Project: Extracellular vesicles

This project is focused on the content of plant extracellular vesicles (EVs). We observed that EVs are highly enriched in very small RNAs, ranging in size from 10 to 17 nucleotides, which we call “tiny RNAs” (tyRNAs) – largely overlooked in past analyses.  Do tyRNAs have a biological function? Are they merely cellular waste products? Other small RNAs are enriched in EVs or in the intercellular wash fluid depleted of EVs, and are additional targets of investigation.

Project: Localizing RNAs

We collaborate with Jeff Caplan at the University of Delaware to develop and apply novel methods for RNA localization. Via this long-running collaboration, we’re utilizing super resolution and single-molecule approaches to quantify and localize RNAs of interest. The primary objective is to determine where and when different small RNAs and mRNAs are localized and accumulating. Most often, we’re focused on anther biology, as represented in these images, which display one lobe or a section of a lobe of a maize anther in cross-section, probed with RNAs involved in anther development. We are asking questions like: Where are these RNAs made? Where do they function? How are 24-nt secondary siRNAs made? And how do these processes vary across diverse plant species?

Project: Genomic analyses of small RNAs

In the Meyers lab, we develop and use a wide variety of bioinformatics tools and pipelines, mainly focused on genomic analyses of plant small RNAs. We have a customized genome browser that is optimized for the display, analysis, and query of small RNA, and mRNA data. We have developed apps for the analysis of small RNA targets, cleavage, genomic distributions, and variation across different genotypes. These tools help promote public access to our data and algorithms. Via collaboration with Hagit Shatkay at the University of Delaware, we have developed machine learning approaches to characterize different classes of plant small RNAs. We are asking questions like: What features distinguish different types of small RNAs? Which mRNAs or other types of RNAs are targets of reproductive small RNAs? Which small RNAs or classes of small RNAs are impacted in different mutants?

Project: Secondary siRNAs in anther development

Much of our current work focuses on phased, secondary siRNAs in anther development, mainly in collaboration with the lab of Virginia Walbot at Stanford University. There are two classes that accumulate to significant levels during grass anther development – premeiotic 21-nt phasiRNAs, and coincident with meiosis, 24-nt phasiRNAs. Together with former post-doc Rui Xia (now at South China Agricultural University), we have shown that the 24-nt class likely emerged coincident with flowering plants. When either class is perturbed, male sterility can result. Our major questions of interest include: What is the molecular basis of environmental sensitivity in grasses with disrupted reproductive phasiRNAs? What do the 24-mers do in tapetal development? What’s the role of the 21-nt reproductive phasiRNAs, and when & how did they evolve? And are there potential applications of these pathways for hybrid production in diverse grasses?

Project: phasiRNAs in wheat and barley

The self-fertilization limits the production of hybrid seeds in species of the Tritceae tribe. The control of pollen production can underpin the production of hybrid seeds in many crops and generates hybrid vigour known as a major factor contributing to high yield. To control pollen production, in wheat and barley, we are investigating the role of reproductive phasiRNAs as a candidate molecular mechanism to achieve  photoperiod/temperature conditional genic male sterility.

The primary emphasis of the Meyers lab is the analysis of small RNAs in plants. With our many collaborators, we have pioneered genomic analysis of small RNAs and their targets, working with “next-gen” sequencing technologies nearly since their invention.

Our work with next-gen sequencing stretches back to ~2001, when Blake was funded by the NSF to apply “MPSS” to the analysis of gene expression in Arabidopsis. This led to the development in the Meyers lab of the first publicly-accessible browser for next-gen data.

After moving to the University of Delaware in 2002, we continued to develop the application to mRNA and small RNA analyses of first MPSS, then 454, and finally the still-current Illumina SBS sequencing.

In 2005, with collaborator Pam Green, we were the first to use next-gen sequencing for the analysis of small RNAs, and in 2008, our labs co-developed “PARE” for the genome-wide analysis of cleaved mRNAs (PARE = Parallel Analysis of RNA Ends).

The Meyers lab has widely applied these methods to study plant genomes and their RNA products, and the lab continues to develop and apply novel informatics approaches for the analysis of RNA function in plants.

Specific areas of research include the use of these technologies to assess small RNA function and biogenesis in a broad range of plants, including Arabidopsis, maize, soybean, rice, and diverse other species. These data are being used to identify novel transcripts, to study small RNAs, microRNA targets, alternatively-polyadenylated transcripts, non-coding RNAs and gene silencing. The data are being analyzed to determine patterns of gene expression under different developmental conditions, for example to identify tissue-specific gene expression.

In 2005, with collaborator Pam Green, we were the first to use next-gen sequencing for the analysis of small RNAs, and in 2008, our labs co-developed “PARE” for the genome-wide analysis of cleaved mRNAs (PARE = Parallel Analysis of RNA Ends).

The Meyers lab has widely applied these methods to study plant genomes and their RNA products, and the lab continues to develop and apply novel informatics approaches for the analysis of RNA function in plants.

Specific areas of research include the use of these technologies to assess small RNA function and biogenesis in a broad range of plants, including Arabidopsis, maize, soybean, rice, and diverse other species. These data are being used to identify novel transcripts, to study small RNAs, microRNA targets, alternatively-polyadenylated transcripts, non-coding RNAs and gene silencing. The data are being analyzed to determine patterns of gene expression under different developmental conditions, for example to identify tissue-specific gene expression.

Our recent work focuses on phased secondary siRNAs in plants, including their function, evolution, and biogenesis. We have also created several databases with query & analysis tools to enable the use of these data for the scientific community. Our most popular site is Arabidopsis SBS, but we’ve also developed a number of other organism-specific variants of this site, available at our website overview page.

These studies are centered on the use of novel genomics tools and bioinformatics approaches to address fundamental questions of small RNA biology, gene expression, and genome regulation in plants.

Phased, secondary, small interfering RNAs (phasiRNAs) are of particular interest to us. Originally designated as trans-acting small interfering RNAs or tasiRNAs, the wider group of phasiRNAs are triggered by microRNAs and produced as siRNAs. Like microRNAs, phasiRNA function in the suppression of target transcript levels.

Data from a broad range of species have demonstrated that the count of phasiRNA generating-loci ranges from tens (Arabidopsis) to hundreds (Medicago, soybean, maize) to thousands (rice). In the dicots, phasiRNA sources and targets include several large or conserved families of genes, such as those encoding NB-LRR disease resistance proteins or transcription factors.

In some plants, NB-LRRs have a particularly high level of redundancy in miRNA and phasiRNA-mediated regulation.

In the grasses, phasiRNAs from non-coding RNAs are prevalent in anthers, during early development and meiosis.

Finally, we retain a bit of a historical interest in disease resistance genes in plants. The Nucleotide Binding Site-Leucine Rich Repeat (NB-LRR) proteins encoded by many resistance genes provide the first line of defense in many specific plant-pathogen interactions.

Approximately 150 of these proteins are encoded in the Arabidopsis Col-0 genome; with variable numbers ranging up to hundreds per plant genome in other species. We study sequence variation, function, and evolution in this class of genes. Connecting two of our long-standing interests, we were the first to describe microRNAs as “master regulators” via direct and indirect targeting (phasiRNAs) of this gene family.

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