The mRNA mobileome: challenges and opportunities for deciphering signals from the noise.

Organismal communication entails encoding a message that is sent over space or time to a recipient cell, where that message is decoded to activate a downstream response. Defining what qualifies as a functional signal is essential for understanding intercellular communication. In this review, we delve into what is known and unknown in the field of long-distance messenger RNA (mRNA) movement and draw inspiration from the field of information theory to provide a perspective on what defines a functional signaling molecule. Although numerous studies support the long-distance movement of hundreds to thousands of mRNAs through the plant vascular system, only a small handful of these transcripts have been associated with signaling functions. Deciphering whether mobile mRNAs generally serve a role in plant communication has been challenging, due to our current lack of understanding regarding the factors that influence mRNA mobility. Further insight into unsolved questions regarding the nature of mobile mRNAs could provide an understanding of the signaling potential of these macromolecules.

Introduction of barnase/barstar in soybean produces a rescuable male sterility system for hybrid breeding.

Hybrid breeding for increased vigour has been used for over a century to boost agricultural outputs without requiring higher inputs. While this approach has led to some of the most substantial gains in crop productivity, breeding barriers have fundamentally limited soybean (Glycine max) from reaping the benefits of hybrid vigour. Soybean flowers self-pollinate prior to opening and thus are not readily amenable to outcrossing. In this study, we demonstrate that the barnase/barstar male sterility/rescue system can be used in soybean to produce hybrid seeds. By expressing the cytotoxic ribonuclease, barnase, under a tapetum-specific promoter in soybean anthers, we are able to completely block pollen maturation, creating male sterile plants. We show that fertility can be rescued in the F1 generation of these barnase-expressing lines when they are crossed with pollen from plants that express the barnase inhibitor, barstar. Importantly, we found that the successful rescue of male fertility is dependent on the relative dosage of barnase and barstar. When barnase and barstar were expressed under the same tapetum-specific promoter, the F1 offspring remained male sterile. When we expressed barstar under a relatively stronger promoter than barnase, we were able to achieve a successful rescue of male fertility in the F1 generation. This work demonstrates the successful implementation of a biotechnology approach to produce fertile hybrid offspring in soybean.

The evolutionary history of small RNAs in Solanaceae.

The Solanaceae or "nightshade" family is an economically important group with remarkable diversity. To gain a better understanding of how the unique biology of the Solanaceae relates to the family's small RNA (sRNA) genomic landscape, we downloaded over 255 publicly available sRNA data sets that comprise over 2.6 billion reads of sequence data. We applied a suite of computational tools to predict and annotate two major sRNA classes: (1) microRNAs (miRNAs), typically 20- to 22-nucleotide (nt) RNAs generated from a hairpin precursor and functioning in gene silencing and (2) short interfering RNAs (siRNAs), including 24-nt heterochromatic siRNAs typically functioning to repress repetitive regions of the genome via RNA-directed DNA methylation, as well as secondary phased siRNAs and trans-acting siRNAs generated via miRNA-directed cleavage of a polymerase II-derived RNA precursor. Our analyses described thousands of sRNA loci, including poorly understood clusters of 22-nt siRNAs that accumulate during viral infection. The birth, death, expansion, and contraction of these sRNA loci are dynamic evolutionary processes that characterize the Solanaceae family. These analyses indicate that individuals within the same genus share similar sRNA landscapes, whereas comparisons between distinct genera within the Solanaceae reveal relatively few commonalities.

Anatomical and biophysical basis for graft incompatibility within the Solanaceae.

Interspecies grafting is an economically relevant technique that allows beneficial shoot and root combinations from separate species to be combined. One hypothesis for the basis of graft compatibility revolves around taxonomic relatedness. To test how phylogenetic distance affects interspecific graft compatibility within the economically important Solanaceae subfamily, Solanoideae, we characterized the anatomical and biophysical integrity of graft junctions between four species: tomato (Solanum lycopersicum), eggplant (Solanum melongena), pepper (Capsicum annuum), and groundcherry (Physalis pubescens). We analyzed the survival, growth, integrity, and cellular composition of the graft junctions. Utilizing various techniques, we were able to quantitatively assess compatibility among the interspecific grafts. Even though most of our graft combinations could survive, we show that only intrageneric combinations between tomato and eggplant are compatible. Unlike incompatible grafts, the formation of substantial vascular reconnections between tomato and eggplant in the intrageneric heterografts likely contributed to biophysically stable grafts. Furthermore, we identified 10 graft combinations that show delayed incompatibility, providing a useful system to pursue deeper work into graft compatibility. This work provides new evidence that graft compatibility may be limited to intrageneric combinations within the Solanoideae subfamily. Further research amongst additional Solanaceous species can be used to test the extent to which our hypothesis applies to this family.

