Autophagy in the control of programmed cell death

Programmed cell death (PCD) is essential for plant development and immunity. Localized PCD is associated with the hypersensitive response (HR), which is a constituent of a successful plant innate immune response. Plants have developed mechanisms to meticulously prevent HR-PCD lesions from spreading. Our understanding of these mechanisms is still in its incipient stages. A recent study demonstrated that autophagy, a universally conserved process of macromolecule turnover, plays a pivotal role in controlling HR-PCD. The molecular identity of the mediators between the PCD and HR pathways is still obscure, but recent work has begun to shed light on the relationship between HR-PCD and autophagy and to suggest possible mechanisms for the regulation of these pathways.     

Self-consuming innate immunity in Arabidopsis

Programmed cell death (PCD) associated with the pathogen-induced hypersensitive response (HR) is a hallmark of plant innate immunity. HR PCD is triggered upon recognition of pathogen effector molecules by host immune receptors either directly or indirectly via effector modulation of host targets. However, it has been unclear by which molecular mechanisms plants execute PCD during innate immune responses. We recently examined HR PCD in autophagy-deficient Arabidopsis knockout mutants (atg) and find that PCD conditioned by one class of plant innate immune receptors is suppressed in atg mutants. Intriguingly, HR triggered by another class of immune receptors with different genetic requirements is not compromised, indicating that only a specific subset of immune receptors engage the autophagy pathway for HR execution. Thus, our work provides a primary example of autophagic cell death associated with innate immune responses in eukaryotes as well as of pro-death functions for the autophagy pathway in plants.     

Autophagy and plant innate immunity

Plant innate immunity is often associated with specialized programmed cell death at or near the site of pathogen infection. Despite the isolation of several lesion mimic mutants, the molecular mechanisms that regulate cell death during an immune response remain obscure. Recently, autophagy, an evolutionarily conserved process of bulk protein and organelle turnover, was shown to play an important role in limiting cell death initiated during plant innate immune responses. Consistent with its role in plants, several studies in animals also demonstrate that the autophagic machinery is involved in innate as well as adaptive immunities. Here, we review the role of autophagy in plant innate immunity. Because autophagy is observed in healthy and dying plant cells, we will also examine whether autophagy plays a protective or a destructive role during an immune response.     

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.     

Arabidopsis ATG6 is required to limit the pathogen-associated cell death response

To begin to understand the interplay between autophagy and the hypersensitive response (HR), a type of programmed cell death (PCD) induced during plant innate immunity, we generated ATG6 antisense plants in the genetically tractable Arabidopsis thaliana system. AtATG6 antisense (AtATG6-AS) plants senesce early and are sensitive to nutrient starvation, suggestive of impairment of autophagic function in these plants. Additionally, these plants exhibited multiple developmental abnormalities, a phenomenon not observed in other AtATG mutants. AtATG6-AS plants produced fewer Monodansylcadaverine (MDC) and LysoTracker (LT) stained-autolysosomes in response to carbon and nitrogen starvation indicating that AtATG6 plays a role in the autophagic pathway in Arabidopsis. Interestingly, the level of AtATG6 mRNA in wild type Col-0 Arabidopsis plants is increased during the early phase of virulent and avirulent Pseudomonas syringae pv tomato (Pst) DC3000 infection suggesting that AtATG6 plays an important role during pathogen infection. In AtATG6-AS plants, HR-PCD induced upon infection with avirulent Pst DC3000 carrying the AvrRpm1 effector protein is not able to be contained at the infection site and spreads into uninfected tissue. Additionally, the disease-associated cell death induced by the infection of virulent Pst DC3000 bacteria is also partially misregulated in AtATG6-AS plants. Therefore, the AtATG6 antisense plants characterized here provide an excellent genetic model system to elucidate the molecular mechanisms by which autophagy regulates pathogen-induced cell death.     

