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Abstract Short-read RNA-seq studies of grafted plants have led to the proposal that thousands of messenger RNAs (mRNAs) move over long distances between plant tissues^1–7, potentially acting as signals^8–12. Transport of mRNAs between cells and tissues has been shown to play a role in several physiological and developmental processes in plants, such as tuberization¹³, leaf development¹⁴and meristem maintenance¹⁵; yet for most mobile mRNAs, the biological relevance of transport remains to be determined^16–19. Here we perform a meta-analysis of existing mobile mRNA datasets and examine the associated bioinformatic pipelines. Taking technological noise, biological variation, potential contamination and incomplete genome assemblies into account, we find that a high percentage of currently annotated graft-mobile transcripts are left without statistical support from available RNA-seq data. This meta-analysis challenges the findings of previous studies and current views on mRNA communication. 

Abstract 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 viability stains. Transcriptomic analysis of 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 nucleotide-binding and leucine-rich repeat receptors (NLR) upregulation and genetic signatures indicative of an immune 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 hypothesize that tomato and pepper graft incompatibility is characterized by an immune response that triggers cell death which interferes with junction formation. 

Abstract 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. 

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. 

Abstract 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. E. 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 an 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 E. 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 to 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. 

Abstract 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. 

Abstract Grafting has been adopted for a wide range of crops to enhance productivity and resilience; for example, grafting of Solanaceous crops couples disease-resistant rootstocks with scions that produce high-quality fruit. However, incompatibility severely limits the application of grafting and graft incompatibility remains poorly understood. In grafts, immediate incompatibility results in rapid death, but delayed incompatibility can take months or even years to manifest, creating a significant economic burden for perennial crop production. To gain insight into the genetic mechanisms underlying this phenomenon, we developed a model system using heterografting of tomato (Solanum lycopersicum) and pepper (Capsicum annuum). These grafted plants express signs of anatomical junction failure within the first week of grafting. By generating a detailed timeline for junction formation, we were able to pinpoint the cellular basis for this delayed incompatibility. Furthermore, we inferred gene regulatory networks for compatible self-grafts and incompatible heterografts based on these key anatomical events, which predict core regulators for grafting. Finally, we examined the role of vascular development in graft formation and uncovered SlWOX4 as a potential regulator of graft compatibility. Following this predicted regulator up with functional analysis, we show that Slwox4 homografts fail to form xylem bridges across the junction, demonstrating that indeed, SlWOX4 is essential for vascular reconnection during grafting, and may function as an early indicator of graft failure.