Chromosome movements during meiosis are essential for accurate homolog recognition and segregation. However, two striking phenomena, rapid prophase chromosome movements (RPMs) and the formation of unusual nuclear structures, both involving large-scale chromatin dynamics and occurring simultaneously during early meiotic prophase I, remain poorly understood in plants. RPMs are known to facilitate homolog pairing across diverse taxa, but their regulation and robustness in plants remain unclear. Similarly, a phenomenon characterized by the formation of nuclear protrusions, that extend through intercellular channels into the cytoplasm of neighboring meiocytes, is frequently reported during plant meiosis. As a rule, this process is referred to as cytomixis, yet its nature and biological significance remain unresolved. Here, we integrate the study of these processes by investigating Arabidopsis thaliana male meiocytes at early prophase I using live-cell imaging with ASY1:GFP as a reporter for chromosome dynamics. We show that chromatin in nuclear protrusions remains active and displays RPMs indistinguishable from those of intact nuclei. Remarkably, a process unavoidably involving mechanical stress and disruption of cytoskeletal connections, does not interrupt RPMs. These findings demonstrate that meiotic RPMs are inherently robust and largely independent of the local cytoplasmic environment and support the view that cytomixis is a normal cytological process rather than a pathology or artifact.

ABSTRACT Autophagy, a cellular degradation pathway, and the phytohormone ethylene function in plant development, senescence, and stress responses. However, the manner of their interaction is mostly unknown. We reasoned that this may be revealed by studying autophagy in a climacteric fruit ripening context, for which ethylene is crucial. During ripening, fruits undergo softening, color change, toxic compound degradation, volatile production, and sugar assembly by fine-tuning synthesis and degradation of their cellular content. For autophagy activity assessment, we analyzed autophagy-related 8 (ATG8) lipidation and GFP-ATG8-labeled autophagosome flux in tomato fruit cells. Autophagy activity increased sharply from ripening initiation, climaxed at its middle stage, and declined towards its end, resembling ethylene production dynamics. Silencing the core-autophagy genes SlATG2 , SlATG7 , and SlATG4 separately in mature fruits resulted in early ethylene production and ripening onset, which was abrogated by 1-methylcyclopropene (1-MCP), an ethylene signaling inhibitor. Beyond ripening, Arabidopsis atg5 and atg7 mutant seedlings exhibited elevated ethylene production and sensitivity to 1-Aminocyclopropane 1-carboxylic acid (ACC), ethylenès precursor, which induces autophagy. This research demonstrates that autophagy limits tomato fruit ripening via a general role in ethylene restriction, opening the path for a mechanistic understanding of autophagy-ethylene crosstalk and harnessing autophagy for fruit shelf-life extension.

BACKGROUND:Plant growth depends on the integration of environmental signals and nutrient availability. Under stress conditions, growth is often attenuated to prioritise defense, creating a trade-off between growth and stress responses. Autophagy, a conserved degradation and recycling mechanism in eukaryotes, plays a central role in maintaining cellular homeostasis during stress. Enhancing autophagy has been shown to improve growth, yield, and stress tolerance in plants, yet the molecular triggers that initiate this process are not fully understood. RESULTS:We identified raffinose, a plant-derived sugar associated with stress responses, as a novel inducer of autophagy in plants. Exogenous application of raffinose stimulated autophagic activity and promoted biomass accumulation and seed yield in an autophagy-dependent manner across multiple plant species. Mechanistic analysis revealed that raffinose activates autophagy through SnRK1 in a TOR-independent manner, and that it upregulates the expression of autophagy-related genes ATG5 and ATG7. The growth-promoting effect of raffinose was specific and not replicated by equivalent carbon supplementation with glucose, while the raffinose precursor galactinol showed similar autophagy-dependent growth enhancement. Finally, we pointed to possible downstream candidates operating autophagy-related biomass accumulation, as identified by metabolic profiling. CONCLUSIONS:Our findings position raffinose as a signalling molecule capable of activating autophagy and enhancing plant growth and yield in a targeted, species-conserved manner. By linking a stress-associated sugar to the activation of a central catabolic pathway, this work reveals a potential mechanism by which plants may optimise the balance between growth and defense. Understanding raffinose-mediated autophagy induction can potentially provide new opportunities for developing strategies to improve crop performance under variable environmental conditions.