The perplexing role of autophagy in plant innate immune responses

Autophagy is a major intracellular process for the degradation of cytosolic macromolecules and organelles in the lysosomes or vacuoles for the purposes of regulating cellular homeostasis and protein and organelle quality control. In complex metazoan organisms, autophagy is highly engaged during the immune responses through interfaces either directly with intracellular pathogens or indirectly with immune signalling molecules. Studies over the last decade or so have also revealed a number of important ways in which autophagy shapes plant innate immune responses. First, autophagy promotes defence‐associated hypersensitive cell death induced by avirulent or related pathogens, but restricts unnecessary or disease‐associated spread of cell death. This elaborate regulation of plant host cell death by autophagy is critical during plant immune responses to the types of plant pathogens that induce cell death, which include avirulent biotrophic pathogens and necrotrophic pathogens. Second, autophagy modulates defence responses regulated by salicylic acid and jasmonic acid, thereby influencing plant basal resistance to both biotrophic and necrotrophic pathogens. Third, there is an emerging role of autophagy in virus‐induced RNA silencing, either as an antiviral collaborator for targeted degradation of viral RNA silencing suppressors or an accomplice of viral RNA silencing suppressors for targeted degradation of key components of plant cellular RNA silencing machinery. In this review, we summarize this important progress and discuss the potential significance of the perplexing role of autophagy in plant innate immunity.     

活性氧对植物自噬调控的研究进展

自噬是一种在真核生物中高度保守的降解细胞组分的生物过程, 在饥饿、衰老和病菌感染等过程中起关键作用。而活性氧是有氧生物在正常或胁迫条件下产生的一种代谢副产物, 在植物的生长发育、胁迫适应和程序性细胞死亡过程中起重要作用。最新研究结果表明, 当植物受到病菌感染产生超敏反应时活性氧和自噬在程序性细胞死亡、生长发育和胁迫适应过程中起重要调控作用。因此, 该文结合最新的研究进展, 从活性氧的种类及特点、自噬的分子基础以及活性氧在植物自噬中的作用等方面, 探讨了活性氧与植物自噬之间的信号转导关系。     

Autophagy in antiviral innate immunity

Several autonomous arms of innate immunity help cells to combat viral infections. One of these is autophagy, a central cytosolic lysosomal‐dependent catabolic process constitutively competent to destroy infectious viruses as well as essential viral components that links virus detection to antiviral innate immune signals. Ongoing autophagy can be upregulated upon virus detection by pathogen receptors, including membrane bound and cytosolic pattern recognition receptors, and may further facilitate pattern recognition receptor‐dependent signalling. Autophagy or autophagy proteins also contribute to the synthesis of antiviral innate type I interferon cytokines as well as to antiviral interferon γ signalling. Additionally, autophagy may play a crucial role during viral infections in containing an excessive cellular response by regulating the intensity of the inflammatory response. As a consequence, viruses have evolved strategies to counteract antiviral innate immunity through manipulation of autophagy. This review highlights recent findings on the cross‐talk between autophagy and innate immunity during viral infections.     

From Arabidopsis to cereal crops: Conservation of chloroplast protein degradation by autophagy indicates its fundamental role in plant productivity

Autophagy is an evolutionarily conserved process leading to the degradation of intracellular components in eukaryotes, which is important for nutrient recycling especially in response to starvation conditions. Nutrient recycling is an essential process that underpins productivity in crop plants, such that remobilized nitrogen derived from older organs supports the formation of new organs or grain-filling within a plant. We extended our understanding of autophagy in a model plant, Arabidopsis thaliana, to an important cereal, rice (Oryza sativa). Through analysis of transgenic rice plants stably expressing fluorescent marker proteins for autophagy or chloroplast stroma, we revealed that chloroplast proteins are partially degraded in the vacuole via Rubisco-containing bodies (RCBs), a type of autophagosomes containing stroma. We further reported evidence that the RCB pathway functions during natural leaf senescence to facilitate subsequent nitrogen remobilization into newly expanding leaves. Thus, our recent studies establish the importance of autophagy in biomass production of cereals.     

