Robert Schmitz

University of Georgia | UGA · Department of Genetics

Molecular clocks are the basis for dating the divergence between lineages over macro-evolutionary timescales (~10^5-10^8 years). However, classical DNA-based clocks tick too slowly to inform us about the recent past. Here, we demonstrate that stochastic DNA methylation changes at a subset of cytosines in plant genomes possess a clock-like behavior....

Cytosine methylation and its machinery influence the gene expression of many eukaryotes; however, insects are an exception to this general tenet despite many lineages retaining methyltransferases and methylated genomes. Here, we tested the a priori hypothesis that perturbed genetic pathways will be associated with meiosis using transcriptomics beca...

The large size and complexity of most fern genomes have hampered efforts to elucidate fundamental aspects of fern biology and land plant evolution through genome-enabled research. Here we present a chromosomal genome assembly and associated methylome, transcriptome and metabolome analyses for the model fern species Ceratopteris richardii . The asse...

PRECISE AND ACCURATE IDENTIFICATION AND QUANTIFICATION OF TRANSCRIPTIONAL START SITES (TSSS) IS KEY TO UNDERSTANDING THE SPATIOTEMPORAL CONTROL OF TRANSCRIPTION. THE CORE PROMOTER CONSISTS OF THE TSS AND PROXIMAL NONCODING SEQUENCES, WHICH ARE CRITICAL IN TRANSCRIPTIONAL REGULATION. THEREFORE, THE ACCURATE IDENTIFICATION OF TSSS IS IMPORTANT FOR UN...

Machine learning approaches have been applied to identify transcription factor (TF)-DNA interaction important for gene regulation and expression. However, due to the enormous search space of the genome, it is challenging to build models capable of surveying entire reference genomes, especially in species where models were not trained. In this study...

As the focus for CRISPR-edited plants moves from proof-of-concept to real-world applications, precise gene manipulation will increasingly require concurrent multiplex editing for polygenic traits. A common approach for editing across multiple sites is to design one gRNA per target; however, this complicates construct assembly and increases the poss...

Machine learning approaches have been applied to identify transcription factor (TF)-DNA interaction important for gene regulation and expression. However, due to the enormous search space of the genome, it is challenging to build models capable of surveying entire reference genomes, especially in species where models were not trained. In this study...

Understanding how cis-regulatory elements facilitate gene expression is a key question in biology. Recent advances in single-cell genomics have led to the discovery of cell-specific chromatin landscapes that underlie transcription programs in animal models. However, the high equipment and reagent costs of commercial systems limit their applications...

Plant tissues and organs are composed of functionally discrete cell types that are all defined by the same genome sequence. Cell-type variation in part arises from differential accessibility of cis-regulatory elements that encode the blueprints for transcriptional programs underlying cell identity and function. Owing to technical limitations, the r...

Mistakes in the maintenance of CG methylation are a source of heritable epimutations in plants. Multigenerational surveys indicate that the rate of these stochastic events varies substantially across the genome, with some regions harbouring localized ‘epimutation hotspots’. Using Arabidopsis as a model, we show that epimutation hotspots are indexed...

Chromatin accessibility, or the physical access to chromatinized DNA, is a widely studied characteristic of the eukaryotic genome. As active regulatory DNA elements are generally ‘accessible’, the genome-wide profiling of chromatin accessibility can be used to identify candidate regulatory genomic regions in a tissue or cell type. Multiple biochemi...

Epialleles are meiotically heritable variations in expression states that are independent from changes in DNA sequence. Although they are common in plant genomes, their molecular origins are unknown. Here we show, using mutant and experimental populations, that epialleles in Arabidopsis thaliana that result from ectopic hypermethylation are due to...

Understanding how cis-regulatory elements facilitate gene expression is a key question in biology. Recent advances in single-cell genomics have led to the discovery of cell-specific chromatin landscapes that underlie transcription programs. However, the high equipment and reagent costs of commercial systems limit their applications for many laborat...