Proteolysis, including post-translational proteolytic processing as well as protein degradation and amino acid recycling, is an essential component of the growth and development of living organisms. In this article, experts in plant proteolysis pose and discuss compelling open questions in their areas of research. Topics covered include the role of proteolysis in the cell cycle, DNA damage response, mitochondrial function, the generation of N-terminal signals (degrons) that mark many proteins for degradation (N-terminal acetylation, the Arg/N-degron pathway, and the chloroplast N-degron pathway), developmental and metabolic signaling (photomorphogenesis, abscisic acid and strigolactone signaling, sugar metabolism, and postharvest regulation), plant responses to environmental signals (endoplasmic-reticulum-associated degradation, chloroplast-associated degradation, drought tolerance, and the growth-defense trade-off), and the functional diversification of peptidases. We hope these thought-provoking discussions help to stimulate further research.

Autophagy is a vital process in eukaryotes, maintaining cellular balance by degrading and recycling cellular components. Autophagy is triggered by various nutrient-deprivation conditions and both biotic and abiotic stresses in plants. Autophagy-deficient mutants exhibit early senescence, reduced yield, and hyper-sensitivity to starvation and abiotic stress. Over-expressing autophagy-related genes in various plant species resulted in increased plant size, yield, and stress resistance. Yet, despite the considerable promise of autophagy modulation for improved plant performance, the molecular mechanisms governing its induction remain partially understood. In the current work, we identified raffinose, a sugar linked to plant stress responses, as a novel plant autophagy inducer. Raffinose treatment resulted in increased biomass and yield in an autophagy-dependent manner in several plant species. We also show that raffinose activates autophagy through the SnRK1 kinase complex, independent of TOR signaling Our findings highlight the potential of raffinose as a tool for enhancing crop resilience and productivity. ### Competing Interest Statement The authors have declared no competing interest.

Autophagy, an intracellular process that facilitates the degradation of cytoplasmic materials, plays a dominant role in plant fitness and immunity. While autophagy was shown to be involved in plant response to fungi, bacteria, and viruses, its role in response to insect herbivory is as yet unknown. In this study, we demonstrate a role of autophagy in plant defense against herbivory using Arabidopsis thaliana and the green peach aphid, Myzus persicae . Following six hours of aphid infestation of wildtype plants, we observed high expression of the autophagy-related genes ATG8a and ATG8f , as well as NBR1 ( Next to BRCA1 gene 1 ), a selective autophagy receptor. Moreover, the number of autophagosomes detected by the overexpression of GFP-fused ATG8f in Arabidopsis increased upon aphid infestation. Following this, atg5.1 and atg7.2 mutants were used to study the effect of autophagy on aphid reproduction and feeding behavior. While aphid reproduction on both mutants was lower than on wildtype, feeding behavior was only affected by atg7.2 mutants. Moreover, upon aphid feeding, the Phytoalexin-deficient 4 ( PAD4 ) defense gene was upregulated in wildtype plants but not affected in the mutants. By contrast, the hydrogen peroxide content was much higher in the mutants relative to wildtype, which might have disturbed aphid reproduction and interfered with their feeding. Additionally, an analysis of the phloem sap metabolite profile revealed that atg7.2 mutant plants have lower levels of amino acids and sugars. These findings, together with the high hydrogen peroxide levels, suggest that aphids might exploit the plant autophagy mechanism for their survival.

Fruit taste is determined by sugars, acids and in some species, bitter chemicals. Attraction of seed-dispersing organisms in nature and breeding for consumer preferences requires reduced fruit bitterness. A key metabolic shift during ripening prevents tomato fruit bitterness by eliminating α-tomatine, a renowned defence-associated Solanum alkaloid. Here, we combined fine mapping with information from 150 resequenced genomes and genotyping a 650-tomato core collection to identify nine bitter-tasting accessions including the 'high tomatine' Peruvian landraces reported in the literature. These 'bitter' accessions contain a deletion in GORKY, a nitrate/peptide family transporter mediating α-tomatine subcellular localization during fruit ripening. GORKY exports α-tomatine and its derivatives from the vacuole to the cytosol and this facilitates the conversion of the entire α-tomatine pool to non-bitter forms, rendering the fruit palatable. Hence, GORKY activity was a notable innovation in the process of tomato fruit domestication and breeding.

