Biological membranes play a crucial role in actively hosting, modulating and coordinating a wide range of molecular events essential for cellular function. Membranes are organized into diverse domains giving rise to dynamic molecular patchworks. However, the very definition of membrane domains has been the subject of continuous debate. For example, in the plant field, membrane domains are often referred to as nanodomains, nanoclusters, microdomains, lipid rafts, membrane rafts, signalling platforms, foci or liquid-ordered membranes without any clear rationale. In the context of plant-microbe interactions, microdomains have sometimes been used to refer to the large area at the plant-microbe interface. Some of these terms have partially overlapping meanings at best, but they are often used interchangeably in the literature. This situation generates much confusion and limits conceptual progress. There is thus an urgent need for us as a scientific community to resolve these semantic and conceptual controversies by defining an unambiguous nomenclature of membrane domains. In this Review, experts in the field get together to provide explicit definitions of plasma membrane domains in plant systems and experimental guidelines for their study. We propose that plasma membrane domains should not be considered on the basis of their size alone but rather according to the biological system being considered, such as the local membrane environment or the entire cell.

Plant legumains are Asn/Asp-specific endopeptidases that have diverse functions in plants. Peptide asparaginyl ligases (PALs) are a special legumain subtype that primarily catalyze peptide bond formation rather than hydrolysis. PALs are versatile protein engineering tools but are rarely found in nature. To overcome this limitation, here we describe a two-step method to design and engineer a high-yield and efficient recombinant PAL based on commonly found asparaginyl endopeptidases. We first constructed a consensus sequence derived from 1500 plant legumains to design the evolutionarily stable legumain conLEG that could be produced in E. coli with 20-fold higher yield relative to that for natural legumains. We then applied the ligase-activity determinant hypothesis to exploit conserved residues in PAL substrate-binding pockets and convert conLEG into conPAL1-3. Functional studies showed that conLEG is primarily a hydrolase, whereas conPALs are ligases. Importantly, conPAL3 is a superefficient and broadly active PAL for protein cyclization and ligation.

CRISPR (clustered regularly interspaced short palindromic repeats)/Cas (CRISPR-associated) technology is a versatile genome editing tool that has been used to improve agriculturally important plant traits. Due to its precision, CRISPR/Cas9 is more effective than either conventional plant breeding methods or standard genetic engineering approaches for the rapid development of new varieties resilient to climate change. In addition to knowledge in tissue culture-based plant transformation, effective gene-specific single guide RNA (sgRNA) design, prediction of its off-target effect and utilization of vectors, promoters, Cas proteins and terminators is required for CRISPR/Cas9. Various bioinformatics tools are available for the best sgRNA design and screening of the off-targets. Various tools are used in the delivery of CRISPR/Cas components into cells and the genome. Moreover, some recent studies proved the simultaneous silencing of different paralogs in the same family or several genes working in the same pathway by using multiple-target sgRNA designs. This review summarizes the type of promoters, Cas proteins, recognition sequences, and terminators available for the development of knock-out and overexpression plant lines. It also provides a general guideline for the development of genome-edited plants from the design of sgRNAs to the selection of non-transgenic genome-edited T2 generation.

Assessing species\' extinction risk is vital to setting conservation priorities. However, assessment endeavors, such as those used to produce the IUCN Red List of Threatened Species, have significant gaps in taxonomic coverage. Automated assessment (AA) methods are gaining popularity to fill these gaps. Choices made in developing, using, and reporting results of AA methods could hinder their successful adoption or lead to poor allocation of conservation resources. We explored how choice of data cleaning type and level, taxonomic group, training sample, and automation method affect performance of threat status predictions for plant species. We used occurrences from the Global Biodiversity Information Facility (GBIF) to generate assessments for species in 3 taxonomic groups based on 6 different occurrence-based AA methods. We measured each method\'s performance and coverage following increasingly stringent occurrence cleaning. Automatically cleaned data from GBIF performed comparably to occurrence records cleaned manually by experts. However, all types of data cleaning limited the coverage of AAs. Overall, machine-learning-based methods performed well across taxa, even with minimal data cleaning. Results suggest a machine-learning-based method applied to minimally cleaned data offers the best compromise between performance and species coverage. However, optimal data cleaning, training sample, and automation methods depend on the study group, intended applications, and expertise.
La valoración del riesgo de extinción de las especies es vital para el establecimiento de prioridades de conservación. Sin embargo, los esfuerzos de valoración, como los que se usan para generar la Lista Roja de Especies Amenazadas de la UICN, tienen brechas importantes en la cobertura taxonómica. Los métodos de valoración automatizada (VA) están ganando popularidad como reductores de estas brechas. Las elecciones realizadas en el desarrollo, uso y reporte de resultados de los métodos de VA podrían obstaculizar su adopción exitosa o derivar en una asignación deficiente de recursos para la conservación. Exploramos cómo la selección del tipo de limpieza de datos y el nivel, grupo taxonómico, muestra de entrenamiento y el método de automatización afectan el desempeño de las predicciones del estado de amenaza de las especies de plantas. Usamos los registros de la Global Biodiversity Information Facility (GBIF) para generar las valoraciones de las especies de tres grupos taxonómicos con base en seis métodos diferentes de VA basados en la presencia de las especies. Medimos el desempeño de cada método y cobertura después de una limpieza de presencia cada vez más estricta. La información de la GBIF limpiada automáticamente tuvo un desempeño comparable con los registros de presencia limpiados manualmente por expertos. Sin embargo, todos los tipos de limpieza de datos limitaron la cobertura de las valoraciones automatizadas. En general, los métodos basados en el aprendizaje automático tuvieron un buen desempeño en todos los taxones, incluso con una limpieza mínima de datos. Los resultados sugieren que un método basado en el aprendizaje automático aplicado a información con la mínima limpieza ofrece el mejor equilibrio entre el desempeño y la cobertura de la especie. A pesar de esto, la limpieza óptima de datos, la muestra de entrenamiento y los métodos de automatización dependen del grupo de estudio, las aplicaciones deseadas y la experiencia.

