optic nerve transection (ONT) in mice. Our results demonstrate activation of autophagy shortly after axotomy with autophagosome formation, upregulation of the autophagy regulator Atg5 and apoptotic death of 50% of the retinal ganglion cells (RGCs) after 5 days. Genetic downregulation of autophagy using knockout mice for Atg4B (another regulator of autophagy) or with specific deletion of Atg5 in retinal ganglion cells, using the Atg5(flox/flox) mice reduces cell survival after ONT, whereas pharmacological induction of autophagy in vivo increases the number of surviving cells. In conclusion, our data support that autophagy has a cytoprotective role in RGCs after traumatic injury and may provide a new therapeutic strategy to ameliorate retinal diseases.     

Endoplasmic reticulum-targeting photosensitizer Hypericin confers chemo-sensitization towards oxaliplatin through inducing pro-death autophagy

Hypericin is an endoplasmic reticulum (ER)-located photosensitizer, which causes oxidative damage to ER during photodynamic therapy (PDT). Hypericin-mediated PDT (HY-PDT) has been confirmed to enhance chemo-sensitivity of oxaliplatin (L-OHP) in colon cancer cells. The present study reveals that autophagy plays a key role in chemosensitization during HY-PDT. We proved pro-death autophagy was required for sensitization and HY-PDT/L-OHP antitumor synergism. High dosage of HY-PDT induced autophagic cell death; while low dose of HY-PDT predominantly triggered protective autophagy and promoted cell proliferation. Low dose of HY-PDT reduced the cytotoxicity of L-OHP in oxaliplatin-resistant colon cancer cells. Different level of autophagy therefore contributed to the opposite effect of HY-PDT on cell fate and chemo-sensitivity. Furthermore, we revealed the role of CHOP as a regulator connecting pro-survival and pro-death autophagy under ER damage. High dose of HY-PDT induced massive ROS generation and severe ER stress, which then led to induction of CHOP. CHOP thereby activated CHOP/TRIB3/Akt/mTOR cascade and triggered autophagic cell death. Additionally, when apoptotic pathway was blocked, cells treated with high dose of HY-PDT preferentially underwent death through autophagic pathway. On the other hand, suppression of autophagy made cells more vulnerable to apoptosis under low dose of HY-PDT. These results provided new evidences for the clinical application of ER-targeting PDT in modifying chemosensitivity of colorectal cancer therapy.     

Survival or death: disequilibrating the oncogenic and tumor suppressive autophagy in cancer

Autophagy (macroautophagy) is an evolutionarily conserved lysosomal degradation process, in which a cell degrades long-lived proteins and damaged organelles. Recently, accumulating evidence has revealed the core molecular machinery of autophagy in carcinogenesis; however, the intricate relationship between autophagy and cancer continue to remain an enigma. Why does autophagy have either pro-survival (oncogenic) or pro-death (tumor suppressive) role at different cancer stages, including cancer stem cell, initiation and progression, invasion and metastasis, as well as dormancy? How does autophagy modulate a series of oncogenic and/or tumor suppressive pathways, implicated in microRNA (miRNA) involvement? Whether would targeting the oncogenic and tumor suppressive autophagic network be a novel strategy for drug discovery? To address these problems, we focus on summarizing the dynamic oncogenic and tumor suppressive roles of autophagy and their relevant small-molecule drugs, which would provide a new clue to elucidate the oncosuppressive (survival or death) autophagic network as a potential therapeutic target.     

