Mutations in the DDHD2 (DDHD domain containing 2) gene cause autosomal recessive spastic paraplegia type 54 (SPG54), a rare neurodegenerative disorder characterized by the early childhood onset of progressive spastic paraplegia. DDHD2 is reported as the principal brain triacylglycerol (TAG) lipase whose dysfunction causes massive lipid droplet (LD) accumulation in the brains of SPG54 patients. However, the precise functions of DDHD2 in regulating LD catabolism are not yet fully understood. In a recent study, we demonstrate that DDHD2 interacts with multiple members of the Atg8-family proteins (MAP1LC3/LC3s, GABARAPs), which play crucial roles in lipophagy. DDHD2 possesses two LC3-interacting region (LIR) motifs that contribute to its LD-eliminating activity. Moreover, DDHD2 enhances the colocalization between LC3B and LDs to promote lipophagy. LD·ATTEC, a compound that tethers LC3 to LDs to enhance their macroautophagic/autophagic clearance, effectively counteracts DDHD2 deficiency-induced LD accumulation. These findings provide insights into the dual functions of DDHD2 as a TAG lipase and cargo receptor for lipophagy in neuronal LD catabolism, and also suggest a potential therapeutic approach for treating SPG54 patients.

Autophagy, a conserved cellular recycling process, plays a crucial role in maintaining homeostasis under stress conditions. It also regulates the development and virulence of numerous filamentous fungi. In this study, we investigated the specific function of ATG8, a reliable autophagic marker, in the opportunistic pathogen Aspergillus flavus. To investigate the role of atg8 in A. flavus, the deletion and complemented mutants of atg8 were generated according to the homologous recombination principle. Deletion of atg8 showed a significant decrease in conidiation, spore germination, and sclerotia formation compared to the WT and atg8C strains. Additionally, aflatoxin production was found severely impaired in the ∆atg8 mutant. The stress assays demonstrated that ATG8 was important for A. flavus response to oxidative stress. The fluorescence microscopy showed increased levels of reactive oxygen species in the ∆atg8 mutant cells, and the transcriptional result also indicated that genes related to the antioxidant system were significantly reduced in the ∆atg8 mutant. We further found that ATG8 participated in regulating the pathogenicity of A. flavus on crop seeds. These results revealed the biological role of ATG8 in A. flavus, which might provide a potential target for the control of A. flavus and AFB1 biosynthesis.

Atg8 family proteins play crucial roles in autophagy to maintain cellular homeostasis. However, the physiological roles of Atg8 family proteins have not been systematically determined. In this study, we generated Atg8a and Atg8b (homologs of Atg8 in Drosophila melanogaster) knockout flies. We found that the loss of Atg8a affected autophagy and resulted in partial lethality, abnormal wings, decreased lifespan, and decreased climbing ability in flies. Furthermore, the loss of Atg8a resulted in reduced muscle integrity and the progressive degeneration of the neuron system. We also found that the phosphorylation at Ser88 of Atg8a is important for autophagy and neuronal integrity. The loss of Atg8b did not affect autophagy but induced male sterility in flies. Here, we take full advantage of the fly system to elucidate the physiological function of Atg8a and Atg8b in Drosophila.

Mitophagy is a selective autophagy for the degradation of damaged or excessive mitochondria to maintain intracellular homeostasis. In Magnaporthe oryzae, a filamentous ascomycetous fungus that causes rice blast, the most devastating disease of rice, mitophagy occurs in the invasive hyphae to promote infection. To date, only a few proteins are known to participate in mitophagy and the mechanisms of mitophagy are largely unknown in pathogenic fungi. Here, by a yeast two-hybrid screen with the core autophagy-related protein MoAtg8 as a bait, we obtained a MoAtg8 interactor MoAti1 (MoAtg8-interacting protein 1). Fluorescent observations and protease digestion analyses revealed that MoAti1 is primarily localized to the peripheral mitochondrial outer membrane and is responsible for recruiting MoAtg8 to mitochondria under mitophagy induction conditions. MoAti1 is specifically required for mitophagy, but not for macroautophagy and pexophagy. Infection assays suggested that MoAti1 is required for mitophagy in invasive hyphae during pathogenesis. Notably, no homologues of MoAti1 were found in rice and human protein databases, indicating that MoAti1 may be used as a potential target to control rice blast. By the host-induced gene silencing (HIGS) strategy, transgenic rice plants targeted to silencing MoATI1 showed enhanced resistance against M. oryzae with unchanged agronomic traits. Our results suggest that MoATI1 is required for mitophagy and pathogenicity in M. oryzae and can be used as a target for reducing rice blast.