Modifications to a LATE MERISTEM IDENTITY1 gene are responsible for the major leaf shapes of Upland cotton (Gossypium hirsutum L.).

Leaf shape varies spectacularly among plants. Leaves are the primary source of photoassimilate in crop plants, and understanding the genetic basis of variation in leaf morphology is critical to improving agricultural productivity. Leaf shape played a unique role in cotton improvement, as breeders have selected for entire and lobed leaf morphs resulting from a single locus, okra (l-D1), which is responsible for the major leaf shapes in cotton. The l-D1 locus is not only of agricultural importance in cotton, but through pioneering chimeric and morphometric studies, it has contributed to fundamental knowledge about leaf development. Here we show that an HD-Zip transcription factor homologous to the LATE MERISTEM IDENTITY1 (LMI1) gene of Arabidopsis is the causal gene underlying the l-D1 locus. The classical okra leaf shape allele has a 133-bp tandem duplication in the promoter, correlated with elevated expression, whereas an 8-bp deletion in the third exon of the presumed wild-type normal allele causes a frame-shifted and truncated coding sequence. Our results indicate that subokra is the ancestral leaf shape of tetraploid cotton that gave rise to the okra allele and that normal is a derived mutant allele that came to predominate and define the leaf shape of cultivated cotton. Virus-induced gene silencing (VIGS) of the LMI1-like gene in an okra variety was sufficient to induce normal leaf formation. The developmental changes in leaves conferred by this gene are associated with a photosynthetic transcriptomic signature, substantiating its use by breeders to produce a superior cotton ideotype.

Connecting the pieces: uncovering the molecular basis for long-distance communication through plant grafting.

Vascular plants are wired with a remarkable long-distance communication system. This network can span from as little as a few centimeters (or less) in species like Arabidopsis, up to 100 m in the tallest giant sequoia, linking distant organ systems into a unified, multicellular organism. Grafting is a fundamental technique that allows researchers to physically break apart and reassemble the long-distance transport system, enabling the discovery of molecular signals that underlie intraorganismal communication. In this review, we highlight how plant grafting has facilitated the discovery of new long-distance signaling molecules that function in coordinating developmental transitions, abiotic and biotic responses, and cross-species interactions. This rapidly expanding area of research offers sustainable approaches for improving plant performance in the laboratory, the field, the orchard, and beyond.

The Persistent Homology Mathematical Framework Provides Enhanced Genotype-to-Phenotype Associations for Plant Morphology.

Efforts to understand the genetic and environmental conditioning of plant morphology are hindered by the lack of flexible and effective tools for quantifying morphology. Here, we demonstrate that persistent-homology-based topological methods can improve measurement of variation in leaf shape, serrations, and root architecture. We apply these methods to 2D images of leaves and root systems in field-grown plants of a domesticated introgression line population of tomato (Solanum pennellii). We find that compared with some commonly used conventional traits, (1) persistent-homology-based methods can more comprehensively capture morphological variation; (2) these techniques discriminate between genotypes with a larger normalized effect size and detect a greater number of unique quantitative trait loci (QTLs); (3) multivariate traits, whether statistically derived from univariate or persistent-homology-based traits, improve our ability to understand the genetic basis of phenotype; and (4) persistent-homology-based techniques detect unique QTLs compared to conventional traits or their multivariate derivatives, indicating that previously unmeasured aspects of morphology are now detectable. The QTL results further imply that genetic contributions to morphology can affect both the shoot and root, revealing a pleiotropic basis to natural variation in tomato. Persistent homology is a versatile framework to quantify plant morphology and developmental processes that complements and extends existing methods.

Genetic Architecture and Molecular Networks Underlying Leaf Thickness in Desert-Adapted Tomato Solanum pennellii.

