Apoptosis (2014) 19:905–921 DOI 10.1007/s10495-014-0981-4

ORIGINAL PAPER

The roles of autophagy in development and stress responses in Arabidopsis thaliana Xin Lv • Xiaojun Pu • Gongwei Qin Tong Zhu • Honghui Lin

•

Published online: 30 March 2014 Ó Springer Science+Business Media New York 2014

Abstract Autophagy is a dynamic process that involves the recycling process of the degradation of intracellular materials. Over the past decade, our molecular and physiological understanding of plant autophagy has greatly been increased. Most essential autophagic machineries are conserved from yeast to plants. The roles that autophagyrelated genes (ATGs) family play in the lifecycle of the Arabidopsis are proved to be similar to that in mammal. Autophagy is activated during certain stages of development, senescence or in response to starvation, or environmental stress in Arabidopsis. In the progression of autophagy, ATGs act as central signaling regulators and could develop sophisticated mechanisms to survive when plants are suffering unfavorable environments. It will facilitate further understanding of the molecular mechanisms of autophagy in plant. In this review, we will discuss recent advances in our understanding of autophagy in Arabidopsis, areas of controversy, and highlight potential future directions in autophagy research. Keywords Autophagy
Arabidopsis
ATGs
PCD
Biotic and abiotic stress Abbreviation ACBP3 Acyl-CoA-binding protein Atg Autophagy-related gene CMA Chaperon-mediated-autophagy HR Hypersensitive response 3-MA 3-Methyladenine

X. Lv
X. Pu
G. Qin
T. Zhu
H. Lin (&) Key Laboratory of Bio-resources & Eco-environment (Ministry of Education), College of Life Science, Sichuan University, Chengdu 610064, China e-mail: [email protected]

PAS PCD PE PI3P PKA RCB ROS SNF TOR TSPO VPS PO

Pre-autophagosomal structure Programmed cell death Phosphatidylethanolamine Phosphatidyl-inositol-3 phosphate Protein kinase A Rubisco-containing bodies Reactive oxygen species Sucrose non-fermenting Target of rapamycin Tryptophan-rich sensory protein-related Vacuolar protein sorting Peroxisomes

Introduction Autophagy, a term from the Greek ‘auto’ (self) and ‘phagein’ (to eat), is a macromolecule degradation nonselectively pathway that toxic components and recycling of needed nutrients during specific developmental processes or encountering stress [1]. Over the past decade, our molecular and physiological understanding of autophagy in plants has greatly developed [2–4]. Most of the essential machinery required for autophagy seems to be conserved from yeast to plants. Depending on the different mechanisms of transport to lysosome/vacuole, three classes of autophagy have been described: macro-autophagy, micro-autophagy and chaperone-mediated autophagy (CMA) [5, 6]. ‘Macro-autophagy’ is characterized by two markedly features: travelling of autophagosomes and fusing of autophagosomes with lysosome. And macroautophagic uptake of intracellular proteins is mediated by the evagination of membrane associated with the rough endoplasmic reticulum (ER) [7]. ‘Micro-autophagy’ is another form of autophagy which makes cytosolic

123

906

Apoptosis (2014) 19:905–921

Fig. 1 Illustration of electron micrographs of representative Arabidopsis cells grown for 12 h in control medium supplemented with 1.5 % sucrose (a) or in sucrose-free medium (b). Cells were fixed, embedded in epoxy resin, sectioned and stained for EM as described in the ‘‘Materials and methods’’ section [21]. a Control cell. The cytoplasm contains globular vacuoles and numerous organelles including ER and Golgi stacks; *starchgrains in the plastid.

b Sucrose starving cell. Numerous vesicles are accumulated in the cytoplasm (black arrowheads) together with typical double-membrane-bound autophagosomes (enlarged inset). Note the disappearance of starch grains and the scarce ER profiles. Inset white arrowheads point to the inner and outer membranes of the autophagosome. G Golgi, V vacuole. Scalebar 1 lm (a, b) and 0. 25 lm (inset)

components directly engulf by the lysosome itself through invagination of the lysosomal membrane [8]. CMA is a selective process during which proteins with a KFERQ consensus peptide sequence are recognized by a chaperone/ co-chaperone complex and delivered to the lytic compartment in an unfolded state [9–11]. The process of macroautophagy (‘autophagy’ hereafter) the most studied form of autophagy, could be divided into five stages: initiation, nucleation, elongation and completion, fusing and degradation [12]. Typical triggers of autophagy include nutrient-deprivation and intracellular stress, especially reactive oxygen species (ROS) [13]. When autophagy occurred, complex mechanisms take part in forming a portion of cytoplasm including organelles, known as the autophagosome. Later, the autophagosome fuses with lysosome which undergoes degradation. Finally, once macromolecules are degraded in the lysosome/vacuole, monomeric units such as amino acids were released into the cytosol for reuse. Most plant cells at least have two different types of vacuoles, lytic and storage. The lytic vacuole is considered to be functionally equivalent to the yeast vacuole or animal lysosome. In plants, autophagy can be induced by a variety of stresses, including pathogen infection and senescence [14–18]. The cargo which comes from the autophagy is degraded into its building blocks, helping the cell to economize its resources, eliminate old/ damaged organelles, and survive in types of stress [19–21]. Here is an illustration of electron microscope which shows

the autophagy-related structures in the cytoplasm of cells deprived of sucrose for 12 h (Fig. 1b), while such clusters were not seen in the control cells (Fig. 1a) [21]. The core autophagy genes (Atgs), are capable of orchestra the formation of autophagosome during the process of autophagy. Atgs were originally founded in yeast when it was regarded as a model to study autophagy in eukaryotic cells [22, 23], homologues to Atg genes have previously discussed in lower eukaryotes like fungi and algae to plants and metazoan [4, 24]. Almost 36 Atgs have been identified with their respectively unique role in promoting autophagosome formation in yeast [3, 25]. Atgs can be divided into mainly three classes due to their different functions [26]: (1) the ATG1-ATG13 kinase complex; (2) ATG9 complex and ATG6/vps30 complex; (3) ATG5ATG12 complex and ATG8-PE complex (two ubiquitinlike conjugation systems). These studies in yeast have greatly facilitated the identification of homologous genes in plants that are required for autophagy, ATGs and the investigation of their molecular functions [2, 18, 27]. In this review, we will briefly summarize the recent advances in our understanding of plant autophagy, including the molecular mechanisms of autophagy and particularly Arabidopsis autophagy with a timeline (Fig. 2) and the factors which induce autophagy of Arabidopsis (Fig. 3). We will list the physiological roles of the Atgs and related proteins in different unfavorable environments, and finally discuss their mechanisms.

123

Apoptosis (2014) 19:905–921

907

Fig. 2 Timeline: a history of the autophagy in plants

Fig. 3 Several factors induce autophagy in Arabidopsis

Function of autophagy in the stress response Role of autophagy in Arabidopsis abiotic stresses Nutrient and starvation Autophagy is required for maintenance of the cellular viability under nutrient-limited conditions. The first and most common abiotic stress shown to induce autophagy was nutrient deprivation [28, 29]. In addition, autophagy is required for the degradation of damaged or toxic material that can be generated as a result of ROS accumulation during oxidative stress. Vacuolar autophagy was observed within 24 h of sucrose starvation in Arabidopsis suspension cells, with increased expression of vacuolar proteases. It is similar to be required for degradation of cytoplasmic components delivered to the vacuole, and thus for nutrient recycling [30]. Autophagy is activated under starvation conditions in Arabidopsis root cells [31] and accompanying the formation of autolysosomes [32]. Hydrotropic stimulation confers water stress in the roots, which also triggers an autophagic response responsible to the degradation of amyloplasts in columella cells of Arabidopsis roots [33]. In addition, the ATG1/13 complex

in Arabidopsis also showed that it is both a regulator and a target of autophagy. It seems that the turnover of the ATG1/13 kinase provides a dynamic mechanism to tightly connect autophagy to a nutritional status in Arabidopsis [34]. In most plants, a large fraction of photo-assimilated carbon is stored in the chloroplasts during the day such as starch and is remobilized during the subsequent night to support metabolism. T-DNA insertion of AtAtg7 mutant plants grow like the wild type under normal conditions. They show accelerated senescence and a highly sensitive response to nutrient limiting growth conditions [28, 35, 36]. Chloroplast autophagy in living cells and direct evidence of chloroplast transportation into the vacuole in an autophagy-defective Arabidopsis mutant, Atg4a4b-1 has been reported [37]. Under nitrogen or carbon-starvation conditions, chlorosis was observed earlier in AtAtg9-1 cotyledons and rosette leaves compared with wild-type plants, which was due to that the senescence-associated genes SEN1 and YSL4 were up-regulated in AtAtg9-1 before induction of senescence in AtAtg9-1 plants [29]. A close relationship between the degradation of chloroplast proteins via Rubisco-containing bodies (RCBs) and leaf carbon is specifically controlled in autophagy [38].