Approaches, values, and perceptions in invasion science are highly dynamic, and like in other disciplines, views among different people can diverge. This has led to debate in the field specifically surrounding the core themes of values, management, impacts, and terminology. Considering these debates, we surveyed 698 scientists and practitioners globally to assess levels of polarization (opposing views) on core and contentious topics. The survey was distributed online (via Google Forms) and promoted through listservs and social media. Although there were generally high levels of consensus among respondents, there was some polarization (scores of ≥0.39 [top quartile]). Relating to values, there was high polarization regarding claims of invasive species denialism, whether invasive species contribute to biodiversity, and how biodiversity reporting should be conducted. With regard to management, there were polarized views on banning the commercial use of beneficial invasive species, the extent to which stakeholders\' perceptions should influence management, whether invasive species use alone is an appropriate control strategy, and whether eradication of invasive plants is possible. For impacts, there was high polarization concerning whether invasive species drive or are a side effect of degradation and whether invasive species benefits are understated. For terminology, polarized views related to defining invasive species based only on spread, whether species can be labeled as invasive in their native ranges, and whether language used is too xenophobic. Factor and regression analysis revealed that views were particularly divergent between people working on different invasive taxa (plants and mammals) and in different disciplines (between biologists and social scientists), between academics and practitioners, and between world regions (between Africa and the Global North). Unlike in other studies, age and gender had a limited influence on response patterns. Better integration globally and between disciplines, taxa, and sectors (e.g., academic vs. practitioners) could help build broader understanding and consensus.
Los enfoques, valores y percepciones en el campo de las invasiones biológicas son muy dinámicos, y como en otras disciplinas científicas, los expertos pueden tener distintas opiniones. Esto ha creado debates, especialmente sobre temas relacionados con valores, gestión, impactos y terminología. Considerando estos debates, encuestamos a 698 científicos y gestores de todo el mundo para evaluar sus niveles de polarización (opiniones opuestas) sobre una serie de temas fundamentales y polémicos. La encuesta fue distribuida a través de internet (a través de Google Forms) y promovida por medio de listas de correo electrónico y redes sociales. Aunque, en general, hubo consenso entre los encuestados, hubo cierta polarización (puntuaciones de ≥ 0.39 [cuartil más alto]). En relación con valores, hubo una gran polarización sobre aquellas declaraciones relacionadas con el negacionismo de especies invasoras, si las especies invasoras contribuyen a aumentar la biodiversidad y cómo se deberían llevar a cabo los informes sobre biodiversidad. En relación con la gestión, hubo opiniones polarizadas sobre la prohibición del uso comercial de especies invasoras beneficiosas, si la opinión de las partes interesadas debería influir en la gestión, si el uso de especies invasoras por sí solo es una estrategia de control adecuada y si la erradicación de plantas invasoras es factible. En cuanto a impactos, hubo gran polarización en cuanto a sí las especies invasoras conducen a o son un efecto lateral de la degradación de ecosistemas y ssi los beneficios de las especies invasoras están subestimados. En cuanto a terminología, encontramos opiniones polarizadas relacionadas con definir especies invasoras exclusivamente en base a su expansión, si las especies se pueden considerar invasoras en sus rangos de distribución nativos y si el lenguaje utilizado en el campo de las invasiones biológicas es xenofóbico. Los análisis factoriales y de regresión revelaron que las opiniones de los expertos encuestados fueron particularmente divergentes entre personas que trabajan con diferentes taxones (plantas y mamíferos) en diferentes disciplinas (entre biólogos y sociólogos), entre científicos y gestores y entre regiones del mundo (entre países de África y del hemisferio Norte). A diferencia de otros estudios, la edad y el género tuvieron una influencia limitada sobre lass respuestas obtenidas. Una mejor integración global y entre disciplinas, taxones y sectores (o. e., investigadores vs. gestores) podría contribuir a alcanzar un mayor entendimiento y consenso.