Banana is one of the most important fruits in the world due to its status as a major food source for more than 400 million people. Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4) causes substantial losses of banana crops every year, and molecular host resistance mechanisms are currently unknown. We here performed a genomewide analysis of the autophagy-related protein 8 (ATG8) family in a wild banana species. The banana genome was found to contain 10 MaATG8 genes. Four MaATG8s formed a gene cluster in the distal part of chromosome 4. Phylogenetic analysis of ATG8 families in banana, Arabidopsis thaliana, citrus, rice, and ginger revealed five major phylogenetic clades shared by all of these plant species, demonstrating evolutionary conservation of the MaATG8 families. The transcriptomic analysis of plants infected with Foc TR4 showed that nine of the MaATG8 genes were more highly induced in resistant cultivars than in susceptible cultivars. Finally, MaATG8F was found to interact with MaATG4B in vitro (with yeast two-hybrid assays), and MaATG8F and MaATG4B all positively regulated banana resistance to Foc TR4. Our study provides novel insights into the structure, distribution, evolution, and expression of the MaATG8 family in bananas. Furthermore, the discovery of interactions between MaATG8F and MaATG4B could facilitate future research of disease resistance genes for the genetic improvement of bananas.

Autophagy is a eukaryote-specific cellular process that can engulf unwanted targets with double-membrane autophagosomes and subject them to the vacuole or lysosome for breaking down and recycling, playing dual roles in plant growth and environmental adaptions. However, perception of specific environmental signals for autophagy induction is largely unknown, limiting its application in agricultural usage. Identification of plant-unique DUF641 family COST1 (Constitutively Stressed 1) protein directly links drought perception and autophagy induction, shedding light on manipulating autophagy for breeding stress tolerant crops. In this study, we performed a genome-wide analysis of DUF641/COST family in tomato, and identified five SlCOST genes SlCOST1, -2, -3, -4, and -5. SlCOST genes show both overlapping and distinct expression patterns in plant growth and stress responding. In addition, SlCOST1, -3, -4, -5 proteins demonstrate co-localization with autophagy adaptor protein ATG8e, and all five SlCOST proteins show interactions ATG8e in planta. However, only SlCOST1, the closest ortholog of Arabidopsis AtCOST1, can restore cost1 mutant to WT level, suggesting conserved role of COST1 and functional diversification of SlCOST family in tomato. Our study provides clues for future investigation of autophagy-related COST family and its promising implementations in breeding crops with robust environmental plasticity.

The Atg8 family of ubiquitin-like proteins play pivotal roles in autophagy and other processes involving vesicle fusion and transport where the lysosome/vacuole is the end station. Nuclear roles of Atg8 proteins are also emerging. Here, we review the structural and functional features of Atg8 family proteins and their protein-protein interaction modes in model organisms such as yeast, Arabidopsis, C. elegans and Drosophila to humans. Although varying in number of homologs, from one in yeast to seven in humans, and more than ten in some plants, there is a strong evolutionary conservation of structural features and interaction modes. The most prominent interaction mode is between the LC3 interacting region (LIR), also called Atg8 interacting motif (AIM), binding to the LIR docking site (LDS) in Atg8 homologs. There are variants of these motifs like \"half-LIRs\" and helical LIRs. We discuss details of the binding modes and how selectivity is achieved as well as the role of multivalent LIR-LDS interactions in selective autophagy. A number of LIR-LDS interactions are known to be regulated by phosphorylation. New methods to predict LIR motifs in proteins have emerged that will aid in discovery and analyses. There are also other interaction surfaces than the LDS becoming known where we presently lack detailed structural information, like the N-terminal arm region and the UIM-docking site (UDS). More interaction modes are likely to be discovered in future studies.