123

908

Growth and development As we know most of the Atg mutants are able to complete their life cycles [15, 28, 39, 40]. But, recent evidences show that deficiency of autophagy leads to significant changes of metabolic profiles in Arabidopsis [41]. Although autophagy has been investigated most extensively during nutrient-deficient conditions, plants maintain a basal level of autophagy even under favorable growth conditions [31, 42]. This basal autophagy may continually eliminate garbage proteins and organelles under normal growth conditions. Increasing evidence has demonstrated that autophagy is essential for the viability in plants. Transgenic lines with reduced AtAtg18a expression show hypersensitivity to sucrose and nitrogen starvation and premature senescence, both during natural senescence of leaves and in a detached leaf assay in Arabidopsis [16]. In addition, stromal protein degradation is incomplete in two autophagy T-DNA insertion mutants (Atatg5 and Atatg7) undergoing natural senescence [43]. Both ATG2 and ATG5 proteins are essential for autophagy while ATG9 protein contributes to, but is not essential for autophagy in Arabidopsis root tip cells. Autophagy is sensitive to 3-methyladenine and dependent on constitutive occurrence of ATG proteins in the root tip cells of Arabidopsis [42]. In addition, Arabidopsis in homologs deleted on chromosome 10 (PTEN), a protein and lipid dual phosphatase homologous to animal PTENs, regulates autophagy in pollen tubes by disrupting the dynamics of phosphatidylinositol 3-phosphate (PI3P) [44]. Autophagy is crucial to plant development. AtAtg6, the Arabidopsis homolog of yeast Atg6/Vps30, and mammalian and tobacco Beclin 1 proteins, lead to pollen germination defects when disrupted [40, 45]. In addition for root cells, autophagy is also observed and crucial to the differentiation of tracheary elements. With an Arabidopsis cell culture in vitro system, autophagy is activated during tracheary elements differentiation [46]. RabG3b, as a component of autophagy, regulates TE differentiation [47]. Besides, ATG8-interacting proteins (ATI1 and ATI2) are partially associated with the ER membrane network overexpressed or suppressed in both ATI1 and ATI2, respectively, stimulating or inhibiting seed germination in the presence of the germination-inhibiting hormone abscisic acid [48]. Salt, drought and other abiotic stress In Arabidopsis, autophagy is induced by multiple abiotic stresses, including nutrient deficiency, oxidative, salt, drought and other stresses [10, 16, 49, 50]. In plants, high salt and drought stress are the most common environmental stresses. In contrast to ion stress resulting from high salt,

123

Apoptosis (2014) 19:905–921

drought stress leads to osmotic stress in plants [51]. In addition, ER stress responses have been well characterized in animals and yeast [52], and autophagy has been suggested to play an important role in recovery from ER stress [53]. These stresses can create oxidation damage, leading to the accumulation of ROS and oxidized proteins [54]. The ability of autophagy to scavenge oxidized proteins and regulate the ROS levels suggests that autophagy may also be involved in abiotic stresses. Autophagy is regulated by distinct pathways during different abiotic stresses. Under these conditions, one common characteristic is the production of ROS, which can act as signal molecules to activate stress response and defense pathways. High salinity and drought stress are two of the most common environmental stresses encountered by plants, and have some common features but are regulated differently [55, 56]. High salinity differs from drought stress in creating ionic stress in addition to osmotic stress, including Na? toxicity and K? deficiency; osmotic stress can cause membrane disruption and enzyme dysfunction [51, 57]. AtAtg8 in Arabidopsis and OsAtg10b in rice function effectively in response to salt stress and osmotic stress [58]. In addition, autophagy-deprived AtAtg18a-RNAi plants show high sensitivity to these two stresses. ROS may function in the induction of autophagy by salt and osmotic stress, they tried to block ROS formation by adding an NADPH oxidase inhibitor. The results show that the NADPH oxidase inhibitors inhibit autophagy induction in plants under high salts stress, but unexpectedly not under osmotic stress; this proves the existence of NADPH oxidase dependent and -independent pathways for regulating autophagy [49]. At the same time, both stresses can cause oxidative damage to the cell, probably increasing the production of ROS and damaged proteins. Evidences show that autophagy is induced by high salinity and osmotic stresses, and that autophagy-defective plants are more sensitive to these conditions. Role of autophagy in Arabidopsis biotic stresses Innate immunity: hypersensitive response (HR) and programmed cell death (PCD) Plants, unlike mammals, are lack of mobile defender cells and a somatic adaptive immune system. Instead, they rely on the innate immunity of each cell and on systemic signals emanating from infection site. It is well known that there are two essential branches of the plant innate immune system. One is the transmembrane pattern recognition receptors that respond to slowly evolved microbial- or pathogen-associated molecular patterns. The other acts largely inside the cells, using the polymorphic protein products encoded by most plant resistance (R) proteins. In

Apoptosis (2014) 19:905–921

animals, PCD can be morphologically divided into three classes: apoptosis, autophagic cell death, and programmed necrosis. Some studies show that PCD occurred with concomitant activation of autophagy, rather than that the PCD process carried out by autophagy [59]. A significant conundrum that has emerged in recent years revolves around the observation that autophagy as it plays a Janus role in tumor cells, present itself as double edge of sword in plant life. Autophagy may have been assigned ‘pro-death’ and ‘pro-survival’ roles in controlling PCD associated with microbial effector-triggered immunity [2]. And it contributes to plant immunity to different pathogens that can be mechanistically diverse, thus resembling the complex role of this process in animal innate immunity [60]. Recent researches have indicated that autophagy cooperates with jasmonate-mediated and WRKY33-mediated signaling pathways in the regulation of plant defense responding to necrotrophic pathogens [61]. Autophagy has also been reported to play important roles in suppression of cell death and defense response to the biotrophic pathogen, the powdery mildew fungus through ATG2 and ATG18 in Arabidopsis [62]. In addition, ATG7 contributes to resistance to fungal pathogens. ATG7dependent autophagy constitutes an ‘‘anti-death’’ plant mechanism to control the containment of cell death and immunity to necrophic fungal infection [63]. The HR is to prevent the spread of infection by microbial pathogens in plants. The HR is characterized by the rapid death of cells in the local region surrounding an infection. Recently, HR-PCD was observed in autophagydeficient Arabidopsis knockout mutants (Atgs) and the data showed that HR induced PCD conditioned by a specific subset of immune receptors is suppressed in Atg mutants [64]. More specifically, HR cell death initiated by Toll Interleukin-1 (TIR)-type immune receptors through the defense regulator EDS1 is suppressed in Atg mutant plants. Furthermore, the reports demonstrated that PCD triggered by coiled-coil-type immune receptors via NDR1 is either autophagy-independent or engages autophagic components with cathepsins and other unidentified cell death mediators [65]. In AtAtg6 antisense plants, HR-PCD induced by infection with avirulent Pseudomonas syringae pv. tomato DC3000(PstDC3000) carrying the AvrRpm1 effector protein is not able to be limited to the infection site, but to spreads into uninfected tissue [39]. Recent studies have indicated that plant autophagy operates a novel negative feedback loop modulating salicylic acid (SA) signaling to negatively regulate senescence and immunity-related PCD [66]. In AtAtg2-2 mutant it showed enhanced resistance to powdery mildew and dramatic mildew-induced cell death as well as early senescence phenotypes in the absence of pathogens [67]. Genotypes lacking ATG5, ATG10 or ATG18a develop

909

spreading necrosis and enhanced disease susceptibility upon infection with toxin-producing pathogens preferring a necrotrophic lifestyle. In contrast, autophagy-deficient genotypes exhibit markedly increased immunity to infections by biotrophic pathogens through altered homeostasis of the plant hormone SA, thus suggesting an additional negative regulatory role of autophagy in Arabidopsis basal immunity [68]. Research on plant PCD has focused mainly on two categories: PCD during normal development and PCD during the HR triggering by pathogen infection. Evidence implies that autophagy plays critical roles in both processes. An understanding of this dual role of autophagy is essential for learning how the process is controlled [69]. Besides, transgenic plants over expressing a constitutively active RabG3b (RabG3bCA) displayed accelerated, unrestricted HR PCD within 1d of infection, in contrast to the autophagy-defective AtAtg5-1 mutant, which gradually developed chlorotic cell death through uninfected sites over several days [70]. As noted by Codogno, ‘Our knowledge of the balanced role of autophagy in cell survival and cell death will benefit studies in regulation.’ [71]. ROS-modulated autophagy ROS are highly reactive oxygen free radicals or non-radical molecules that are generated by multiple mechanisms, with the nicotinamide adenine dinucleotide phosphate oxidases (NOX) [72]. These ROS, as important multifaceted signaling molecules, can regulate a number of cellular pathways and thus playing critical roles [73]. ROS and autophagy have been historically associated with cell death. However, more recent evidence indicates that both ROS and autophagy play important roles in signaling and cellular adaptation to stress. ROS are chemically reactive molecules containing oxygen, and highly toxic materials that accumulate in large amounts under various environmental stress conditions and/or during developmental stages. For example, when a plant recognizes an attacking pathogen, one of the first induced reactions is to rapidly produce superoxide or hydrogen peroxide to strengthen the cell wall [67]. Studies in plants and algae have demonstrated that autophagy is structurally and functionally conserved in photosynthetic organisms and plays an important role in the cellular response and adaptation to different stress conditions that involve the generation of ROS, such as oxidative and drought stresses, pathogen infection, or photooxidative damage. In the cell, mitochondria act as the primary energy-generating source that regulates several critical cellular processes. As we know that mitochondria are known to play the key roles in triggering cell death via disrupting electron transport and energy metabolism, releasing or activating autophagy-related proteins, as well