Signaling pathway of autophagy associated with innate immunity

Autophagy has recently been shown to be an important component of the innate immune response. The signaling pathways leading to activation of autophagy in innate immunity are not well studied. Our recent study shows that Toll-like receptor 4 (TLR 4) serves as an environmental sensor for autophagy. We define a new molecular pathway in which lipopolysaccharide (LPS) induces autophagy in human and murine macrophages by a pathway regulated through Toll-interleukin 1 receptor domain-containing adaptor-inducing interferon-beta (TRIF)-dependent, myeloid differentiation factor 88 (MyD88)-independent TLR4 signaling. Receptor-interacting protein (RIP1) and p38 mitogen-activated protein-kinase (MAPK) are downstream components of this pathway. This signaling pathway does not affect cell viability, indicating that it is distinct from an autophagic death signaling pathway. We further show that LPS-induced autophagy can enhance mycobacterial co-localization with the autophagosomes. The above study raises important questions. (1) What is the complete signaling pathway for LPS-induced autophagy? (2) Does TLR3 mediate autophagy? (3) What are the mechanisms that determine whether autophagy acts as a pro-death or pro-survival pathway? (4) What are the physiological functions of LPS-induced autophagosomes? Future studies examining the above questions should provide us with important clues as to how autophagy is regulated in innate immunity, and how autophagy can be utilized in pathogen clearance.     

昆虫细胞自噬的生物学意义和自噬体膜的来源

细胞自噬(autophagy)是生物体广泛存在的细胞内自主降解过程。该过程通过自我吞噬细胞质成分和细胞器形成具有双层膜结构的自噬体, 与溶酶体融合实现细胞内物质的循环利用。细胞自噬在饥饿、 缺氧、 内质网胁迫、 病原入侵、 蛋白聚集等不良环境条件下实现自我挽救, 而细胞自噬的大量发生也是程序性细胞死亡(PCD)的启动和执行者之一。目前人们对自噬体分子组装和自噬发生的分子通路已有较深入的了解, 但仍然在很多重要问题上难以达成共识。本文结合我们的研究进展, 对昆虫细胞自噬的生物学意义和自噬体膜的来源问题进行综述和探讨。昆虫在营养相对匮乏的情况下发生低水平自噬(常态自噬), 用于维持细胞内的新陈代谢和继续生存的需要。昆虫在摄食阶段受到过度饥饿的刺激, 在变态发育时期受到蜕皮激素(20E)的诱导, 幼虫组织细胞发生高水平自噬和凋亡(apoptosis), 细胞表现为不可逆死亡, 过度饥饿导致幼虫发育迟缓或者死亡, 而20E导致幼虫蜕皮和幼虫组织退化或消亡。不同于酵母和高等动物细胞中的深入研究, 病原入侵是否和如何诱导昆虫细胞发生自噬, 目前尚缺乏足够的文献依据, 值得深入探讨。几乎所有的细胞器(内质网、 高尔基体、 线粒体)膜都可能是自噬体膜的来源, 这一问题在昆虫中也有待进一步诠释。     

MAPK/JNK signalling: a potential autophagy regulation pathway

Autophagy refers to a lysosomal degradative pathway or a process of self-cannibalization. This pathway maintains nutrients levels for vital cellular functions during periods of starvation and it provides cells with survival advantages under various stress situations. However, the mechanisms responsible for the induction and regulation of autophagy are poorly understood. The c-Jun NH2-terminal kinase (JNK) signal transduction pathway functions to induce defence mechanisms that protect organisms against acute oxidative and xenobiotic insults. This pathway has also been repeatedly linked to the molecular events involved in autophagy regulation. The present review will focus on recent advances in understanding of the relationship between mitogen-activated protein kinase (MAPK)/JNK signalling and autophagic cell death.     

Guttiferone K induces autophagy and sensitizes cancer cells to nutrient stress-induced cell death