Syntrichia caninervis is the dominant species of biological soil crust in the desert, including the Gurbantunggut Desert in China. It is widely distributed in drylands and considered to be a new model of vegetative desiccation tolerance moss. Here, we cloned an ATG8 gene from S. caninervis and confirmed its function under multiple abiotic stresses, both in situ and in Physcomitrium patens. The results showed that the ScATG8 gene encoded a protein with a highly conserved ATG8 functional domain. ScATG8 gene was increasingly expressed under different abiotic stresses. Under desiccation stress, the overexpression of ScATG8 enhanced the tolerance of S. caninervis and its ability to scavenge ROS. In addition, ScATG8 overexpression promoted the growth of P. patens under multiple stress conditions. Thus, ScATG8 may be a multifunctional gene, and it plays a critical role in the survival of S. caninervis under various abiotic stresses. Our results provide new insights into the function of ATG8 in enabling desiccation tolerance and open up more possibilities for subsequent plant molecular breeding and the mining of the resistance genes of S. caninervis and other moss species.

ATG8/LC3-mediated autophagosome formation is a key rate-limiting step in the process of autophagy. The parasitic protist Toxoplasma gondii possesses a single ATG8 homolog (TgATG8), which can localize to either cytosolic autophagosome involved in delivery of autophagic material in bradyzoites, or the outermost membrane of apicoplast, a nonphotosynthetic plastid-like organelle, responsible for maintaining homeostasis in tachyzoites. However, mechanisms that regulate TgATG8 remain insufficiently understood. Here, a TgATG7 conditional knockdown line that we have generated is severely impaired in parasite\'s growth and exhibits significant defects in the organelle level, strikingly with a fragmentation of the mitochondrial network and a loss of the apicoplast. Specific TgATG7C1133S point mutant complemented line showed that these defects were associated with its E1-type enzyme activity. Both depletion of TgATG7 and mutation of its catalytic cysteine 1133 hindered TgATG8 lipidation and apicoplast localization. Unexpectedly, we also found that depletion of TgATG7 reduced the unlipidated TgATG8 protein level. Subsequently, we determined that TgATG7 was able to interact with TgATG8 directly via its C-terminal domain and multi-monoubiquitination stimulated proteasome-dependent degradation of TgATG8, while TgATG7 could inhibit the degradation through stabilization of TgATG8. Additionally, we identified a putative TgATG8 interacting fragment of TgATG7, 1281-1290aa. Depletion of the fragment impaired the parasite growth and apicoplast inheritance. To our knowledge, our study is the first to elucidate the role of TgATG7 and the ubiquitin-proteasome system in synergistically regulating the non-lipidated pool of TgATG8, suggesting a potential homeostatic mechanism responsible for balancing autophagic activity in T. gondii.

High temperature stress poses significant adverse effects on crop yield and quality. Yet the molecular mechanisms underlying heat stress tolerance in plants/crops, especially regarding the organellar remodeling and homeostasis, are largely unknown. In a recent study, Zhou et al. reported that autophagy-related 8 (ATG8), a famous regulator involved in autophagy, plays a new role in Golgi restoration upon heat stress. Golgi apparatus is vacuolated following short-term acute heat stress, and ATG8 is translocated to the dilated Golgi membrane and interacts with CLATHRIN LIGHT CHAIN 2 (CLC2) to facilitate Golgi restoration, which is dependent on the ATG conjugation system, but not of the upstream autophagic initiators. These exciting findings broaden the fundamental role of ATG8, and elucidate the organelle-level restoration mechanism of Golgi upon heat stress in plants.