123

910

as altering cellular redox potential [74]. Upon infection, ROS are generated at the plasmamembrane by NOX, although other organelles such asmitochondria, chloroplasts, and/or peroxisomes maintain enhanced ROS production [75, 76]. Evidences show that the mitochondrial electron transport chains are the primary sources of ROS production in most eukaryotes. Autophagy dependent chloroplast degradation may be the other source of ROS as well as the pathogen-response signaling molecules that induce the defense response. Autophagy plays a role in chloroplast degradation in Arabidopsis during avirulent Pst DC3000 (AvrRps4) infection [77]. In addition, peroxisomes are the other primary sources of ROS. The central role of peroxisomes in the generation and scavenging of H2O2 has been well known ever since their discovery almost five decades ago [78]. In plants, peroxisomes may play an important role in redox signaling and in the control of redox homeostasis, since they constitute a major source of H2O2 but can also produce O2radicals or nitric oxide [79, 80]. Pexophagy has not yet been demonstrated in plants; therefore, the physiological role of peroxisomes degradation and its implication in redox signaling remains unclear. The chloroplast is a primary source of ROS in plants and algae, and it may be targeted for degradation in a process called chlorophagy [37, 81]. It is less complicated to sum up the pivotal functions of mitochondria or lysosomes than of peroxisomes (PO). The former are notably the powerhouse of a cell providing it with ATP, and contribute to the initiation of apoptosis when this vital task becomes severely impaired. Lysosomes are the rubbish chute of a cell degrading. Both external metabolites taken up by endocytosis, cellular constituents like organelles or protein complexes internalized by autophagy. An example is seen during pexophagy in the yeast Pichiapastoris, where the receptor protein for peroxisomes, PpAtg30, is phosphorylated. Through phosphorylated residues, PpAtg30 interacts with other autophagy machinery components, such as PpAtg11, targeting peroxisomes for autophagic breakdown [82]. A number of studies have been carried out on ROS and its various functions in plants, especially in the realm of Arabidopsis and autophagy. Recently, accumulating data have demonstrated that massive ROS generated may critically mediate several core autophagic pathways in plants. For example, multiple atg mutants are known to be hypersensitive to abiotic stresses [27, 83–85]. Oxidative stress induces autophagy in Arabidopsis seedlings with an AtAtg18a-dependent manner and that RNAi-AtAtg18a transgenic lines, which are defective in autophagosome formation are hypersensitive to ROS [86]. Besides, new technologies have been used that two macroautophagy markers, monodansylcadaverine and green fluorescent

123

Apoptosis (2014) 19:905–921

protein-AtAtg8e, whereas how that application of hydrogen peroxide or the reactive oxidative species inducer methyl viologen can induce macroautophagy in Arabidopsis [10]. Conversely, under a presumably normal condition, AtAtg2 and AtAtg5 mutant leaves accumulate a higher level of hydrogen peroxide than wild-type leaves, implying that autophagy normally limits ROS [66]. The accumulation of ROS may indicate cellular damage and tissue necrosis, because of the lack of autophagy. ROS can elevate to a terrible level when the plants are in additional abiotic stresses. The mechanism of this hypersensitivity is not clear, but ROS may be an important link, which is leading to an improved understanding of the multifaceted redox interactions with the plant hormone signaling network. ROS is a common mediator of plant responses to control plant growth and defense [73, 87]. The mechanisms underlying the induction of autophagy by ROS likely involve one or several target proteins that undergo ROStriggered redox post translational modifications affecting their activity and resulting in the induction of autophagy. Identifying these ROS-controlled autophagy regulators and delineating the underlying molecular mechanism will likely represent a major challenge in the field for coming years. Roles of autophagy-related genes (Atgs) in Arabidopsis A milestone in our understanding of the regulation of autophagy was the identification of the Atgs. To date, more than 30 kinds Atgs of the yeast have been well characterized, while there are 36 core autophagy genes identified in Arabidopsis [3] In yeast, some of the encoded proteins assemble together in a dynamic pre-autophagosomal structure (PAS) that binds to, and organizes, the phagophore [88, 89]. The stages of autophagosome formation and delivery to the vacuole has suggested that two distinct pathways exist in which autophagosomes can either fuse directly with the vacuole or can first be engulfed by a smaller lysosome-like or endosome-like organelle in the plant [90, 91]. And then, the autophagosome then transports the cargo to the vacuole. During the fusion process, the outer autophagosome membrane fuses with the vacuole membrane, and the remaining single-membrane structure (termed an autophagic vacuole) is delivered inside the vacuole. The autophagic vacuoles are then broken down by vacuolar hydrolases, and the products are exported from the vacuole to the cytoplasm for reuse (Fig. 4). And it is showed that the pathway of autophagy in Arabidopsis. Biochemical and cell biological characterization of Atgs resealed a model of autophagic processes that are controlled by several classes of Atgs (Fig. 4). There are three main classes of core ATG proteins: ATG1-13 complex, ATG6/vps30 complex and Ubiquitin-like Systems. They

Apoptosis (2014) 19:905–921

911

Fig. 4 Schematic representation of known or proposed steps within the autophagy (ATG)-mediated autophagic system in Arabidopsis. a Autophagy in Arabidopsis proceeds through a series of steps, including initiation at the induction, elongation, closure and completion of the autophagosome, autophagosome maturation via docking and fusion, breakdown and degradation of the autophagosome inner membrane and cargo, and recycling of the resulting macromolecules. b The pathway of autophagy in Arabidopsis

123

912

are highly conserved in various eukaryotes [92]. All of them are significant for Arabidopsis autophagy in the life. ATG1-13 complex The prior studies using Arabidopsis thaliana as the model have identified a mechanistically similar ATG autophagic system in plants [34, 93]. It is described that the ATG1 and ATG13 subunits show their regulation by starvation. When the plant detects the signals, it transmits the signals to the autophagosomes, generating apparatus known as the PAS. Studies from yeast identified the ATG1/13 kinase complex as a key positive regulator of autophagy [88, 94]. The core components are the Ser/Thr kinase ATG1 and its accessory proteins ATG2, ATG13 and ATG18 [34, 95]. The activity of the ATG1-13 complex is regulated by various upstream kinase cascades that affect the interaction strength among the subunits and the kinase activity of ATG1. One of them is the target of rapamycin (TOR). And its target recognition cofactor RAPTOR, protein kinase A (PKA), and the sucrose non-fermenting (SNF)-1 kinase (AKIN10/AKIN11 in Arabidopsis), converge to control the pivotal ATG1/ ATG13 Ser/Thr kinase complex [96–98]. The assembled and ‘activated’ ATG1-ATG13 kinase complex stimulates several steps related to autophagic vesiculation either directly or indirectly. One step involves the vacuolar-protein-sorting (VPS)-34/ATG6 (Beclin1)/ ATG14/VPS15 lipid kinase complex that presumably decorates the phagophore with phosphatidyl-inositol-3 phosphate (PI3P). The PI3-kinase inhibitors, wortmannin, LY294002 and 3-methyladenine (3-MA), impair phagophore formation by blocking VPS34 action [99]. Why PI3P is needed but not yet clear, but one possibility is that it helps distinguish the developing phagophore from other endomembrane compartments during assembly, and then provides a mark recognized by the machinery that fuses autophagosomes with the tonoplast. Another step engages the transmembrane protein ATG9, which helps deliver lipids to the expanding phagophore in conjunction with the peripheral ATG2 and ATG18 proteins. In yeast and animals, TOR kinase acted as a negative regulator of autophagy [100]. The TOR protein is conserved in plants and RAPTOR homologues have also been identified in Arabidopsis [101, 102]. However, disruption of the TOR gene is lethal and causes an early block in embryo development [103], and there is one TOR gene named AtTORC1 in Arabidopsis [96]. During starvation, TOR is inhibited, which triggers hypophosphorylation of ATG13 and hyperphosphorylation of ATG1, followed by the association of ATG1 and ATG13 with each other and several other factors [Atg11p and Atg17p in yeast, and the focal adhesion kinase family-interacting protein of 200 kDa (FIP200) and ATG101 in mammals and probably plants] [104].