Reticulophagy, the selective autophagy of endoplasmic reticulum (ER) components, is known to operate in eukaryotes from yeast and unicellular algae to animals and plants. Thus far, only ER-stress induced reticulophagy was reported and analyzed in plants. In this study we characterize a reticulophagy pathway in Arabidopsis thaliana that is triggered by dark-induced starvation but not by ER stress. This pathway is defined by the previously reported ATG8-interacting proteins, ATI1 and ATI2. We further identified the ER-localized MSBP1 (Membrane Steroid Binding Protein 1) as an ATI1- and ATI2-interacting protein and an autophagy cargo, and show that ATI1 and ATI2 serve as its cargo receptors. Together, these findings expand our knowledge on plant responses during energy deprivation and highlight the role of this special type of reticulophagy in this process.Abbreviations: AGO1: ARGONAUTE 1; ATI: ATG8-Interacting Protein; BiFC: Bimolecular Fluorescence Complementation; BR: brassinosteroid; conA: concanamycin A; DMSO: dimethyl sulfoxid; DTT: dithiothreitol; ER: endoplasmic reticulum; GFP: green fluorescent protein; MAPR: Membrane-Associated Progesterone Binding Protein; MSBP: Membrane Steroid Binding Protein; SD: standard deviation; SE: standard error; TM: tunicamycin; TOR: target of rapamycin; Y2H: yeast two-hybrid.

This article comments on: Hickl D, Drews F, Girke C, Zimmer D, Mühlhaus T, Hauth J, Nordström K, Trentmann O, Neuhaus EH, Scheuring D, Fehlmann T, Keller A, Simon M, Möhlmann T. 2021. Differential degradation of RNA species by autophagy-related pathways in Arabidopsis. Journal of Experimental Botany 72, 6867–6881.

ER-phagy, the selective autophagy of endoplasmic reticulum (ER) components, is known to operate in eukaryotes from yeast and unicellular algae to animals and plants. Thus far, only ER-stress derived ER-phagy was reported and analyzed in plants. In this study we characterize an ER-phagy pathway in Arabidopsis thaliana that is triggered by dark-induced starvation and not by ER-stress. This pathway is defined by the previously reported ATG8-interacting proteins, ATI1 and ATI2 and is regulated by the TOR signaling pathway. We further identified ER-localized Membrane Steroid Binding Protein 1 (MSBP1) as an ATI1 and 2 interacting protein and an autophagy cargo, and show that ATI1 and 2 serve as its cargo receptors. Together, these findings expand our knowledge on plants responses during energy deprivation and highlight the role of this special type of ER-phagy in this process.

RNA silencing is a major antiviral defense mechanism in plants and invertebrates. Plant ARGONAUTE1 (AGO1) is pivotal in RNA silencing, and hence is a major target for counteracting viral suppressors of RNA-silencing proteins (VSRs). P0 from Turnip yellows virus (TuYV) is a VSR that was previously shown to trigger AGO1 degradation via an autophagy-like process. However, the identity of host proteins involved and the cellular site at which AGO1 and P0 interact were unknown. Here we report that P0 and AGO1 associate on the endoplasmic reticulum (ER), resulting in their loading into ER-associated vesicles that are mobilized to the vacuole in an ATG5- and ATG7-dependent manner. We further identified ATG8-Interacting proteins 1 and 2 (ATI1 and ATI2) as proteins that associate with P0 and interact with AGO1 on the ER up to the vacuole. Notably, ATI1 and ATI2 belong to an endogenous degradation pathway of ER-associated AGO1 that is significantly induced following P0 expression. Accordingly, ATI1 and ATI2 deficiency causes a significant increase in posttranscriptional gene silencing (PTGS) activity. Collectively, we identify ATI1 and ATI2 as components of an ER-associated AGO1 turnover and proper PTGS maintenance and further show how the VSR P0 manipulates this pathway.

In metazoans, autophagy is an essential component of host defense against viruses, orchestrating their degradation. Such antiviral functions for autophagy have also been long suspected in the green lineage. Two recent reports provide molecular insights on how plants selectively send viral proteins and even particles to the vacuole.