Autophagy controls plant basal immunity in a pathogenic lifestyle-dependent manner

Plant genomes harbor autophagy-related (ATG) genes that encode major components of the eukaryotic autophagic machinery. Autophagy in plants has been functionally linked to senescence, oxidative stress adaptation and the nutrient starvation response. In addition, plant autophagy has been assigned negative (‘anti-death’) and positive (‘pro-death’) regulatory functions in controlling cell death programs that establish sufficient immunity to microbial infection. The role of autophagy in plant disease and basal immunity to microbial infection has, however, not been studied in detail. We have employed a series of autophagy-deficient genotypes of the genetic model plant Arabidopsis thaliana in various infection systems. Genotypes lacking ATG5, ATG10 or ATG18a develop spreading necrosis and enhanced disease susceptibility upon infection with toxin-producing pathogens preferring a necrotrophic lifestyle. These findings suggest that autophagy positively controls the containment of host tissue integrity upon infections by host-destructive microbes. In contrast, autophagy-deficient genotypes exhibit markedly increased immunity to infections by biotrophic pathogens through altered homeostasis of the plant hormone salicylic acid, thus suggesting an additional negative regulatory role of autophagy in plant basal immunity. In sum, our findings suggest that the role of plant autophagy in immunity cannot be generalized, and depends critically on the lifestyle and infection strategy of invading microbes.     

ATG5 is required to limit cell death induced by <Emphasis Type="Italic">Pseudomonas syringae</Emphasis> in <Emphasis Type="Italic">Arabidopsis</Emphasis> and may be mediated by the salicylic acid pathway

Autophagy can be regarded as a protection mechanism to restrict programmed cell death (PCD) induced by pathogen infection during plant innate immunity in the early stages. Autophagy related 5 (ATG5) plays an important role in autophagy in Arabidopsis. We investigated the function of ATG5 in Arabidopsis in the hypersensitive response (HR)-PCD elicited by both virulent and avirulent strains of Pseudomonas syringae pv. tomato bacteria DC3000. Results show that ATG5 plays a vital role in limiting HR induced by P. syringae strains and colocalizes with autophagic bodies during the early phase of bacterial infection. In addition, the P. syringae-induced response is mediated by the salicylic acid (SA) signaling pathway. In summary, ATG5 is required for limiting HR-PCD induced in Arabidopsis by P. syringae strains and may be mediated by SA signaling.     

Variations on a theme: plant autophagy in comparison to yeast and mammals

Autophagy is an evolutionary conserved process of bulk degradation and nutrient sequestration that occurs in all eukaryotic cells. Yet, in recent years, autophagy has also been shown to play a role in the specific degradation of individual proteins or protein aggregates as well as of damaged organelles. The process was initially discovered in yeast and has also been very well studied in mammals and, to a lesser extent, in plants. In this review, we summarize what is known regarding the various functions of autopahgy in plants but also attempt to address some specific issues concerning plant autophagy, such as the insufficient knowledge regarding autophagy in various plant species other than Arabidopsis, the fact that some genes belonging to the core autophagy machinery in various organisms are still missing in plants, the existence of autophagy multigene families in plants and the possible operation of selective autophagy in plants, a study that is still in its infancy. In addition, we point to plant-specific autophagy processes, such as the participation of autophagy during development and germination of the seed, a unique plant organ. Throughout this review, we demonstrate that the use of innovative bioinformatic resources, together with recent biological discoveries (such as the ATG8-interacting motif), should pave the way to a more comprehensive understanding of the multiple functions of plant autophagy.     

Mitophagy in plants

Mitochondria play a central role in primary metabolism in plants as well as in heterotrophic eukaryotes. Plants must control the quality and number of mitochondria in response to a changing environment, across cell types and developmental stages. Mitophagy is defined as the degradation of mitochondria by autophagy, an evolutionarily conserved system for the removal and recycling of intracellular components. Recent studies have highlighted the importance of mitophagy in plant stress responses. This review article summarizes our current knowledge of plant mitophagy and discusses the underlying mechanisms. In plants, chloroplasts cooperate with mitochondria for energy production, and autophagy also targets chloroplasts through a process known as chlorophagy. Advances in plant autophagy studies now allow a comparative analysis of the autophagic turnover of mitochondria and chloroplasts, via the selective degradation of their soluble proteins, fragments, or entire organelles.     