123

Apoptosis (2014) 19:905–921

ATG 6/vps30 complex Yeast autophagy protein 6 (ATG6/Vps30) has been reported to be essential for the formation of autophagosomes during starvation [105]. Mammalian homolog of yeast ATG6 is recognized as Beclin-1, which has been reported to be involved in the regulation of autophagy and anti-apoptotic pathways [106]. The plant ortholog of yeast ATG6 was first identified in Nicotiana tobacco during the screening of genes affecting hypersensitive-response-PCD induced by tobacco mosaic virus; this suggests that ATG6 is a negative regulator of PCD and also that it is responsible for controlling virus replication [107]. Following the induction, ATG6 protein helps the elongation of the membrane to enclose autophagosomes. However, Arabidopsis ATG6 (AtAtg6) T-DNA insertion mutants exhibited defects in pollen germination [40, 45]. Otherwise, AtATG6 antisense (AtAtg6-AS) plants senesce early and are sensitive to nutrient starvation, suggestive of impairment of autophagic function in these plants. Above mentioned information provide an excellent genetic model system to elucidate the molecular mechanisms by which autophagy regulates pathogen-induced cell death [108]. It is found that three homologs in the rice (Oryza sativa) genome (OsAtg6a, OsAtg6b, OsAtg6c) [83]. The analysis of gene expression in the different abiotic stresses and hormones describe an expression database in rice which can be considered to imitate in Arabidopsis. Ubiquitin-like systems In contrast to yeast, which has a single Atg8, Atg4, and Atg12 gene, Arabidopsis contains nine members of the AtAtg8 family (AtAtg8a–AtAtg8i), two members of the AtAtg4 family (AtAtg4a, AtAtg4b), and two members of the AtAtg12 family (AtAtg12a, AtAtg12b) [28]. And there is two ubiquitin-like systems: ATG8 complex and ATG12-5 complex. Following the expanding phagophore, the next step revolves around decorating the phagophore with ATG8. It becomes anchored in the membrane by a signature conjugation pathway analogous to, but separate from, ubiquitylation, which attaches the lipid phosphati-dylethanolamine (PE) to its carboxyl terminus. Similar to other members of the Ub-fold family, ATG8 (or LC3/GABARAP/GATE-16 in mammals) contains a b-grasp fold at its core, comprising a concave five-stranded b sheet cradling a diagonal helix, from which protrudes a short carboxy-terminal extension ending in the catalytically essential glycine. ATG8 is typically synthesized as a longer precursor that requires processing by the ATG4 protease to expose this residue. Mature ATG8 is activated by an ATPdependent E1-activating enzyme ATG7, which subsequently binds ATG8 to a conserved cysteine within ATG7

Apoptosis (2014) 19:905–921

via a thioester linkage. The bound ATG8 is then donated from ATG7 to the E2-conjugating enzyme ATG3 by transesterification, and finally attached to PE with the aid of an ATG8-specific E3 ligase complex [109]. The ATG8PE ligase is uniquely assembled by a parallel conjugation pathway involving another Ub-fold protein ATG12, its target ATG5, and the ATG16 scaffold (Fig. 4). Sequential actions of ATG7 and the ATG12-specific E2 ATG10 connect ATG12 to ATG5 via an isopeptide bond between its C-terminal glycine and a conserved lysine in ATG5. The ATG12–ATG5 conjugate associates with the phagophore through the dimeric ATG16 protein, and the resulting hexameric complex then directs lipidation of ATG3bound ATG8 [28, 34]. The ATG–PE adduct coats the expanding phagophore membrane, whereas the ATG12– ATG5/ATG16 multimer, along with the rest of the PAS, dissociates after the autophagosome is formed. ATG8–PE lining the outer membrane is eventually delipidated by ATG4 and released, whereas ATG8–PE lining the inner membrane is consumed in the vacuole [110]. Although ATG5 is considered the main target of ATG12, recent data in mammalian cell suggest that ATG3 is also modified [111]. Among all the Atg genes, ATG8s appear to be functional and useful markers to track autophagosome formation and

913

docking in the yeast and mammal [112, 113]. Their dynamics in vivo suggest that formation of the ATG12– ATG5 conjugate precedes formation of the ATG8–PE conjugate, and enhances this lipidation reaction via crosstalk between the two conjugation reactions [114, 115]. The actual function of the ATG12–ATG5 and ATG8–PE conjugates are unknown, but at the morphological level they are necessary for autophagic engulfment. The Arabidopsis counterpart of ATG8 was reported to interact with microtubules, suggesting that the ATG8–PE conjugate assists in transporting autophagosomes to the vacuole via interaction with the microtubule to skeleton. In Arabidopsis, a family of nine Atg8 genes, termed AtAtg8a to 8i, has been identified, while AtAtg12 has 2 family members AtAtg12a and AtAtg12b [29]. The proteins encoded by this family show between high identity(average 70 %) with each other and from high (average 60 %) identity with the yeast protein. The yeast two-hybrid experiment of the AtAtg8a and AtAtg8d shows a similar pathway occurs in plants. Moreover, AtAtg8 be involved in linking the autophagy pathway to the microtubule network and that the microtubules are responsible for the relocation of autophagosomes to the vacuole [116]. All Arabidopsis Atg8 genes have identical expression patterns and are induced by nitrogen or sucrose

Fig. 5 Amino acid sequences of yeast, mammals and A. thaliana ATG8 proteins

123

Gene names

At2g37840

At3g53930

At3g6196

At3g19190

At5g61500

At2g44140

At3g59950

At5g17290

At3g61710

At5g45900

ATG family

ATG1a

123

ATG1b

ATG1c

ATG2

ATG3

ATG4a

ATG4b

ATG5

ATG6

ATG7

Ubiquitin-like modifier activating enzyme atg7

Beclin-1-like protein

Autophagy protein 5

Cysteine protease ATG4a Cysteine protease ATG4b

Autophagy-related protein3

Autophagy-related protein 2

Protein kinase family protein

Unc51-like kinase

Unc51-like kinase

Protein names

ATG7_ARATH

F4JFF7_ARATH

BECN1_ARATH

A8MRV3_ARATH

ATG5_ARATH

F4J9I3_ARATH

ATG4B_ARATH

ATG4A_ARATH

Q56WG4_ARATH

ATG3_ARATH

F4JB41_ARATH F8S296_ARATH

Q9LJL0_ARATH

F4IX14_ARATH

Q94C95_ARATH

C0Z2C5_ARATH

Q8GWX7_ARATH

F4JBP3_ARATH

A8MR56_ARATH

F4IRW0_ARATH

Autophagy, defense response to fungus, leaf senescence, protein lipidation protein transport

Autophagic vacuole assembly, defense response to fungus, pollen germination, protein targeting to vacuole

Autophagy

Autophagy, protein transport

Autophagy, protein transport

Autophagy, protein transport

Autophagy, Protein transport

Q8RWS7_ARATH

Biological process

Gene ontology (GO)

Q5E914_ARATH

Entry name

Table 1 Gene and protein information about AtAtg gene family

Cytosol

Pre-autophagosomal structure

Cytoplasm

Autophagic vacuole, vacuolar lumen, chloroplast

Cytosol

Autophagic vacuole

Autophagic vacuole

Cellular component

Fungal pathogens; nutrient starvation; powdery mildew pathogen;Celldeath;PCD;HR

Cell death response HR PCD; autophagy, VPS and the glycosylphosphatidylinositol anchor system

Mediates vesicle nucleation;germinate the pollen;pathogen-associated

Fungal pathogen; powdery mildew resistanc; nutrient starvation

Cell death; PCD;HR necrotrophic

Expanding phagophore membrane;

Autophagy protein transport autophagosomesUbl conjugation pathway

Autophagy protein transport autophagosomes Ubl conjugation pathway

Cell death; PCD; HR; nutrient starvation

Promote phagophore expansion;

ATG2, ATG9, and ATG18

Nutrient starvation; ATG1 and its accessory proteins

Function

AtNBR1

ABA pathway

AtTSPO;RCBs; TIR, EDS1;

RCBs

Interaction gene

[63, 118, 125]

[42, 62, 65, 81, 128, 129]

[37, 116]

[34]

[34, 94]

Reference

914 Apoptosis (2014) 19:905–921

Gene names

At4g21980

At4g04620

At1g62040

At2g05630

At2g45170

At4g16520

At3g60640

At3g06420

At3g15580

At2g31260

At3g07525

At1g54210

At3g13970

At3g49590

At3g18770

At5g50230

ATG family

ATG8a

ATG8b

ATG8c

ATG8d

ATG8e

ATG8f

ATG8g

ATG8h

ATG8i

ATG9

ATG10

ATG12a

ATG12b

ATG13a

ATG13b

ATG16

Table 1 continued

At5g50230

Autophagy-related protein 13

Autophagy-related protein 13

Ubiquitin-like protein ATG12B

Ubiquitin-like protein ATG12A

Autophagocytosisassociated family protein

Protein autophagy 9

Autophagy-related protein 8i

Autophagy-related protein 8h

Autophagy-related protein 8g

Autophagy-related protein 8f

Autophagy-related protein 8e

Autophagy-related protein 8d

Autophagy-related protein 8c

Autophagy-related protein 8b

Autophagy-related protein 8a

Protein names

Q9FGS2_ARATH

Q6NNP0_ARATH

Q9LSA0_ARATH F4J8V5_ARATH

Q0WQ00_ARATH

F4IXZ6_ARATH

Q9SCK0_ARATH

AT12B_ARATH

AT12A_ARATH

Q67YB5_ARATH

F4JEH1_ARATH

Q8VZ52_ARATH

Q8RUS5_ARATH

Q9SJX0_ARATH

ATG8I_ARATH

ATG8H_ARATH

ATG8G_ARATH

ATG8F_ARATH

D7LBW5_ARALL

ATG8E_ARATH

F4IHC1_ARATH

ATG8D_ARATH

F4HX35_ARATH

ATG8C_ARATH

Autophagic vacuole assembly, autophagy, protein transport

Autophagy, defense response to fungus

Autophagy

Autophagy, protein transport

A8MS84_ARATH

Biological process

Gene ontology (GO)