BackgroundMedicinal plants have long been an excellent source of pharmaceutical agents. Autophagy, a catabolic degradation process through lysosomes, plays an important role in tumorigenesis and cancer therapy.PurposeThrough a screen designed to identify autophagic regulators from a library of natural compounds, we found that Guttiferone K (GUTK) can activate autophagy in several cancer cell lines. The objective of this study is to investigate the mechanism by which GUTK sensitizes cancer cells to cell death in nutrient starvation condition.MethodsCell death analysis was performed by propidium iodide staining with flow cytometry or Annexin V-FITC/PI staining assay. DCFH-DA staining was used for intracellular ROS measurement. Protein levels were analyzed by western blot analysis. Cell viability was measured by MTT assay.ResultsExposure to GUTK was observed to markedly induce GFP-LC3 puncta formation and activate the accumulation of LC3-II and the degradation of p62 in HeLa cells, suggesting that GUTK is an autophagy inducer. Importantly, hydroxychloroquine, an autophagy inhibitor, was found to significantly prevent GUTK-induced cell death in nutrient starvation conditions, suggesting that the cell death observed is largely dependent on autophagy. We further provide evidence that GUTK inhibits Akt phosphorylation, thereby inhibiting the mTOR pathway in cancer cells during nutrient starvation. In addition, GUTK causes the accumulation of reactive oxygen species (ROS) and the phosphorylation of JNK in EBSS, which may mediate both autophagy and apoptosis.ConclusionThese data indicate that GUTK sensitizes cancer cells to nutrient stress-induced cell death though Akt/mTOR dependent autophagy pathway.     

Autophagic and apoptotic response to stress signals in mammalian cells

Autophagy is a highly conserved catabolic programme for degrading proteins and organelles. This process has been shown to act as a pro-survival or pro-death mechanism in different physiological and pathological conditions. Several stress stimuli can induce autophagy, such as nutrient deprivation or critical steps in development of lower and higher eukaryotes. Apoptosis is an orchestrated form of cell death in which cells are actively involved in their own demise. Again, stress is a positive regulator of apoptosis and, in particular, of its apoptosome-mediated mitochondrial pathway. Besides discussing the individual roles played by the key molecules involved in autophagy in mammals in response to stress signals, we discuss here the interrelations between autophagy and apoptosis under these conditions.     

Molecular regulation of autophagy and apoptosis during ischemic and non-ischemic cardiomyopathy

A significant understanding of the genetic signaling pathways governing the extrinsic and intrinsic apoptotic pathways has been established. In recent years, the role of apoptosis in the heart during ischemic and non-ischemic cardiomyopathies has been under investigation and reported to contribute to ventricular remodeling and heart failure. Autophagy has been recently characterized as an essential cellular process in the heart, but whether autophagy functions as a pro-death or pro-survival program during disease conditions is still not completely understood. The mitochondrial death protein Bnip3 has been implicated in both apoptosis and autophagy, and its role in both processes is also discussed.     

Molecular regulation of autophagy and apoptosis during ischemic and non-ischemic cardiomyopathy

A significant understanding of the genetic signaling pathways governing the extrinsic and intrinsic apoptotic pathways has been established. In recent years, the role of apoptosis in the heart during ischemic and non-ischemic cardiomyopathies has been under investigation and reported to contribute to ventricular remodeling and heart failure. Autophagy has been recently characterized as an essential cellular process in the heart, but whether autophagy functions as a pro-death or pro-survival program during disease conditions is still not completely understood. The mitochondrial death protein Bnip3 has been implicated in both apoptosis and autophagy, and its role in both processes is also discussed.     