Autophagy in plants

Autophagy is a highly conserved processing mechanism in eukaryotes whereby cytoplasmic components are engulfed in double-membrane vesicles called autophagosomes and are delivered into organelles such as lysosomes (mammal) or vacuoles (yeast/plant) for degradation and recycling of the resulting molecules. Isolation of yeastAUTOPHAGY (ATG) genes has facilitated the identification of correspondingArabidopsis ATG genes based on sequence similarity. Genetic and molecular analyses using knockout and/or knockdown mutants of those genes have unraveled the biological functions of autophagy during plant development, nutrient recycling, and environmental stress responses. Additional roles for autophagy have been suggested in the degradation of oxidized proteins during oxidative stress and the regulation of hypersensitive response (HR)-programmed cell death (PCD) during innate immunity. Our review summarizes knowledge about the structure and function of autophagic pathways andATG components, and the biological roles of autophagy in plants.     

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.     

Autophagy regulates programmed cell death during the plant innate immune response

The plant innate immune response includes the hypersensitive response (HR), a form of programmed cell death (PCD). PCD must be restricted to infection sites to prevent the HR from playing a pathologic rather than protective role. Here we show that plant BECLIN 1, an ortholog of the yeast and mammalian autophagy gene ATG6/VPS30/beclin 1, functions to restrict HR PCD to infection sites. Initiation of HR PCD is normal in BECLIN 1-deficient plants, but remarkably, healthy uninfected tissue adjacent to HR lesions and leaves distal to the inoculated leaf undergo unrestricted PCD. In the HR PCD response, autophagy is induced in both pathogen-infected cells and distal uninfected cells; this is reduced in BECLIN 1-deficient plants. The restriction of HR PCD also requires orthologs of other autophagy-related genes including PI3K/VPS34, ATG3, and ATG7. Thus, the evolutionarily conserved autophagy pathway plays an essential role in plant innate immunity and negatively regulates PCD.     

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.     

Autophagy is required for tolerance of drought and salt stress in plants

Autophagy is a protein degradation process in which cells recycle cytoplasmic contents when subjected to environmental stress conditions or during certain stages of development. Upon the induction of autophagy, a double membrane autophagosome forms around cytoplasmic components and delivers them to the vacuole or lysosome for degradation. In plants, autophagy has been shown previously to be induced during abiotic stresses including nutrient starvation and oxidative stress. In this paper, we demonstrate the induction of autophagy in high salt and osmotic stress conditions, concomitant with the upregulation of expression of an Arabidopsis thaliana autophagy-related gene AtATG18a. Autophagy-defective RNAi-AtATG18a plants are more sensitive to salt and drought conditions than wild-type plants, demonstrating a role for autophagy in the response to these stresses. NADPH oxidase inhibitors block autophagy induction upon nutrient starvation and salt stress, but not during osmotic stress, indicating that autophagy can be activated by NADPH oxidase-dependent or -independent pathways. Together our results indicate that diverse environmental stresses can induce autophagy and that autophagy is regulated by distinct signaling pathways in different conditions.     

Relationship between growth arrest and autophagy in midgut programmed cell death in Drosophila

Autophagy has been implicated in both cell survival and programmed cell death (PCD), and this may explain the apparently complex role of this catabolic process in tumourigenesis. Our previous studies have shown that caspases have little influence on Drosophila larval midgut PCD, whereas inhibition of autophagy severely delays midgut removal. To assess upstream signals that regulate autophagy and larval midgut degradation, we have examined the requirement of growth signalling pathways. Inhibition of the class I phosphoinositide-3-kinase (PI3K) pathway prevents midgut growth, whereas ectopic PI3K and Ras signalling results in larger cells with decreased autophagy and delayed midgut degradation. Furthermore, premature induction of autophagy is sufficient to induce early midgut degradation. These data indicate that autophagy and the growth regulatory pathways have an important relationship during midgut PCD. Despite the roles of autophagy in both survival and death, our findings suggest that autophagy induction occurs in response to similar signals in both scenarios.     

Autophagy and innate immunity: Triggering,targeting and tuning

Autophagy is a conserved catabolic stress response pathway that is increasingly recognized as an important component of both innate and acquired immunity to pathogens. The activation of autophagy during infection not only provides cell-autonomous protection through lysosomal degradation of invading pathogens (xenophagy), but also regulates signaling by other innate immune pathways. This review will focus on recent advances in our understanding of three major areas of the interrelationship between autophagy and innate immunity, including how autophagy is triggered during infection, how invading pathogens are targeted to autophagosomes, and how the autophagy pathway participates in “tuning” the innate immune response.