ATG8A_ARATH ATG8B_ARATH

Entry name

Cytoplasm

Vacuolar lumen

Autophagic vacuole, mitochondrion CVT vesicle membrane, autophagic vacuole membrane, microtubule, mitochondrion,

Cellular component

Nutrient starvation

Expanding phagophore membrane; promoting atg8 lipidation with atg5; nutrient starvation

Involved in ATG12-ATG5 conjugate

Promote phagophoreexpansion; Celldeath; PCD; HR; nutrient starvation

Cell death; PCD;HR; nutrient starvation

(ATG12-ATG5,ATG8-PE) senescence;

Two conjugation systems

A marker protein of the PAS; vesicle expansion and fusion; conjugated to phosphatidylethanol-amine (PE) in a manner dependent on AtATG7, AtATG3 a tool with which to investigate the crosstalk mechanism between

Function

RCBs

ATI1and ATI2; Adi3;

RCBs

ACBP3; AtNBR1;

TSPO;

Interaction gene

[34, 94, 135]

[65, 117, 129, 135]

[14, 118]

[10, 34]

[10, 21, 38, 48, 117, 125, 130– 134]

Reference

Apoptosis (2014) 19:905–921 915

123

Gene names

At3g62770

At4g30510

At2g40810

At3g56440

At5g0515

At5g54730

At1g03380

At1g54710

At4g29380

At1g60490

ATG family

ATG18a

ATG18b

ATG18c

ATG18d

ATG18e

ATG18f

ATG18g

ATG18h

VPS15

VPS34

Table 1 continued

123

Phosphatidylinositol 3-kinase

Phosphoinositide-3kinase, regulatory subunit 4, p150

Autophagy-related protein 18h

Autophagy-related protein 18g

Autophagy-related protein 18e Autophagy-related protein 18f

Autophagy-related protein 18d

Autophagy-related protein 18c

Autophagy-related protein 18b

Autophagy-related protein 18a

Protein names

Phosphatidylinositol phosphorylation, phosphatidylinositolmediated signaling

Q8RXR0_ARATH Q56YM3_ARATH

Endocytosis, PI3P biosynthetic process, phosphatidylinositolmediated signaling, reactive oxygen species metabolic process, response to salt stress

pollen germination

Pollen development,

Response to starvation

Response to starvation

Autophagy, response to starvation

Autophagic cell death, autophagy, defense response to fungus, leaf senescence, response to starvation Macroautophagy

Biological process

Gene ontology (GO)

PI3K_ARATH

Q9M0E5_ARATH

Q8H1Q5_ARATH

Q9C5I7_ARATH

Q56W94_ARATH

Q8GUL1_ARATH

Q9FH32_ARATH

Q9FHK8_ARATH

Q0WPK3_ARATH

Q8GYD7_ARATH

O22195_ARATH

F4JQB6_ARATH

Q8H1Q8_ARATH

Q9M0A9_ARATH

F4IZI7_ARATH

Q9LZI8_ARATH D7LT96_ARALL

Q93VB2_ARATH

Entry name

Cul4-RING ubiquitin ligase complex

Cytosol, nucleus, organelle membrane PAS complex, cytosol organelle membrane

Cellular component Autophagosomeformation; necrotrophic fungal pathogen; oxidativestress; salt and osmotic stress; water stress; NADPH oxidase-dependent or independent pathways.

Function

PFD1.2

WRKY33;

Interaction gene

[10, 16, 33, 49, 60–62, 68, 86]

Reference

916 Apoptosis (2014) 19:905–921

Apoptosis (2014) 19:905–921

starvation [17, 21]. While the formation of the ATG12– ATG5 adduct is essential to Atg8-mediated autophagy in Arabidopsis by promoting Atg8 lipidation [115]. Additionally, the Arabidopsis acyl-CoA-binding protein ACBP3 has been reported to be a crucial regulator of the ATG8-PE complex through its interaction with PE [117]. The above statements support the hypothesis that the ATG conjugation systems in Arabidopsis are highly similar to those in yeast and mammals, in vitro conjugation systems of Arabidopsis will also afford a tool [118]. The cellular processes involving the Atg8 genes functions efficiently in young, non-senescing tissues, both under favorable growth conditions and starvation stresses in A. thaliana [119]. Furthermore, starvation stress stimulates the expression of GmAtg8i and ethylene signal-related genes. Since the ethylene signal is involved in senescence and various environmental stresses, it is possible that starvation stress-induced autophagy is partly mediated by the ethylene signaling in soybean [120]. Among all the evidence, we focused on ATG8 and ATG12 proteins in Arabidopsis. On the one hand, it is assumed that AtAtg8 have the analogous function with yeast and mammals. And the multiple-sequence alignment of ATG8 proteins with yeast and mammals counterparts showed higher similarity among themselves and to other plants than to animal/yeast homologs. On the other hand, it is hypothesized that ubiquitin-like system is similar with yeast and mammals (Fig. 5). ATG12 is conjugated to ATG5 in requires ATG7 and ATG10. ATG3, E2-like enzyme and ATG4 are conjugated to PE in a reaction [121]. The lapidated form of ATG8 is attached to the phagophore membrane for the autophagosome formation and fusion with a late lysosome. Taken together, the results suggest that ATG8 and ATG12 proteins have a high degree of homology domains with yeast and mammals. Despite of being considered as the marker of the autophagy, it hypothesizes that ATG8 proteins may have considerable functional homology. ATG12-5 complex and Atg8-PE complex are ubiquitinlike conjugation systems in mammals and yeast. Homology LC3 in mammals indicates that the ATG8-PE conjugate are essential for autophagosome formation and completion while ATG8-PE also mediates fusion of the autophagosome to the vacuole membrane [122]. Besides, this study shows some gene and protein information about ATG8 and related ATGs (Table 1) from NCBI database (http://www.ncbi.nlm.nih.gov/) and Uniprot database (http://www.uniprot.org/). According to all the findings mentioned previously, the role of AtAtg8s is significant to the autophagy in Arabidopsis within a variety of biological process.

917

Others Besides, autophagy has more roles in Arabidopsis. Various mutants that interfere with the autophagic process (Atg mutants) display lower numbers of Anthocyanic Vacuolar Inclusions (AVIs), in addition to a reduced accumulation of anthocyanins. Interestingly, vanadate increases the numbers of AVIs in the Atg mutants, suggesting that several pathways might participate in AVI formation. Taken together, the results suggest novel mechanisms for the formation of sub-vacuolar compartments capable of accumulating anthocyanin pigments [123]. In addition, a case study with the A. thaliana tryptophan-rich sensory proteinrelated (At-TSPO), a membrane protein transiently induced by abiotic stress, revealed a complex interaction between heme metabolism and selective autophagy resulting in down regulation of At-TSPO [124]. Arabidopsis NBR1 was reported to be an autophagy substrate degraded in the vacuole dependent on the polymerization property of the PB1 domain and expression of Arabidopsis Atg7 [125, 126]. Recently, it shows that the de-ubiquitinating enzyme AMSH1 and the ESCRT-III subunit VPS2.1 are required for autophagic degradation in Arabidopsis [127]. Autophagosomes are transported to, and ultimately fuse with, the tonoplast via a v-SNARE-type mechanism to discharge the internal vesicle as an autophagic body (Fig. 3). Eventual degradation of the limiting autophagic body membrane releases the cargo into the vacuolar lumen; the CVT cargo accumulates undigested, whereas autophagic cargo is catabolized by the diverse array of vacuolar hydrolases [17].

Conclusions and future directions One of the most recent advances in the molecular era of autophagy is the structure analysis of autophagy proteins, which began with the structure of the mammalian ATG8 homologue. And the structure of ATG12 has been detected in Arabidopsis. Structure–function analyses will probably be useful in understanding the mechanism of autophagy. Subsequently, combined with continued studies of the regulatory process, they might allow the rational design of drugs and the precise modulation that will be needed to use autophagy effectively in therapeutic intervention. In another aspect, the mechanism of autophagy in Arabidopsis is remained to be defined. Our understanding of the roles of autophagy in Arabidopsis has been benefited from the availability of all the above-mentioned data; however, there are significant unclear factors inherent in the complex molecular mechanisms and pathway of autophagy, and thus there is an urgent need for more additional key information. Therefore, further discoveries are being driven by an

123

918

abundance of structural information on the potential biotic and abiotic stresses resistance. Thereby, this model pathway that we hypothesize helps us to experimental verification. Intriguingly, an ongoing emergence of sophisticated mathematics models for the disruption of protein–protein interactions would be screened as potential key protein. These findings would provide a comprehensive perspective of further explanation of the roles of plant autophagy that may target PCD and other related pathways as potential stress resistant mechanisms in A. thaliana. Acknowledgments We thank Prof. Jinku Bao (Sichuan University) and Zhi Shi (Sichuan University) for their useful suggestions. This work was supported by the National Nature Science Foundation of China (91017004) and Doctoral Foundation of the Ministry of Education (20110181110059 and 20120181130008).