Thicker leaves allow plants to grow in water-limited conditions. However, our understanding of the genetic underpinnings of this highly functional leaf shape trait is poor. We used a custom-built confocal profilometer to directly measure leaf thickness in a set of introgression lines (ILs) derived from the desert tomato Solanum pennellii and identified quantitative trait loci. We report evidence of a complex genetic architecture of this trait and roles for both genetic and environmental factors. Several ILs with thick leaves have dramatically elongated palisade mesophyll cells and, in some cases, increased leaf ploidy. We characterized the thick IL2-5 and IL4-3 in detail and found increased mesophyll cell size and leaf ploidy levels, suggesting that endoreduplication underpins leaf thickness in tomato. Next, we queried the transcriptomes and inferred dynamic Bayesian networks of gene expression across early leaf ontogeny in these lines to compare the molecular networks that pattern leaf thickness. We show that thick ILs share S. pennellii-like expression profiles for putative regulators of cell shape and meristem determinacy as well as a general signature of cell cycle-related gene expression. However, our network data suggest that leaf thickness in these two lines is patterned at least partially by distinct mechanisms. Consistent with this hypothesis, double homozygote lines combining introgression segments from these two ILs show additive phenotypes, including thick leaves, higher ploidy levels, and larger palisade mesophyll cells. Collectively, these data establish a framework of genetic, anatomical, and molecular mechanisms that pattern leaf thickness in desert-adapted tomato.

Plant chimeras: The good, the bad, and the 'Bizzaria'.

Chimeras - organisms that are composed of cells of more than one genotype - captured the human imagination long before they were formally described and used in the laboratory. These organisms owe their namesake to a fire-breathing monster from Greek mythology that has the head of a lion, the body of a goat, and the tail of a serpent. The first description of a non-fictional chimera dates back to the middle of the seventeenth century when the Florentine gardener Pietro Nati discovered an adventitious shoot growing from the graft junction between sour orange (Citrus aurantium) and citron (Citrus medica). This perplexing chimera that grows with sectors phenotypically resembling each of the citrus progenitors inspired discussion and wonder from the scientific community and was fittingly named the 'Bizzaria'. Initially, the 'Bizzaria' was believed to be an asexual hybrid that formed from a cellular fusion between the grafted parents; however, in-depth cellular analyses carried out centuries later demonstrated that the 'Bizzaria', along with other chimeras, owe their unique sectored appearance to a conglomeration of cells from the two donors. Since this pivotal discovery at the turn of the twentieth century, chimeras have served both as tools and as unique biological phenomena that have contributed to our understanding of plant development at the cellular, tissue, and organismal level. Rapid advancements in genome sequencing technologies have enabled the establishment of new model species with novel morphological and developmental features that enable the generation of chimeric organisms. In this review, we show that genetic mosaic and chimera studies provide a technologically simple way to delve into the organismal, genetic, and genomic inner workings underlying the development of diverse model organisms. Moreover, we discuss the unique opportunity that chimeras present to explore universal principles governing intercellular communication and the coordination of organismal biology in a heterogenomic landscape.

Cell-specific transcriptomic analyses of three-dimensional shoot development in the moss Physcomitrella patens.

Haploid moss gametophytes harbor distinct stem cell types, including tip cells that divide in single planes to generate filamentous protonemata, and bud cells that divide in three planes to yield axial gametophore shoots. This transition from filamentous to triplanar growth occurs progressively during the moss life cycle, and is thought to mirror evolution of the first terrestrial plants from Charophycean green algal ancestors. The innovation of morphologically complex plant body plans facilitated colonization of the vertical landscape, and enabled development of complex vegetative and reproductive plant morphologies. Despite its profound evolutionary significance, the molecular programs involved in this transition from filamentous to triplanar meristematic plant growth are poorly understood. In this study, we used single-cell type transcriptomics to identify more than 4000 differentially expressed genes that distinguish uniplanar protonematal tip cells from multiplanar gametophore bud cells in the moss Physcomitrella patens. While the transcriptomes of both tip and bud cells show molecular signatures of proliferative cells, the bud cell transcriptome exhibits a wider variety of genes with significantly increased transcript abundances. Our data suggest that combined expression of genes involved in shoot patterning and asymmetric cell division accompanies the transition from uniplanar to triplanar meristematic growth in moss.

Transcriptomic evidence for the evolution of shoot meristem function in sporophyte-dominant land plants through concerted selection of ancestral gametophytic and sporophytic genetic programs.

