Abstract Plants, with their unique evolutionary trajectory and complex physiological adaptations, have developed organ-specific mechanisms to cope with various environmental stresses. Autophagy, an essential catabolic process, plays an important role in maintaining plant growth, immunity, and overall fitness. In this study, we reveal the spatio-temporal dynamics of autophagic responses in Arabidopsis thaliana roots and shoots under different stress conditions, including AZD8055 treatment and carbon and nitrogen depletion. Our findings demonstrate that roots exhibit stronger autophagic activity than shoots under all three conditions, highlighting the unique adaptations of these two organs. Furthermore, we dissect the selectivity of autophagy in targeting organelles, revealing immediate, delayed, and no uptake categories. Additionally, we observe organelles coexisting within autophagic bodies, shedding light on the complexity of cargo selection. This study enhances our understanding of plant specific autophagy dynamics, emphasizing its role in sustaining the source-sink functions and offering insights into plant adaptation to diverse stressors.

The ability for stress to modify development is common in plants; yet, how external cues determine phenotypic outputs and developmental responses is not fully understood. Here, we uncovered a ZINC FINGER OF ARABIDOPSIS THALIANA14 (ZAT14) transcription factor whose expression was enhanced in differentiating xylem through its positive regulation by VASCULAR RELATED NAC-DOMAIN PROTEIN7 (VND7), yet, decreased in root tips through its negative regulation by PLETHORA2 (PLT2) in Arabidopsis (Arabidopsis thaliana). Mutating ZAT14 and its closely related homologs, ZAT5, ZAT14L, and ZAT15, disrupted vascular patterning and inhibited xylem differentiation indicating that ZATs are important for xylem formation. A transcriptome analysis of zat triple and quadruple mutants found that many cell wall-related genes were differentially expressed. In particular, 10 expansin genes were repressed by ZATs and several were direct targets of the ZATs. We uncovered that salinity repressed ZAT14, ZAT14L, and ZAT15 vascular expression, whereas zat mutants improved salinity tolerance, decreased xylem differentiation, and reduced cell death mediated by salt. Furthermore, expansin mutants decreased salinity tolerance and increased xylem differentiation under salinity stress. We propose that ZATs are key regulators of programmed cell death that promote xylem formation, yet upon salinity stress, ZATs are repressed to inhibit cell death and improve salt tolerance, thus modifying developmental outputs in response to stress.

Plants possess remarkable regenerative abilities to form de novo vasculature after damage and in response to pathogens that invade and withdraw nutrients.To identify common factors that affect vascular formation upon stress,we searched for Arabidopsis thaliana genes differentially expressed upon Agrobacterium infection,nematode infection,and plant grafting.One such gene is cell wall-related and highly induced by all three stresses,which we named ENHANCED XYLEM AND GRAFTING1(EXG1),since its mutations promote ectopic xylem formation in a vascular cell induction system and enhance graft formation.Further observations revealed that exg1 mutants show inhibited cambium development and callus formation but enhanced tissue attachment,syncytium size,phloem reconnection,and xylem formation.Given that bras-sinosteroids also promote xylem differentiation,we analyzed brassinosteroid-related genes and found that mutations in RLP44 encoding a receptor-like protein cause similar regeneration-related phenotypes as mu-tations in EXG1.Like EXG1,RLP44 expression is also induced by grafting and wounding.Mutations in EXG1 and RLP44 affect the expression of many genes in common,including those related to cell walls and genes important for vascular regeneration.Our results suggest that EXG1 integrates information from wounding or pathogen stress and functions with RLP44 to suppress vascular differentiation during regeneration and healing.

AbstractPlants possess remarkable regenerative abilities to formde novovasculature after damage and in response to pathogens that invade and withdraw nutrients. To look for common factors that affect vascular formation upon stress, we searched forArabidopsis thalianagenes differentially expressed duringAgrobacteriuminfection, nematode infection and plant grafting. One such gene was cell-wall associated and highly induced by all three stresses. Mutations in it enhanced ectopic xylem formation in Vascular cell Induction culture System Using Arabidopsis Leaves (VISUAL) and enhanced graft formation and was thus namedENHANCER OF VISUAL AND GRAFTING 1(EVG1). Mutatedevg1inhibited cambium development and callus formation yet promoted tissue attachment, syncytium size, phloem reconnection and xylem formation.evg1affected abscisic acid and cell wall responses and was itself down regulated by ABA. We found mutations in a receptor-like gene,RLP44, had the same regeneration phenotype asEVG1mutations including enhancing VISUAL and grafting.evg1andrlp44mutants affected the expression of many genes in common including those important for successful regeneration and vascular formation. We propose thatEVG1integrates information from cutting, wounding or parasitism stresses and functions withRLP44to suppress vascular differentiation during regeneration.