References 1. Deter RL, de Duve C (1967) Influence of glucagon, an inducer of cellular autophagy, on some physical properties of rat liver lysosomes. J Cell Biol 33:437–449 2. Liu Y, Bassham DC (2012) Autophagy: pathways for self-eating in plant cells. Annu Rev Plant Biol 63:215–237 3. Avin-Wittenberg T, Honig A, Galili G (2012) Variations on a theme: plant autophagy in comparison to yeast and mammals. Protoplasma 249:285–299 4. Diaz-Troya S, Perez-Perez ME, Florencio FJ, Crespo JL (2008) The role of TOR in autophagy regulation from yeast to plants and mammals. Autophagy 4:851–865 5. Massey AC, Zhang C, Cuervo AM (2006) Chaperone-mediated autophagy in aging and disease. Curr Top Dev Biol 73:205–235 6. Cuervo AM, Dice JF (2000) Age-related decline in chaperonemediated autophagy. J Biol Chem 275:31505–31513 7. Blommaart E, Luiken J, Meijer A (1997) Autophagic proteolysis: control and specificity. Histochem J 29:365–385 8. van Doorn WG, Woltering EJ (2005) Many ways to exit? Cell death categories in plants. Trends Plant Sci 10:117–122 9. Till A, Lakhani R, Burnett SF, Subramani S (2012) Pexophagy: the selective degradation of peroxisomes. Int J Cell Biol 2012:512721 10. Xiong Y, Contento AL, Nguyen PQ, Bassham DC (2007) Degradation of oxidized proteins by autophagy during oxidative stress in Arabidopsis. Plant Physiol 143:291–299 11. Johansen T, Lamark T (2011) Selective autophagy mediated by autophagic adapter proteins. Autophagy. 7:279–296 12. Mizushima N (2007) Autophagy: process and function. Genes Dev 21:2861–2873 13. Li ZY, Yang Y, Ming M, Liu B (2011) Mitochondrial ROS generation for regulation of autophagic pathways in cancer. Biochem Biophys Res Commun 414:5–8 14. Phillips AR, Suttangkakul A, Vierstra RD (2008) The ATG12conjugating enzyme ATG10 is essential for autophagic vesicle formation in Arabidopsis thaliana. Genetics 178:1339–1353 15. Thompson AR, Doelling JH, Suttangkakul A, Vierstra RD (2005) Autophagic nutrient recycling in Arabidopsis directed by the ATG8 and ATG12 conjugation pathways. Plant Physiol 138:2097–2110 16. Xiong Y, Contento AL, Bassham DC (2005) AtATG18a is required for the formation of autophagosomes during nutrient stress and senescence in Arabidopsis thaliana. Plant J 42:535–546

123

Apoptosis (2014) 19:905–921 17. Yoshimoto K, Hanaoka H, Sato S, Kato T, Tabata S, Noda T et al (2004) Processing of ATG8s, ubiquitin-like proteins, and their deconjugation by ATG4s are essential for plant autophagy. Plant Cell 16:2967–2983 18. Bassham DC (2007) Plant autophagy—more than a starvation response. Curr Opin Plant Biol 10:587–593 19. Bassham DC, Laporte M, Marty F, Moriyasu Y, Ohsumi Y, Olsen LJ et al (2006) Autophagy in development and stress responses of plants. Autophagy. 2:2–11 20. Aubert S, Gout E, Bligny R, Marty-Mazars D, Barrieu F, Alabouvette J et al (1996) Ultrastructural and biochemical characterization of autophagy in higher plant cells subjected to carbon deprivation: control by the supply of mitochondria with respiratory substrates. J Cell Biol 133:1251–1263 21. Rose TL, Bonneau L, Der C, Marty-Mazars D, Marty F (2006) Starvation-induced expression of autophagy-related genes in Arabidopsis. Biol Cell 98:53–67 22. Tsukada M, Ohsumi Y (1993) Isolation and characterization of autophagy-defective mutants of Saccharomyces cerevisiae. FEBS Lett 333:169–174 23. Thompson AR, Vierstra RD (2005) Autophagic recycling: lessons from yeast help define the process in plants. Curr Opin Plant Biol 8:165–173 24. Meijer WH, van der Klei IJ, Veenhuis M, Kiel JA (2007) ATG genes involved in non-selective autophagy are conserved from yeast to man, but the selective Cvt and pexophagy pathways also require organism-specific genes. Autophagy. 3:106–116 25. Pallauf K, Rimbach G (2013) Autophagy, polyphenols and healthy ageing. Ageing Res Rev 12:237–252 26. Yang Z, Klionsky DJ (2010) Eaten alive: a history of macroautophagy. Nat Cell Biol 12:814–822 27. Kwon SI, Park OK (2008) Autophagy in plants. J Plant Biol 51:313–320 28. Doelling JH, Walker JM, Friedman EM, Thompson AR, Vierstra RD (2002) The APG8/12-activating enzyme APG7 is required for proper nutrient recycling and senescence in Arabidopsis thaliana. J Biol Chem 277:33105–33114 29. Hanaoka H, Noda T, Shirano Y, Kato T, Hayashi H, Shibata D et al (2002) Leaf senescence and starvation-induced chlorosis are accelerated by the disruption of an Arabidopsis autophagy gene. Plant Physiol 129:1181–1193 30. Contento AL, Kim SJ, Bassham DC (2004) Transcriptome profiling of the response of Arabidopsis suspension culture cells to Suc starvation. Plant Physiol 135:2330–2347 31. Yano K, Suzuki T, Moriyasu Y (2007) Constitutive autophagy in plant root cells. Autophagy. 3:360–362 32. Oh-ye Y, Inoue Y, Moriyasu Y (2011) Detecting autophagy in Arabidopsis roots by membrane-permeable cysteine protease inhibitor E-64d and endocytosis tracer FM4-64. Plant Signal Behav 6:1946–1949 33. Nakayama M, Kaneko Y, Miyazawa Y, Fujii N, Higashitani N, Wada S et al (2012) A possible involvement of autophagy in amyloplast degradation in columella cells during hydrotropic response of Arabidopsis roots. Planta 236:999–1012 34. Suttangkakul A, Li F, Chung T, Vierstra RD (2011) The ATG1/ ATG13 protein kinase complex is both a regulator and a target of autophagic recycling in Arabidopsis. Plant Cell 23:3761–3779 35. Wang Y, Yu B, Zhao J, Guo J, Li Y, Han S et al (2013) Autophagy contributes to leaf starch degradation. Plant Cell 25:1383–1399 36. Wang Y, Liu Y (2013) Autophagic degradation of leaf starch in plants. Autophagy. 9:1247–1248 37. Wada S, Ishida H, Izumi M, Yoshimoto K, Ohsumi Y, Mae T et al (2009) Autophagy plays a role in chloroplast degradation during senescence in individually darkened leaves. Plant Physiol 149:885–893