The genus \'Candidatus Phytoplasma\' was proposed to accommodate cell wall-less bacteria that are molecularly and biochemically incompletely characterized, and colonize plant phloem and insect vector tissues. This provisional classification is highly relevant due to its application in epidemiological and ecological studies, mainly aimed at keeping the severe phytoplasma plant diseases under control worldwide. Given the increasing discovery of molecular diversity within the genus \'Ca. Phytoplasma\', the proposed guidelines were revised and clarified to accommodate those \'Ca. Phytoplasma\' species strains sharing >98.65 % sequence identity of their full or nearly full 16S rRNA gene sequences, obtained with at least twofold coverage of the sequence, compared with those of the reference strain of such species. Strains sharing <98.65 % sequence identity with the reference strain but >98.65 % with other strain(s) within the same \'Ca. Phytoplasma\' species should be considered related strains to that \'Ca. Phytoplasma\' species. The guidelines herein, keep the original published reference strains. However, to improve \'Ca. Phytoplasma\' species assignment, complementary strains are suggested as an alternative to the reference strains. This will be implemented when only a partial 16S rRNA gene and/or a few other genes have been sequenced, or the strain is no longer available for further molecular characterization. Lists of \'Ca. Phytoplasma\' species and alternative reference strains described are reported. For new \'Ca. Phytoplasma\' species that will be assigned with identity ≥98.65 % of their 16S rRNA gene sequences, a threshold of 95 % genome-wide average nucleotide identity is suggested. When the whole genome sequences are unavailable, two among conserved housekeeping genes could be used. There are 49 officially published \'Candidatus Phytoplasma\' species, including \'Ca. P. cocostanzaniae\' and \'Ca. P. palmae\' described in this manuscript.

In the era of big and omics data, good organization, management, and description of experimental data are crucial for achieving high-quality datasets. This, in turn, is essential for the export of robust results, to publish reliable papers, make data more easily available, and unlock the huge potential of data reuse. Lately, more and more journals now require authors to share data and metadata according to the FAIR (Findable, Accessible, Interoperable, Reusable) principles. This work aims to provide a step-by-step guideline for the FAIR data and metadata management specific to grapevine and wine science. In detail, the guidelines include recommendations for the organization of data and metadata regarding (i) meaningful information on experimental design and phenotyping, (ii) sample collection, (iii) sample preparation, (iv) chemotype analysis, (v) data analysis (vi) metabolite annotation, and (vii) basic ontologies. We hope that these guidelines will be helpful for the grapevine and wine metabolomics community and that it will benefit from the true potential of data usage in creating new knowledge being revealed.

This review outlines the advances achieved in monitoring natural and anthropogenic plant stressors by hyperspectral remote sensing over the last 50 years. A broad diversity of methods based on field and imaging spectroscopy were developed in that field for precision farming and environmental monitoring purposes. From the 466 articles reviewed, we identified the main factors to consider to achieve accurate monitoring of plant stress, namely: The plant species and the stressor to monitor, the goal (detection or quantification), and scale (field or broad-scale) of monitoring, and the need for controlled experiments. Based on these factors, we then provide recommendations and guidelines for the development of reliable methods to monitor 11 major biotic and abiotic plant stressors. For each stressor, the effects on plant health and reflectance are described and the most suited spectral regions, scale, spatial resolution, and processing approaches to achieve accurate monitoring are presented. As a perspective, we discuss two major components that should be implemented in future methods to improve stress monitoring: The discrimination of plant stressors with similar effects on plants and the transferability of the methods across scales.

Clustered regularly interspaced short palindromic repeat (CRISPR)-associated systems have revolutionized genome engineering by facilitating a wide range of targeted DNA perturbations. These systems have resulted in the development of powerful new screens to test gene functions at the genomic scale. While there is tremendous potential to map and interrogate gene regulatory networks at unprecedented speed and scale using CRISPR screens, their implementation in plants remains in its infancy. Here we discuss the general concepts, tools, and workflows for establishing CRISPR screens in plants and analyze the handful of recent reports describing the use of this strategy to generate mutant knockout collections or to diversify DNA sequences. In addition, we provide insight into how to design CRISPR knockout screens in plants given the current challenges and limitations and examine multiple design options. Finally, we discuss the unique multiplexing capabilities of CRISPR screens to investigate redundant gene functions in highly duplicated plant genomes. Combinatorial mutant screens have the potential to routinely generate higher-order mutant collections and facilitate the characterization of gene networks. By integrating this approach with the numerous genomic profiles that have been generated over the past two decades, the implementation of CRISPR screens offers new opportunities to analyze plant genomes at deeper resolution and will lead to great advances in functional and synthetic biology.

This chapter presents an overview of the major plant DNA sequences and molecular methods available for plant taxonomy. Guidelines are provided for the choice of sequences and methods to be used, based on the DNA compartment (nuclear, chloroplastic, mitochondrial), evolutionary mechanisms, and the level of taxonomic differentiation of the plants under survey.