Plants possess remarkable regenerative abilities to form de novo vasculature after damage and in response to pathogens that invade and withdraw nutrients. To look for common factors that affect vascular formation upon stress, we searched for Arabidopsis thaliana genes differentially expressed during Agrobacterium infection, nematode infection and plant grafting. One such gene was cell-wall associated and highly induced by all three stresses. Mutations in it enhanced ectopic xylem formation in Vascular cell Induction culture System Using Arabidopsis Leaves (VISUAL) and enhanced graft formation and was thus named ENHANCER OF VISUAL AND GRAFTING 1 ( EVG1 ). Mutated evg1 inhibited cambium development and callus formation yet promoted tissue attachment, syncytium size, phloem reconnection and xylem formation. evg1 affected abscisic acid and cell wall responses and was itself down regulated by ABA. We found mutations in a receptor-like gene, RLP44 , had the same regeneration phenotype as EVG1 mutations including enhancing VISUAL and grafting. evg1 and rlp44 mutants affected the expression of many genes in common including those important for successful regeneration and vascular formation. We propose that EVG1 integrates information from cutting, wounding or parasitism stresses and functions with RLP44 to suppress vascular differentiation during regeneration.### Competing Interest StatementThe authors have declared no competing interest.

Plants show an unparalleled regenerative capacity, allowing them to survive severe stress conditions, such as injury, herbivory attack, and harsh weather conditions. This potential not only replenishes tissues and restores damaged organs but can also give rise to whole plant bodies. Despite the intertwined nature of development and regeneration, common upstream cues and signaling mechanisms are largely unknown. Here, we demonstrate that in addition to being activators of regeneration, ETHYLENE RESPONSE FACTOR 114 (ERF114) and ERF115 govern developmental growth in the absence of wounding or injury. Increased ERF114 and ERF115 activity enhances auxin sensitivity, which is correlated with enhanced xylem maturation and lateral root formation, whereas their knockout results in a decrease in lateral roots. More -over, we provide evidence that mechanical cues contribute to ERF114 and ERF115 expression in correlation with BZR1-mediated brassinosteroid signaling under both regenerative and developmental conditions. Antagonistically, cell wall integrity surveillance via mechanosensory FERONIA signaling suppresses their expression under both conditions. Taken together, our data suggest a molecular framework in which cell wall signals and mechanical strains regulate organ development and regenerative responses via ERF114-and ERF115-mediated auxin signaling.

Plants show an unparalleled regenerative capacity, allowing them to survive severe stress conditions, such as injury, herbivory attack and harsh weather conditions. This potential not only replenishes tissues and restores damaged organs, but can also give rise to whole plant bodies, highlighting the intertwined nature of development and regeneration. It suggests that regeneration and developmental processes respond to the same upstream signals, but how a cell knows which of the two processes to engage is currently unknown. Here, we demonstrate that next to being regulators of regeneration, ETHYENE RESPONSE FACTOR 114 (ERF114) and ERF115 govern developmental growth in the absence of wounding or injury. Increased ERF114 and ERF115 activity is correlated with enhanced xylem maturation and lateral root formation, whereas their knockout results in a decrease in lateral roots and xylem connectivity following grafting. Moreover, we provide evidence that mechanical cues contribute to ERF114 and ERF115 expression in correlation with BZR1 mediated brassinosteroid signaling under both regenerative and developmental conditions. Antagonistically, negative regulation of cell wall extensibility via cell wall-associated mechanosensory FERONIA signaling suppresses their expression under both conditions. Our data suggest a molecular framework in which mechanical perturbations too great to be compensated by adaptive cell wall remodeling results in strong ERF114 and ERF115 expression, switching their role from developmental to regenerative regulators.

Plants display remarkable abilities to adjust growth and development to environmental conditions, such as the amount of available water. This developmental plasticity is apparent not only in root and shoot growth rates, but also in tissue patterning and cell morphology.(1,2) We have previously shown that in response to limited water availability, Arabidopsis thaliana root displays changes in xylem morphology, mediated by the non-cell-autonomous action of abscisic acid, ABA.(2) Here, we show, through analyses of ABA response reporters and tissue-specific suppression of ABA signaling, that xylem cells themselves act as primary signaling centers governing both xylemcell fate and xylem differentiation rate, revealing the cell-autonomous control of multiple aspects of xylem development by ABA. ABA rapidly activates the expression of genes encoding VASCULAR-RELATED NAC DOMAIN (VND) transcription factors. Molecular and genetic analyses revealed that the two ABA-mediated xylem developmental changes are regulated by distinct members of this transcription factor family, with VND2 and VND3 promoting differentiation rate of metaxylem cells, while VND7 promotes the conversion of metaxylem toward protoxylem morphology. This phenomenon shows how different aspects of developmental plasticity can be interlinked, yet genetically separable. Moreover, similarities in phenotypic and molecular responses to ABA in diverse species indicate evolutionary conservation of the ABA-xylem development regulatory network among eudicots. Hence, this study gives molecular insights into how environmental stress modifies plant vascular anatomy and has potential relevance for water use optimization and adaptation to drought conditions.