Apoptosis (2014) 19:905–921 38. Izumi M, Wada S, Makino A, Ishida H (2010) The autophagic degradation of chloroplasts via rubisco-containing bodies is specifically linked to leaf carbon status but not nitrogen status in Arabidopsis. Plant Physiol 154:1196–1209 39. Patel S, Dinesh-Kumar SP (2008) Arabidopsis ATG6 is required to limit the pathogen-associated cell death response. Autophagy. 4:20–27 40. Qin G, Ma Z, Zhang L, Xing S, Hou X, Deng J et al (2007) Arabidopsis AtBECLIN 1/AtAtg6/AtVps30 is essential for pollen germination and plant development. Cell Res 17:249–263 41. Izumi M, Hidema J, Ishida H. (2013) Deficiency of autophagy leads to significant changes of metabolic profiles in Arabidopsis. Plant Signal Behav 8 42. Inoue Y, Suzuki T, Hattori M, Yoshimoto K, Ohsumi Y, Moriyasu Y (2006) AtATG genes, homologs of yeast autophagy genes, are involved in constitutive autophagy in Arabidopsis root tip cells. Plant Cell Physiol 47:1641–1652 43. Lee TA, Vande Wetering SW, Brusslan JA (2013) Stromal protein degradation is incomplete in Arabidopsis thaliana autophagy mutants undergoing natural senescence. BMC Res Notes 6:17 44. Zhang Y, Li S, Zhou LZ, Fox E, Pao J, Sun W et al (2011) Overexpression of Arabidopsis thaliana PTEN caused accumulation of autophagic bodies in pollen tubes by disrupting phosphatidylinositol 3-phosphate dynamics. Plant J 68: 1081–1092 45. Fujiki Y, Yoshimoto K, Ohsumi Y (2007) An Arabidopsis homolog of yeast ATG6/VPS30 is essential for pollen germination. Plant Physiol 143:1132–1139 46. Kwon SI, Cho HJ, Park OK (2010) Role of Arabidopsis RabG3b and autophagy in tracheary element differentiation. Autophagy. 6:1187–1189 47. Kwon SI, Cho HJ, Jung JH, Yoshimoto K, Shirasu K, Park OK (2010) The Rab GTPase RabG3b functions in autophagy and contributes to tracheary element differentiation in Arabidopsis. Plant J 64:151–164 48. Honig A, Avin-Wittenberg T, Ufaz S, Galili G (2012) A new type of compartment, defined by plant-specific Atg8-interacting proteins, is induced upon exposure of Arabidopsis plants to carbon starvation. Plant Cell 24:288–303 49. Liu Y, Xiong Y, Bassham DC (2009) Autophagy is required for tolerance of drought and salt stress in plants. Autophagy. 5: 954–963 50. Han S, Yu B, Wang Y, Liu Y (2011) Role of plant autophagy in stress response. Protein Cell 2:784–791 51. Zhu J-K (2001) Cell signaling under salt, water and cold stresses. Curr Opin Plant Biol 4:401–406 52. Liu Y, Bassham DC (2013) Degradation of the endoplasmic reticulum by autophagy in plants. Autophagy. 9:622–623 53. Pu Y, Bassham DC (2013) Links between ER stress and autophagy in plants. Plant Signal Behav 8:e24297 54. Tsugane K, Kobayashi K, Niwa Y, Ohba Y, Wada K, Kobayashi H (1999) A recessive Arabidopsis mutant that grows photoautotrophically under salt stress shows enhanced active oxygen detoxification. Plant Cell Online 11:1195–1206 55. Munns R (2002) Comparative physiology of salt and water stress. Plant Cell Environ 25:239–250 56. Song W-Y, Zhang Z-B, Shao H-B, Guo X-L, Cao H-X, Zhao H-B et al (2008) Relationship between calcium decoding elements and plant abiotic-stress resistance. Int J Biol Sci 4:116 57. Mahajan S, Tuteja N (2005) Cold, salinity and drought stresses: an overview. Arch Biochem Biophys 444:139–158 58. Slavikova S, Ufaz S, Avin-Wittenberg T, Levanony H, Galili G (2008) An autophagy-associated Atg8 protein is involved in the responses of Arabidopsis seedlings to hormonal controls and abiotic stresses. J Exp Bot 59:4029–4043

919 59. Kroemer G, El-Deiry W, Golstein P, Peter M, Vaux D, Vandenabeele P et al (2005) Classification of cell death: recommendations of the Nomenclature Committee on Cell Death. Cell Death Differ 12:1463–1467 60. Lenz HD, Haller E, Melzer E, Kober K, Wurster K, Stahl M et al (2011) Autophagy differentially controls plant basal immunity to biotrophic and necrotrophic pathogens. Plant J 66:818–830 61. Lai Z, Wang F, Zheng Z, Fan B, Chen Z (2011) A critical role of autophagy in plant resistance to necrotrophic fungal pathogens. Plant J 66:953–968 62. Wang Y, Wu Y, Tang D (2011) The autophagy gene, ATG18a, plays a negative role in powdery mildew resistance and mildewinduced cell death in Arabidopsis. Plant Signal Behav 6: 1408–1410 63. Lenz HD, Vierstra RD, Nurnberger T, Gust AA (2011) ATG7 contributes to plant basal immunity towards fungal infection. Plant Signal Behav 6:1040–1042 64. Hofius D, Mundy J, Petersen M (2009) Self-consuming innate immunity in Arabidopsis. Autophagy. 5:1206–1207 65. Hofius D, Schultz-Larsen T, Joensen J, Tsitsigiannis DI, Petersen NH, Mattsson O et al (2009) Autophagic components contribute to hypersensitive cell death in Arabidopsis. Cell 137: 773–783 66. Yoshimoto K, Jikumaru Y, Kamiya Y, Kusano M, Consonni C, Panstruga R et al (2009) Autophagy negatively regulates cell death by controlling NPR1-dependent salicylic acid signaling during senescence and the innate immune response in Arabidopsis. Plant Cell 21:2914–2927 67. Wang Y, Nishimura MT, Zhao T, Tang D (2011) ATG2, an autophagy-related protein, negatively affects powdery mildew resistance and mildew-induced cell death in Arabidopsis. Plant J 68:74–87 68. Lenz HD, Haller E, Melzer E, Gust AA, Nurnberger T (2011) Autophagy controls plant basal immunity in a pathogenic lifestyle-dependent manner. Autophagy. 7:773–774 69. Hayward AP, Tsao J, Dinesh-Kumar SP (2009) Autophagy and plant innate immunity: defense through degradation. Semin Cell Dev Biol 20:1041–1047 70. Kwon SI, Cho HJ, Kim SR, Park OK (2013) The Rab GTPase RabG3b positively regulates autophagy and immunity-associated hypersensitive cell death in Arabidopsis. Plant Physiol 161: 1722–1736 71. Klionsky DJ (2007) Autophagy: from phenomenology to molecular understanding in less than a decade. Nat Rev Mol Cell Biol 8:931–937 72. Huang J, Lam GY, Brumell JH (2011) Autophagy signaling through reactive oxygen species. Antioxid Redox Signal 14:2215–2231 73. Foyer CH, Noctor G (2013) Redox signaling in plants. Antioxid Redox Signal 18:2087–2090 74. Heo J-M, Rutter J (2011) Ubiquitin-dependent mitochondrial protein degradation. Int J Biochem Cell Biol 43:1422–1426 75. Torres MA, Jones JD, Dangl JL (2005) Pathogen-induced, NADPH oxidase–-derived reactive oxygen intermediates suppress spread of cell death in Arabidopsis thaliana. Nat Genet 37:1130–1134 76. Torres MA, Jones JD, Dangl JL (2006) Reactive oxygen species signaling in response to pathogens. Plant Physiol 141:373–378 77. Dong J, Chen W (2013) The role of autophagy in chloroplast degradation and chlorophagy in immune defenses during Pst DC3000 (AvrRps4) infection. PLoS One 8:e73091 78. Schrader M (2009) Anniversaries, peroxisomes and reactive oxygen species. Histochem Cell Biol 131:435–436 79. Foyer CH, Noctor G (2009) Redox regulation in photosynthetic organisms: signaling, acclimation, and practical implications. Antioxid Redox Signal 11:861–905

123

920 80. del Rı´o LA (2011) Peroxisomes as a cellular source of reactive nitrogen species signal molecules. Arch Biochem Biophys 506:1–11 81. Ishida H, Yoshimoto K, Izumi M, Reisen D, Yano Y, Makino A et al (2008) Mobilization of Rubisco and stroma-localized fluorescent proteins of chloroplasts to the vacuole by an ATG gene-dependent autophagic process. Plant Physiol 148:142–155 82. Farre J-C, Manjithaya R, Mathewson RD, Subramani S (2008) PpAtg30 tags peroxisomes for turnover by selective autophagy. Dev Cell 14:365–376 83. Rana RM, Dong S, Ali Z, Huang J, Zhang HS (2012) Regulation of ATG6/Beclin-1 homologs by abiotic stresses and hormones in rice (Oryza sativa L.). Genet Mol Res (GMR) 11:3676–3687 84. Izumi M, Hidema J, Makino A, Ishida H (2013) Autophagy contributes to nighttime energy availability for growth in Arabidopsis. Plant Physiol 161:1682–1693 85. Dobrenel T, Marchive C, Azzopardi M, Clement G, Moreau M, Sormani R et al (2013) Sugar metabolism and the plant target of rapamycin kinase: a sweet operaTOR? Front Plant Sci. 4:93 86. Xiong Y, Contento AL, Bassham DC (2007) Disruption of autophagy results in constitutive oxidative stress in Arabidopsis. Autophagy. 3:257–258 87. Mittler R, Vanderauwera S, Gollery M, Van Breusegem F (2004) Reactive oxygen gene network of plants. Trends Plant Sci 9:490–498 88. Suzuki K, Kubota Y, Sekito T, Ohsumi Y (2007) Hierarchy of Atg proteins in pre-autophagosomal structure organization. Genes Cells 12:209–218 89. Suzuki K, Ohsumi Y (2007) Molecular machinery of autophagosome formation in yeast, (Saccharomyces cerevisiae). FEBS Lett 581:2156–2161 90. Toyooka K, Moriyasu Y, Goto Y, Takeuchi M, Fukuda H, Matsuoka K (2006) Protein aggregates are transported to vacuoles by macroautophagic mechanism in nutrient-starved plant cells. Autophagy. 2:96–106 91. van Doorn WG, Papini A (2013) Ultrastructure of autophagy in plant cells: a review. Autophagy. 9:1922–1936 92. Yang Z, Klionsky DJ (2010) Mammalian autophagy: core molecular machinery and signaling regulation. Curr Opin Cell Biol 22:124–131 93. Bassham DC (2009) Function and regulation of macroautophagy in plants. Biochim Biophys Acta (BBA) 1793:1397–1403 94. Mizushima N (2010) The role of the Atg1/ULK1 complex in autophagy regulation. Curr Opin Cell Biol 22:132–139 95. Suzuki K, Ohsumi Y (2010) Current knowledge of the preautophagosomal structure (PAS). FEBS Lett 584:1280–1286 96. Liu Y, Bassham DC (2010) TOR is a negative regulator of autophagy in Arabidopsis thaliana. PLoS One 5:e11883 97. Kim J, Kundu M, Viollet B, Guan KL (2011) AMPK and mTOR regulate autophagy through direct phosphorylation of Ulk1. Nat Cell Biol 13:132–141 98. Robaglia C, Thomas M, Meyer C (2012) Sensing nutrient and energy status by SnRK1 and TOR kinases. Curr Opin Plant Biol 15:301–307 99. Takatsuka C, Inoue Y, Matsuoka K, Moriyasu Y (2004) 3-Methyladenine inhibits autophagy in tobacco culture cells under sucrose starvation conditions. Plant Cell Physiol 45:265–274 100. Pattingre S, Espert L, Biard-Piechaczyk M, Codogno P (2008) Regulation of macroautophagy by mTOR and Beclin 1 complexes. Biochimie 90:313–323 101. Garrett A, Bruce V, Maureen H (2005) The Arabidopsis AtRaptor genes are essential for post-embryonic plant growth. BMC Biol 3:12 102. Deprost D, Truong H-N, Robaglia C, Meyer C (2005) An Arabidopsis homolog of RAPTOR/KOG1 is essential for early