The engulfment receptor Draper is required for autophagy during cell death

Autophagy is a process to degrade and recycle cytoplasmic contents. Autophagy is required for survival in response to starvation, but has also been associated with cell death. How autophagy functions during cell survival in some contexts and cell death in others is unknown. Drosophila larval salivary glands undergo programmed cell death requiring autophagy genes, and are cleared in the absence of known phagocytosis. Recently, we demonstrated that Draper (Drpr), the Drosophila homolog of C. elegans engulfment receptor CED-1, is required for autophagy induction during cell death, but not during cell survival. drpr mutants fail to clear salivary glands. drpr knockdown in salivary glands prevents the induction of autophagy, and Atg1 misexpression in drpr null mutants suppresses salivary gland persistence. Surprisingly, drpr knockdown cell-autonomously prevents autophagy induction in dying salivary gland cells, but not in larval fat body cells following starvation. This is the first engulfment factor shown to function in cellular self-clearance, and the first report of a cell-death-specific autophagy regulator.Key words: autophagy, Draper, programmed cell death, engulfment, developmentProgrammed cell death is required for animal development and tissue homeostasis. Improper cell death leads to pathologies including autoimmunity and cancer. Several morphological forms of cell death occur during animal development, including apoptosis and autophagic cell death. Autophagic cell death is characterized by the presence of autophagosomes in dying cells that are not known to be engulfed by phagocytes. Autophagic cell death is observed during several types of mammalian developmental cell death, including regression of the corpus luteum and involution of mammary and prostate glands.During macroautophagy (autophagy), cytoplasmic components are sequestered by autophagosomes and delivered to the lysosome for degradation. Autophagy is a cellular response to stress required for survival in response to starvation. Whereas autophagy has been associated with cell death, it is unknown how autophagy is distinguished during cell death and cell survival. Autophagy is induced in Drosophila in response to starvation in the fat body where it promotes cell survival, while autophagy is induced by the steroid hormone ecdysone in salivary glands where it promotes cell death. This allows studies of autophagy in different cell types and in response to different stimuli.Drosophila larval salivary glands die with autophagic cell death morphology and autophagy is required for their degradation. Expression of the caspase inhibitor p35 enhances salivary gland persistence in Atg mutants, suggesting that caspases and autophagy function in parallel during salivary gland degradation. Either activation of caspases or Atg1 misexpression is sufficient to induce ectopic salivary gland clearance. We queried genome-wide microarray data from purified dying salivary glands and noted the induction of engulfment genes, those required for a phagocyte to consume and degrade a dying cell. We also noted few detectable changes in engulfment genes in Drosophila larvae during starvation.We found that Drpr, the Drosophila orthologue of C. elegans engulfment receptor CED-1, is enriched in dying salivary glands, and drpr null mutants have persistent salivary glands. Interestingly, whereas knockdown of drpr in phagocytic blood cells fails to influence salivary gland clearance, expression of drpr-RNAi in salivary glands prevents gland clearance. Drosophila drpr is alternatively spliced to produce three isoforms. We found that drpr-I-specific knockdown prevents salivary gland degradation and Drpr-I expression in salivary glands of drpr null mutants rescues salivary gland persistence. Therefore, drpr is autonomously required for salivary gland clearance. However, how Drpr is induced or activated during hormone-regulated cell death remains to be determined.drpr knockdown fails to influence caspase activation, and caspase inhibitor p35 expression in drpr null mutants enhances salivary gland persistence, suggesting that Drpr functions downstream or parallel to caspases in dying salivary glands. Interestingly, we found that drpr knockdown in salivary glands prevents the formation of GFP-LC3 puncta. Further, Atg1 misexpression in salivary glands of drpr null mutants suppresses salivary gland persistence. drpr is therefore required for autophagy induction in salivary glands, and Atg1 functions downstream of Drpr in this tissue. We found that several other engulfment genes are required for salivary gland degradation. However, the Drpr signaling mechanism leading to autophagy induction in salivary glands remains to be elucidated.We tested whether drpr is a general regulator of autophagy. The Drosophila fat body is a nutrient storage and mobilization organ akin to the mammalian liver, and is a well-established model to study starvation-induced autophagy. We found that drpr-RNAi expression in fat body clone cells fails to prevent GFP-Atg8 puncta formation in response to starvation. Similarly, drpr null fat body clone cells form Cherry-Atg8 puncta after starvation. Strikingly, drpr-RNAi expression in salivary gland clone cells inhibits the formation of GFP-Atg8 puncta. Therefore, drpr is cell-autonomously required for autophagy induction in dying salivary gland cells, but not for autophagy induction in fat body cells after starvation. These findings suggest that distinct signaling mechanisms regulate autophagy in response to nutrient deprivation compared to steroid hormone induction. Little is known about what distinguishes autophagy function in cell survival versus death. It is possible that varying levels of autophagy are induced during specific cell contexts and that high levels of autophagy could overwhelm a cell—leading to cell death. Autophagic degradation of specific cargo, such as cell death inhibitors, could also contribute to cell death.Given recent interest in manipulation of autophagy for therapies, it is possible that factors such as Drpr could be used as biomarkers to distinguish autophagy leading to cell death versus cell survival. While it is generally accepted that augmentation of protein clearance by autophagy during neurodegeneration would be beneficial, the role of autophagy in tumor progression is less clear. For example, monoallelic loss of the human Atg6 homolog beclin 1 is prevalent in human cancers, suggesting that autophagy is a tumorsuppressive mechanism. Thus, autophagy enhancers have been proposed for cancer prevention. However, autophagy occurs in tumor cells as a survival mechanism, and autophagy inhibitors have been proposed for anti-cancer therapies. Understanding how autophagy is regulated in different contexts is critical for appropriate therapeutic strategies.     