Summary Plants display a remarkable ability to adjust their growth and development to changes in environmental conditions, such as reduction in water availability. This high degree of plasticity is apparent not only as altered root and shoot growth rates, but also as changes to tissue patterning and cell morphology [1,2]. We have previously shown that Arabidopsis thaliana root xylem displays plastic developmental responses to limited water availability, mediated by non-cell autonomous action of abscisic acid, ABA [2]. Here, we show through analyses of ABA response reporters and tissue specific suppression of ABA signalling that xylem cells act as primary signalling centres for mediation of changes to both xylem cell fate and differentiation rate revealing a cell autonomous control of xylem development by ABA. Transcriptomic changes in response to ABA showed that members of the VASCULAR RELATED NAC DOMAIN (VND) transcription factor family are rapidly activated. Molecular and genetic analyses revealed that the two aspects of xylem developmental changes, cell fate and differentiation rate, are dependent on distinct members of this transcription factor family. Thus, this study provides insights into how different aspects of developmental plasticity can be interlinked, yet genetically independent of each other. Moreover, similarities in phenotypic and molecular responses to ABA in diverse species indicate an evolutionary conservation of the ABA-xylem development regulatory network among eudicots. Hence, this study gives molecular insights on how environmental stress promotes anatomical plasticity to key plant traits with potential relevance for water use optimization and adaptation to drought conditions.

Key message CRISPR-Cas9/Cpf1 system with its unique gene targeting efficiency, could be an important tool for functional study of early developmental genes through the generation of successful knockout plants.Abstract The introduction and utilization of systems biology approaches have identified several genes that are involved in early development of a plant and with such knowledge a robust tool is required for the functional validation of putative candidate genes thus obtained. The development of the CRISPR-Cas9/Cpf1 genome editing system has provided a convenient tool for creating loss of function mutants for genes of interest. The present study utilized CRISPR/Cas9 and CRISPR-Cpf1 technology to knock out an early developmental gene EPFL9 (Epidermal Patterning Factor like-9, a positive regulator of stomatal development in Arabidopsis) orthologue in rice. Germ-line mutants that were generated showed edits that were carried forward into the T2 generation when Cas9-free homozygous mutants were obtained. The homozygous mutant plants showed more than an eightfold reduction in stomatal density on the abaxial leaf surface of the edited rice plants. Potential off-target analysis showed no significant off-target effects. This study also utilized the CRISPR-LbCpf1 (Lachnospiracae bacterium Cpf1) to target the same OsEPFL9 gene to test the activity of this class-2 CRISPR system in rice and found that Cpf1 is also capable of genome editing and edits get transmitted through generations with similar phenotypic changes seen with CRISPR-Cas9. This study demonstrates the application of CRISPR-Cas9/Cpf1 to precisely target genomic locations and develop transgene-free homozygous heritable gene edits and confirms that the loss of function analysis of the candidate genes emerging from different systems biology based approaches, could be performed, and therefore, this system adds value in the validation of gene function studies.

The clustered regularly interspaced short palindromic repeats (CRISPR) system is a prokaryotic adaptive immune system that has the ability to identify specific locations on the bacteriophage (phage) genome to create breaks in it, and internalize the phage genome fragments in its own genome as CRISPR arrays for memory-dependent resistance. Although CRISPR has been used in the dairy industry for a long time, it recently gained importance in the field of genome editing because of its ability to precisely target locations in a genome. This system has further been modified to locate and target any region of a genome of choice due to modifications in the components of the system. By changing the nucleotide sequence of the 20-nucleotide target sequence in the guide RNA, targeting any location is possible. It has found an application in the modification of plant genomes with its ability to generate mutations and insertions, thus helping to create new varieties of plants. With the ability to introduce specific sequences into the plant genome after cleavage by the CRISPR system and subsequent DNA repair through homology-directed repair (HDR), CRISPR ensures that genome editing can be successfully applied in plants, thus generating stronger and more improved traits. Also, the use of the CRISPR editing system can generate plants that are transgene-free and have mutations that are stably inherited, thus helping to circumvent current GMO regulations.

Rice is an important crop for a large portion of the population of the world, being the main source of food and the agriculture of which is the main source of income. Genome editing being the focal point of research in recent times is now rightly being targeted towards improving the quality of rice. Research using the CRISPR genome editing tools has increased the ability to target and modify rice genes for the development of improved varieties. However, current research is focused on improving even further the efficiency of the CRISPR genome editing tools to successfully edit endogenous rice genes. These studies indicate that genome editing is a successful and feasible venture in rice. Newer developments and improvements of CRIPSR tools have further enabled researchers to modify more genes in rice with increased efficiency. The ability of the CRISPR system to generate transgene free genome edited plants is further reason for continued research as it helps to step past genome modification regulatory issues. The successful application and development of CRISPR tools for genome editing in rice will not only help in making site-specific integration events, but will also help in regulating gene expression, gene discovery, rice functional genomics and creating new improved traits in rice.