Alternation of generations, in which the haploid and diploid stages of the life cycle are each represented by multicellular forms that differ in their morphology, is a defining feature of the land plants (embryophytes). Anciently derived lineages of embryophytes grow predominately in the haploid gametophytic generation from apical cells that give rise to the photosynthetic body of the plant. More recently evolved plant lineages have multicellular shoot apical meristems (SAMs), and photosynthetic shoot development is restricted to the sporophyte generation. The molecular genetic basis for this evolutionary shift from gametophyte-dominant to sporophyte-dominant life cycles remains a major question in the study of land plant evolution. We used laser microdissection and next generation RNA sequencing to address whether angiosperm meristem patterning genes expressed in the sporophytic SAM of Zea mays are expressed in the gametophytic apical cells, or in the determinate sporophytes, of the model bryophytes Marchantia polymorpha and Physcomitrella patens. A wealth of upregulated genes involved in stem cell maintenance and organogenesis are identified in the maize SAM and in both the gametophytic apical cell and sporophyte of moss, but not in Marchantia. Significantly, meiosis-specific genetic programs are expressed in bryophyte sporophytes, long before the onset of sporogenesis. Our data suggest that this upregulated accumulation of meiotic gene transcripts suppresses indeterminate cell fate in the Physcomitrella sporophyte, and overrides the observed accumulation of meristem patterning genes. A model for the evolution of indeterminate growth in the sporophytic generation through the concerted selection of ancestral meristem gene programs from gametophyte-dominant lineages is proposed.

Dissecting the molecular signatures of apical cell-type shoot meristems from two ancient land plant lineages.

Shoot apical meristem (SAM) structure varies markedly within the land plants. The SAMs of many seedless vascular plants contain a conspicuous inverted, pyramidal cell called the apical cell (AC), which is unidentified in angiosperms. In this study, we use transcriptomic sequencing with precise laser microdissections of meristem subdomains to define the molecular signatures of anatomically distinct zones from the AC-type SAMs of a lycophyte (Selaginella moellendorffii) and a monilophyte (Equisetum arvense). The two model species for this study represent vascular plant lineages that diverged > 400 million yr ago. Our data comprise comprehensive molecular signatures for the distinct subdomains within AC-type SAMs, an anatomical anomaly whose functional significance has been debated in the botanical literature for over two centuries. Moreover, our data provide molecular support for distinct gene expression programs between the AC-type SAMs of Selaginella and Equisetum, as compared with the SAM transcriptome of the angiosperm maize. The results are discussed in light of the functional significance and evolutionary success of the AC-type SAM within the embryophytes.

Graft incompatibility between pepper and tomato can be attributed to genetic incompatibility between diverged immune systems.

Graft compatibility is the capacity of two plants to form cohesive vascular connections. Tomato and pepper are incompatible graft partners; however, the underlying cause of graft rejection between these two species remains unknown.We diagnosed graft incompatibility between tomato and diverse pepper varieties based on weakened biophysical stability, decreased growth, and persistent cell death using trypan blue and TUNEL assays. Transcriptomic analysis of cell death in the junction was performed using RNA-sequencing, and molecular signatures for incompatible graft response were characterized based on meta-transcriptomic comparisons with other biotic processes.We show that tomato is broadly incompatible with diverse pepper cultivars. These incompatible graft partners activate prolonged transcriptional changes that are highly enriched for defense processes. Amongst these processes was broad NLR upregulation and hypersensitive response. Using transcriptomic datasets for a variety of biotic stress treatments, we identified a significant overlap in the genetic profile of incompatible grafting and plant parasitism. In addition, we found over 1000 genes that are uniquely upregulated in incompatible grafts.Based on NLR overactivity, DNA damage, and prolonged cell death we have determined that tomato and pepper graft incompatibility is likely caused by a form of genetic incompatibility, which triggers a hyperimmune-response.

Chromosome-level Genome Assembly of Euphorbia peplus, a Model System for Plant Latex, Reveals that Relative Lack of Ty3 Transposons Contributed to Its Small Genome Size.

Euphorbia peplus (petty spurge) is a small, fast-growing plant that is native to Eurasia and has become a naturalized weed in North America and Australia. Euphorbia peplus is not only medicinally valuable, serving as a source for the skin cancer drug ingenol mebutate, but also has great potential as a model for latex production owing to its small size, ease of manipulation in the laboratory, and rapid reproductive cycle. To help establish E. peplus as a new model, we generated a 267.2-Mb Hi-C-anchored PacBio HiFi nuclear genome assembly with a BUSCO score of 98.5%, a genome annotation based on RNA-seq data from six organs, and publicly accessible tools including a genome browser and an interactive organ-specific expression atlas. Chromosome number is highly variable across Euphorbia species. Using a comparative analysis of our newly sequenced E. peplus genome with other Euphorbiaceae genomes, we show that variation in Euphorbia chromosome number between E. peplus and Euphorbia lathyris is likely due to fragmentation and rearrangement rather than chromosomal duplication followed by diploidization of the duplicated sequence. Moreover, we found that the E. peplus genome is relatively compact compared with related members of the genus in part due to restricted expansion of the Ty3 transposon family. Finally, we identify a large gene cluster that contains many previously identified enzymes in the putative ingenol mebutate biosynthesis pathway, along with additional gene candidates for this biosynthetic pathway. The genomic resources we have created for E. peplus will help advance research on latex production and ingenol mebutate biosynthesis in the commercially important Euphorbiaceae family.