Autophagy is a major cellular degradation pathway in eukaryotes. Recent studies have revealed the importance of autophagy in many aspects of plant life, including seedling establishment, plant development, stress resistance, metabolism, and reproduction. This is manifested by the dual ability of autophagy to execute bulk degradation under severe environmental conditions, while simultaneously to be highly selective in targeting specific compartments and protein complexes to regulate key cellular processes, even during favorable growth conditions. Delivery of cellular components to the vacuole enables their recycling, affecting the plant metabolome, especially under stress. Recent research in Arabidopsis has further unveiled fundamental mechanistic aspects in autophagy which may have relevance in non-plant systems. We review the most recent discoveries concerning autophagy in plants, touching upon all these aspects.

γ-Aminobutyric acid (GABA) is a non-proteinogenic amino acid that is found in uni- and multi-cellular organisms and is involved in many aspects of plant life cycle. GABA metabolism occurs by the action of evolutionary conserved enzymes that constitute the GABA shunt, bypassing two steps of the TCA cycle. The central position of GABA in the interface between plant carbon and nitrogen metabolism is well established. In parallel, there is evidence to support a role for GABA as a signaling molecule in plants. Here we cover some of the recent findings on GABA metabolism and signaling in plants and further suggest that the metabolic and signaling aspects of GABA may actually be inseparable.

Degradation of chloroplasts is a hallmark of both natural and stress-induced plant senescence. Autophagy and senescence-associated vacuoles are two established cellular pathways for chloroplast degradation. Recently, a third independent pathway for chloroplast degradation was reported. Here we will discuss this new discovery in relation to the other known pathways.

Selective autophagy has been extensively studied in various organisms, but knowledge regarding its functions in plants, particularly in organelle turnover, is limited. We have recently discovered ATG8-INTERACTING PROTEIN1 (ATI1) from Arabidopsis thaliana and showed that following carbon starvation it is localized on endoplasmic reticulum (ER)-associated bodies that are subsequently transported to the vacuole. Here, we show that following carbon starvation ATI1 is also located on bodies associating with plastids, which are distinct from the ER ATI bodies and are detected mainly in senescing cells that exhibit plastid degradation. Additionally, these plastid-localized bodies contain a stroma protein marker as cargo and were observed budding and detaching from plastids. ATI1 interacts with plastid-localized proteins and was further shown to be required for the turnover of one of them, as a representative. ATI1 on the plastid bodies also interacts with ATG8f, which apparently leads to the targeting of the plastid bodies to the vacuole by a process that requires functional autophagy. Finally, we show that ATI1 is involved in Arabidopsis salt stress tolerance. Taken together, our results implicate ATI1 in autophagic plastid-to-vacuole trafficking through its ability to interact with both plastid proteins and ATG8 of the core autophagy machinery.

Trafficking of proteins from the endoplasmic reticulum (ER) to the vacuole is a fundamental process in plants, being involved both in vacuole biogenesis as well as with plant growth and response to environmental stresses. Although the canonical transport of cellular components from the ER to the vacuole includes the Golgi apparatus as an intermediate compartment, there are multiple lines of evidence that support the existence of a direct ER-to-vacuole, Golgi-independent, trafficking route in plants that uses the autophagy machinery. Plant autophagy was initially described by electron microscopy, visualizing cellular structures that are morphologically reminiscent of autophagosomes. In some of these reports these structures were shown to transport vacuole residing proteins, particularly seed storage proteins, directly from the ER to the vacuole. More recently, following the discovery of the proteins of the core autophagy machinery, molecular tools were implemented in deciphering the involvement of autophagy in this special trafficking route. Here we review the relatively older and more recent scientific observations, supporting the involvement of autophagy in the special cellular trafficking pathways of plants.

Macroautophagy (hereafter referred to as autophagy) is a cellular mechanism dedicated to the degradation and recycling of unnecessary cytosolic components by their removal to the lytic compartment of the cell (the vacuole in plants). Autophagy is generally induced by stresses causing energy deprivation and its operation occurs by special vesicles, termed autophagosomes. Autophagy also operates in a selective manner, recycling specific components, such as organelles, protein aggregates or even specific proteins, and selective autophagy is implicated in both cellular housekeeping and response to stresses. In plants, selective autophagy has recently been shown to degrade mitochondria, plastids and peroxisomes, or organelle components such as the endoplasmic-reticulum (ER) membrane and chloroplast-derived proteins such as Rubisco. This ability places selective-autophagy as a major factor in cellular steady-state maintenance, both under stress and favorable environmental conditions. Here we review the recent advances documented in plants for this cellular process and further discuss its impact on plant physiology.