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.     

EDS1-mediated activation of autophagy regulates <Emphasis Type="Italic">Pst</Emphasis> DC3000 (<Emphasis Type="Italic">AvrRps4</Emphasis>)-induced programmed cell death in <Emphasis Type="Italic">Arabidopsis</Emphasis>

Autophagy is a conserved intracellular process through which cytoplasmic components are degraded and recycled under stress conditions. In the innate immunity of higher plants, autophagy has either pro-survival or pro-death functions in pathogen-induced programmed cell death (PCD). In aged leaves, autophagy negatively regulates PCD by eliminating redundant salicylic acid. However, in young leaves, the specific pro-death mechanisms of autophagy and signaling pathways related to the autophagic process have not been elucidated. Here, we demonstrate that enhanced disease susceptibility 1 (EDS1) mediated the activation of autophagy and played a key role in the pro-death mechanism of autophagy during avirulent Pst DC3000 (AvrRps4) infection. The path through which autophagosomes enter the vacuole was blocked. Additionally, formation of the ATG12–ATG5 complex and the level of enzymatic activity associated with ATG8 cleavage decreased in eds1 mutants. The expression of EDS1 in atg5 mutants was also much lower than that in wild-type plants during pathogen-triggered PCD. These findings implied that EDS1 may regulate autophagy by affecting the activities of the two ubiquitin-like protein-conjugating pathways. Moreover, autophagy may regulate immunity-related PCD by affecting the expression of EDS1 in young plants. Our results provide important insights into the mechanisms of EDS1 in autophagy during infection with avirulent Pst DC3000 (AvrRps4) in Arabidopsis.     

AMPK mediates a pro-survival autophagy downstream of PARP-1 activation in response to DNA alkylating agents

In this study we aim to elucidate the signaling pathway and biological function of autophagy induced by MNNG, a commonly used DNA alkylating agent. We first observed that MNNG is able to induce necrotic cell death and autophagy in Bax?/? Bak?/? double knockout MEFs. We analyzed the critical role of PARP-1 activation and ATP depletion in MNNG-mediated cell death and autophagy via AMPK activation and mTOR suppression. We provide evidence that suppression of AMPK blocks MNNG-induced autophagy and enhances cell death, suggesting the pro-survival function of autophagy in MNNG-treated cells. Taken together, data from this study reveal a novel mechanism in controlling MNNG-mediated autophagy via AMPK activation downstream of PARP-1 activation and ATP depletion.     

Oncogene-induced autophagy and the Goldilocks principle

Although several oncogenes enhance autophagic flux, the molecular mechanism and consequences of oncogene-induced autophagy remain to be clarified. We have recently shown that expression of oncogenic H-Ras (V12) promotes autophagy through upregulation of Beclin 1 and the BH3-only protein Noxa. H-Ras-expressing cells undergo autophagic cell death as a result of Noxa-mediated displacement of Mcl-1 and Bcl-xL from Beclin 1. Oncogenic H-Ras-induced death is attenuated through knockdown of BECLIN 1, ATG5, or ATG7, or through overexpression of Mcl-1, Bcl-2, Bcl-xL and their close relatives. These observations suggest that high-intensity oncogene activation may be selected against by promoting excessive autophagy, leading to cell death. Consequently, such oncogenes may select for cells with a reduced capacity for autophagy, either through loss of a BECLIN 1 allele or through upregulation of negative regulators of Beclin 1, such as Bcl-2 family members.