ColourQuant: A High-Throughput Technique to Extract and Quantify Color Phenotypes from Plant Images.

Color patterning contributes to important plant traits that influence ecological interactions, horticultural breeding, and agricultural performance. High-throughput phenotyping of color is valuable for understanding plant biology and selecting for traits related to color during plant breeding. Here we present ColourQuant, an automated high-throughput pipeline that allows users to extract color phenotypes from images. This pipeline includes methods for color phenotyping using mean pixel values, a Gaussian density estimator of CIELAB color, and the analysis of shape-independent color patterning by circular deformation.

Getting to the root of grafting-induced traits.

Grafting is an ancient technique that involves the physical joining of genotypically distinct shoot and root systems, in order to achieve a desirable compound plant. This practice is widely used in modern agriculture to improve biotic and abiotic stress tolerance, modify plant architecture, induce precocious flowering and rejuvenate old perennial varieties, boost yield, and more. Beneficial new rootstock-scion combinations are currently identified through an inefficient trial and error process, which presents a significant bottleneck for the application of grafting to combat new environmental challenges. Identifying the mechanisms that underlie beneficial grafting-induced traits will facilitate rapid breeding and genetic engineering of new rootstock x scion combinations that exhibit superior performance across varying agricultural environments.

Growing a glue factory: Open questions in laticifer development.

Latex-containing cells called laticifers are present in at least 41 flowering plant families and are thought to have convergently evolved at least 12 times. These cells are known to function in defense, but little is known about the molecular genetic mechanisms of their development. The expansion of laticifers into their distinctive tube shape can occur through two distinct mechanisms, cell fusion and intrusive growth. The mechanism and extent of intrusive laticifer growth are still being investigated. Hormonal regulation by jasmonic acid and ethylene is important for both laticifer differentiation and latex biosynthesis. Current evidence suggests that laticifers can be specified independently of latex production, but extensive latex production requires specified laticifers. Laticifers are an emerging system for studying the intersection of cell identity specification and specialized metabolism.

TBtools: An Integrative Toolkit Developed for Interactive Analyses of Big Biological Data.

The rapid development of high-throughput sequencing techniques has led biology into the big-data era. Data analyses using various bioinformatics tools rely on programming and command-line environments, which are challenging and time-consuming for most wet-lab biologists. Here, we present TBtools (a Toolkit for Biologists integrating various biological data-handling tools), a stand-alone software with a user-friendly interface. The toolkit incorporates over 130 functions, which are designed to meet the increasing demand for big-data analyses, ranging from bulk sequence processing to interactive data visualization. A wide variety of graphs can be prepared in TBtools using a new plotting engine ("JIGplot") developed to maximize their interactive ability; this engine allows quick point-and-click modification of almost every graphic feature. TBtools is platform-independent software that can be run under all operating systems with Java Runtime Environment 1.6 or newer. It is freely available to non-commercial users at https://github.com/CJ-Chen/TBtools/releases.

Composite modeling of leaf shape along shoots discriminates Vitis species better than individual leaves.

PREMISE: Leaf morphology is dynamic, continuously deforming during leaf expansion and among leaves within a shoot. Here, we measured the leaf morphology of more than 200 grapevines (Vitis spp.) over four years and modeled changes in leaf shape along the shoot to determine whether a composite leaf shape comprising all the leaves from a single shoot can better capture the variation and predict species identity compared with individual leaves. METHODS: Using homologous universal landmarks found in grapevine leaves, we modeled various morphological features as polynomial functions of leaf nodes. The resulting functions were used to reconstruct modeled leaf shapes across the shoots, generating composite leaves that comprehensively capture the spectrum of leaf morphologies present. RESULTS: We found that composite leaves are better predictors of species identity than individual leaves from the same plant. We were able to use composite leaves to predict the species identity of previously unassigned grapevines, which were verified with genotyping. DISCUSSION: Observations of individual leaf shape fail to capture the true diversity between species. Composite leaf shape-an assemblage of modeled leaf snapshots across the shoot-is a better representation of the dynamic and essential shapes of leaves, in addition to serving as a better predictor of species identity than individual leaves.