123

Apoptosis (2014) 19:905–921

103.

104.

105.

106. 107.

108.

109. 110.

111.

112.

113.

114.

115.

116.

117.

118.

119.

120.

embryo development. Biochem Biophys Res Commun 326: 844–850 Menand B, Desnos T, Nussaume L, Berger F, Bouchez D, Meyer C et al (2002) Expression and disruption of the Arabidopsis TOR (target of rapamycin) gene. Proc Natl Acad Sci USA 99:6422–6427 Alers S, Loffler AS, Wesselborg S, Stork B (2012) Role of AMPK-mTOR-Ulk1/2 in the regulation of autophagy: cross talk, shortcuts, and feedbacks. Mol Cell Biol 32:2–11 Kametaka S, Okano T, Ohsumi M, Ohsumi Y (1998) Apg14p and Apg6/Vps30p form a protein complex essential for autophagy in the yeast Saccharomyces cerevisiae. J Biol Chem 273:22284–22291 Sinha S, Levine B (2008) The autophagy effector Beclin 1: a novel BH3-only protein. Oncogene 27:S137–S148 Liu Y, Schiff M, Czymmek K, Tallo´czy Z, Levine B, DineshKumar S (2005) Autophagy regulates programmed cell death during the plant innate immune response. Cell 121:567–577 Harrison-Lowe NJ, Olsen LJ (2008) Autophagy protein 6 (ATG6) is required for pollen germination in Arabidopsis thaliana. Autophagy 4(3) Ohsumi Y (2001) Molecular dissection of autophagy: two ubiquitin-like systems. Nat Rev Mol Cell Biol 2:211–216 Kirisako T, Ichimura Y, Okada H, Kabeya Y, Mizushima N, Yoshimori T et al (2000) The reversible modification regulates the membrane-binding state of Apg8/Aut7 essential for autophagy and the cytoplasm to vacuole targeting pathway. J Cell Biol 151:263–276 Radoshevich L, Murrow L, Chen N, Fernandez E, Roy S, Fung C et al (2010) ATG12 conjugation to ATG3 regulates mitochondrial homeostasis and cell death. Cell 142:590–600 Mizushima N, Yamamoto A, Matsui M, Yoshimori T, Ohsumi Y (2004) In vivo analysis of autophagy in response to nutrient starvation using transgenic mice expressing a fluorescent autophagosome marker. Mol Biol Cell 15:1101–1111 Suzuki K, Kirisako T, Kamada Y, Mizushima N, Noda T, Ohsumi Y (2001) The pre-autophagosomal structure organized by concerted functions of APG genes is essential for autophagosome formation. EMBO J 20:5971–5981 Hanada T, Noda NN, Satomi Y, Ichimura Y, Fujioka Y, Takao T et al (2007) The Atg12-Atg5 conjugate has a novel E3-like activity for protein lipidation in autophagy. J Biol Chem 282:37298–37302 Chung T, Phillips AR, Vierstra RD (2010) ATG8 lipidation and ATG8-mediated autophagy in Arabidopsis require ATG12 expressed from the differentially controlled ATG12A AND ATG12B loci. Plant J 62:483–493 Ketelaar T, Voss C, Dimmock SA, Thumm M, Hussey PJ (2004) Arabidopsis homologues of the autophagy protein Atg8 are a novel family of microtubule binding proteins. FEBS Lett 567:302–306 Xiao S, Chye ML (2010) The Arabidopsis thaliana ACBP3 regulates leaf senescence by modulating phospholipid metabolism and ATG8 stability. Autophagy. 6:802–804 Fujioka Y, Noda NN, Fujii K, Yoshimoto K, Ohsumi Y, Inagaki F (2008) In vitro reconstitution of plant Atg8 and Atg12 conjugation systems essential for autophagy. J Biol Chem 283:1921–1928 Slavikova S, Shy G, Yao Y, Glozman R, Levanony H, Pietrokovski S et al (2005) The autophagy-associated Atg8 gene family operates both under favourable growth conditions and under starvation stresses in Arabidopsis plants. J Exp Bot 56:2839–2849 Okuda M, Nang MP, Oshima K, Ishibashi Y, Zheng SH, Yuasa T et al (2011) The ethylene signal mediates induction of

Apoptosis (2014) 19:905–921

121. 122.

123.

124.

125.

126.

127.

128.

GmATG8i in soybean plants under starvation stress. Biosci Biotechnol Biochem 75:1408–1412 Kim SH, Kwon C, Lee JH, Chung T (2012) Genes for plant autophagy: functions and interactions. Mol Cells 34:413–423 Mizushima N, Yoshimori T, Ohsumi Y (2011) The role of Atg proteins in autophagosome formation. Annu Rev Cell Dev Biol 27:107–132 Pourcel L, Irani NG, Lu Y, Riedl K, Schwartz S, Grotewold E (2010) The formation of anthocyanic vacuolar inclusions in Arabidopsis thaliana and implications for the sequestration of anthocyanin pigments. Mol Plant 3:78–90 Vanhee C, Batoko H (2011) Arabidopsis TSPO and porphyrins metabolism: a transient signaling connection? Plant Signal Behav 6:1383–1385 Svenning S, Lamark T, Krause K, Johansen T (2011) Plant NBR1 is a selective autophagy substrate and a functional hybrid of the mammalian autophagic adapters NBR1 and p62/ SQSTM1. Autophagy. 7:993–1010 Zhou J, Wang J, Cheng Y, Chi YJ, Fan B, Yu JQ et al (2013) NBR1-mediated selective autophagy targets insoluble ubiquitinated protein aggregates in plant stress responses. PLoS Genet 9:e1003196 Katsiarimpa A, Kalinowska K, Anzenberger F, Weis C, Ostertag M, Tsutsumi C et al (2013) The deubiquitinating enzyme AMSH1 and the ESCRT-III subunit VPS2.1 are required for autophagic degradation in Arabidopsis. Plant Cell 25:2236–2252 Vanhee C, Batoko H (2011) Autophagy involvement in responses to abscisic acid by plant cells. Autophagy. 7:655–656

921 129. Izumi M, Ishida H (2011) The changes of leaf carbohydrate contents as a regulator of autophagic degradation of chloroplasts via Rubisco-containing bodies during leaf senescence. Plant Signal Behav. 6:685–687 130. Vanhee C, Zapotoczny G, Masquelier D, Ghislain M, Batoko H (2011) The Arabidopsis multistress regulator TSPO is a heme binding membrane protein and a potential scavenger of porphyrins via an autophagy-dependent degradation mechanism. Plant Cell. 23:785–805 131. Contento AL, Xiong Y, Bassham DC (2005) Visualization of autophagy in Arabidopsis using the fluorescent dye monodansylcadaverine and a GFP-AtATG8e fusion protein. Plant J. 42:598–608 132. Xiao S, Gao W, Chen QF, Chan SW, Zheng SX, Ma J et al (2010) Overexpression of Arabidopsis acyl-CoA binding protein ACBP3 promotes starvation-induced and age-dependent leaf senescence. Plant Cell. 22:1463–1482 133. Devarenne TP (2011) The plant cell death suppressor Adi3 interacts with the autophagic protein Atg8 h. Biochem Biophys Res Commun. 412:699–703 134. Ishida H, Yoshimoto K (2008) Chloroplasts are partially mobilized to the vacuole by autophagy. Autophagy. 4:961–962 135. Suzuki NN, Yoshimoto K, Fujioka Y, Ohsumi Y, Inagaki F (2005) The crystal structure of plant ATG12 and its biological implication in autophagy. Autophagy. 1:119–126

123