Cellular and gene expression patterns associated with root bifurcation in Selaginella.

The roots of lycophytes branch through dichotomy or bifurcation, during which the root apex splits into two daughter roots. This is morphologically distinct from lateral root (LR) branching in the extant euphyllophytes, with LRs developing along the root axis at different distances from the apex. Although the process of root bifurcation is poorly understood, such knowledge can be important, because it may represent an evolutionarily ancient strategy that roots recruited to form new stem cells or meristems. In this study, we examined root bifurcation in the lycophyte Selaginella moellendorffii. We characterized an in vitro developmental time frame based on repetitive apex bifurcations, allowing us to sample different stages of dichotomous root branching and analyze the root meristem and root branching in S. moellendorffii at the microscopic and transcriptomic level. Our results showed that, in contrast to previous assumptions, initial cells (ICs) in the root meristem are mostly not tetrahedral but rather show an irregular shape. Tracking down the early stages of root branching argues for the occurrence of a symmetric division of the single IC, resulting in two apical stem cells that initiate root meristem bifurcation. Moreover, we generated a S. moellendorffii root branching transcriptome that resulted in the delineation of a subset of core meristem genes. The occurrence of multiple putative orthologs of meristem genes in this dataset suggests the presence of conserved pathways in the control of meristem and root stem cell establishment or maintenance.

Determinants of Host Range Specificity in Legume-Rhizobia Symbiosis.

Leguminous plants possess the almost unique ability to enter symbiosis with soil-resident, nitrogen fixing bacteria called rhizobia. During this symbiosis, the bacteria physically colonize specialized organs on the roots of the host plant called nodules, where they reduce atmospheric nitrogen into forms that can be assimilated by the host plant and receive photosynthates in return. In order for nodule development to occur, there is extensive chemical cross-talk between both parties during the formative stages of the symbiosis. The vast majority of the legume family are capable of forming root nodules and typically rhizobia are only able to fix nitrogen within the context of this symbiotic association. However, many legume species only enter productive symbiosis with a few, or even single rhizobial species or strains, and vice-versa. Permitting symbiosis with only rhizobial strains that will be able to fix nitrogen with high efficiency is a crucial strategy for the host plant to prevent cheating by rhizobia. This selectivity is enforced at all stages of the symbiosis, with partner choice beginning during the initial communication between the plant and rhizobia. However, it can also be influenced even once nitrogen-fixing nodules have developed on the root. This review sets out current knowledge about the molecular mechanisms employed by both parties to influence host range during legume-rhizobia symbiosis.

Comprehensive Chemical Characterization of the Aerosol Emissions of a Vaping Product Based on a New Technology.

Around 10 million people in the United States and 3 million people in the United Kingdom are estimated to use vaping category products. There are some estimates that there will be 75-80 million vapers worldwide by 2020. Most of these products are based on coil-and-wick technology. Because the heating and aerosol formation are separate processes, the system can lead to dry-wicking and elevated emission of carbonyls if designed and/or manufactured poorly. Low-nicotine and low-power coil-and-wick devices have also been linked to increased exposure to formaldehyde due to compensatory behavior by users. We characterized the emissions of a vaping product which uses a fabric-free stainless-steel mesh distiller plate technology that heats and aerosolizes the e-liquid in a single process. The plate has a microporous structure for capillary-induced liquid transformation (wicking) and aerosolization that is optimized to avoid fluid starvation and overheating and improved control. Compared with emissions previously reported for a coil-and-wick nicotine vaping product (e-cigarette), most classes of harmful and potentially harmful constituents (HPHCs) from this vaping product were below the level of detection or quantification. For those that were quantifiable, this vaping product generally had lower levels of emissions than the e-cigarette, including carbonyls. Formaldehyde and methyl glyoxal levels did not differ significantly between vaping products. In this system, the single mode of liquid transfer and vapor formation permits high aerosol mass delivery but further reduces emissions of HPHCs that may be present in conventional e-cigarette aerosol, by lessening the risk of thermal breakdown of the aerosol-generating solvent mixture.

3DCellAtlas Meristem: a tool for the global cellular annotation of shoot apical meristems.

Modern imaging approaches enable the acquisition of 3D and 4D datasets capturing plant organ development at cellular resolution. Computational analyses of these data enable the digitization and analysis of individual cells. In order to fully harness the information encoded within these datasets, annotation of the cell types within organs may be performed. This enables data points to be placed within the context of their position and identity, and for equivalent cell types to be compared between samples. The shoot apical meristem (SAM) in plants is the apical stem cell niche from which all above ground organs are derived. We developed 3DCellAtlas Meristem which enables the complete cellular annotation of all cells within the SAM with up to 96% accuracy across all cell types in Arabidopsis and 99% accuracy in tomato SAMs. Successive layers of cells are identified along with the central stem cells, boundary regions, and layers within developing primordia. Geometric analyses provide insight into the morphogenetic process that occurs during these developmental processes. Coupling these digital analyses with reporter expression will enable multidimensional analyses to be performed at single cell resolution. This provides a rapid and robust means to perform comprehensive cellular annotation of plant SAMs and digital single cell analyses, including cell geometry and gene expression. This fills a key gap in our ability to analyse and understand complex multicellular biology in the apical plant stem cell niche and paves the way for digital cellular atlases and analyses.

Histological Profiling Over Time to Optimize Root Cell Type-Specific Reporter Lines for Cell Sorting.

Cell type-specific marker lines expressing fluorophores such as GFP or GUS can be used as starting material from which single cell types can be isolated by fluorescence-activated cell sorting (FACS) and/or for the study of root development. Establishing the stability of these lines is an essential step prior to further study to ensure that marker expression and localization is stable over time and during environmental perturbations of interest to researchers applying these lines as treatments. Here, we detail the use of root cross sectioning to investigate marker expression throughout the length and width of the root using the model legume Medicago truncatula as an example. In order to deal with the fact that plant cell walls are highly autofluorescent, we also describe the usage of confocal microscopy to conduct a lambda scan to discriminate autofluorescence from marker molecule expression.

Changes in Gene Expression in Space and Time Orchestrate Environmentally Mediated Shaping of Root Architecture.

Shaping of root architecture is a quintessential developmental response that involves the concerted action of many different cell types, is highly dynamic, and underpins root plasticity. To determine to what extent the environmental regulation of lateral root development is a product of cell-type preferential activities, we tracked transcriptomic responses to two different treatments that both change root development in Arabidopsis thaliana at an unprecedented level of temporal detail. We found that individual transcripts are expressed with a very high degree of temporal and spatial specificity, yet biological processes are commonly regulated, in a mechanism we term response nonredundancy. Using causative gene network inference to compare the genes regulated in different cell types and during responses to nitrogen and a biotic interaction, we found that common transcriptional modules often regulate the same gene families but control different individual members of these families, specific to response and cell type. This reinforces that the activity of a gene cannot be defined simply as molecular function; rather, it is a consequence of spatial location, expression timing, and environmental responsiveness.