Epigenomics is the study of molecular signatures associated with discrete regions within genomes, many of which are important for a wide range of nuclear processes. The ability to profile the epigenomic landscape associated with genes, repetitive regions, transposons, transcription, differential expression, cis-regulatory elements, and three-dimens...

Hair-like trichomes cover the aerial organs of many plant species and act as a barrier between a plant and its environment. They function in defense against biotic and abiotic stresses, while also serving as sites for synthesis and storage of secondary metabolites. Previously, the transcription factor PtaMYB186 was identified as a positive regulato...

Coupling assay for transposase-accessible chromatin sequencing (ATAC-seq) with microfluidic separation and cellular barcoding has emerged as a powerful approach to investigate chromatin accessibility of individual cells. Here, we define a protocol for constructing single-cell ATAC-seq libraries from maize seedling nuclei and the preliminary computa...

An a-maize-ing set of genomes Maize is an important crop cultivated worldwide. As maize spread across the world, selection for local environments resulted in variation, but the impact on differences between the genome has not been quantified. By producing high-quality genomic sequences of the 26 lines used in the maize nested association mapping pa...

Accurate genome annotations are essential to modern biology; however, they remain challenging to produce. Variation in gene structure and expression across species, as well as within an organism, make correctly annotating genes arduous; an issue exacerbated by pitfalls in current in-silico methods. These issues necessitate complementary approaches...

Background Regulation of chromatin accessibility and transcription are tightly coordinated processes. Studies in yeast and higher eukaryotes have described accessible chromatin regions, but little work has been done in filamentous fungi. Results Here we present a genome-scale characterization of accessible chromatin regions in Neurospora crassa ,...

Accessible chromatin and unmethylated DNA are associated with many genes and cis-regulatory elements. Attempts to understand natural variation for accessible chromatin regions (ACRs) and unmethylated regions (UMRs) often rely upon alignments to a single reference genome. This limits the ability to assess regions that are absent in the reference gen...

cis-regulatory elements (CREs) encode the genomic blueprints of spatiotemporal gene expression programs enabling highly specialized cell functions. Using single-cell genomics in six maize organs, we determined the cis- and trans-regulatory factors defining diverse cell identities and coordinating chromatin organization by profiling transcription fa...

Stochastic gains and losses of DNA methylation at CG dinucleotides are a frequent occurrence in plants. These spontaneous ‘epimutations’ occur at a rate that is 100 000 times higher than the genetic mutation rate, are effectively neutral at the genome-wide scale, and are stably inherited across mitotic and meiotic cell divisions. Mathematical model...

Given the importance of DNA methylation in protection of the genome against transposable elements and transcriptional regulation in other taxonomic groups, the diversity in both levels and patterns of DNA methylation in the insects raises questions about its function and evolution. We show that the maintenance DNA methyltransferase, DNMT1, affects...

Accessible chromatin and unmethylated DNA are associated with many genes and cis-regulatory elements. Attempts to understand natural variation for accessible chromatin regions (ACRs) and unmethylated regions (UMRs) often rely upon alignments to a single reference genome. This limits the ability to assess regions that are absent in the reference gen...

Recent pangenome studies have revealed a large fraction of the gene content within a species exhibits presence-absence variation (PAV). However, coding regions alone provide an incomplete assessment of functional genomic sequence variation at the species level. Little to no attention has been paid to noncoding regulatory regions in pangenome studie...

Transposable elements (TEs) have the potential to create regulatory variation both through the disruption of existing DNA regulatory elements and through the creation of novel DNA regulatory elements. In a species with a large genome, such as maize, many TEs interspersed with genes create opportunities for significant allelic variation due to TE pr...

We report de novo genome assemblies, transcriptomes, annotations, and methylomes for the 26 inbreds that serve as the founders for the maize nested association mapping population. The data indicate that the number of pan-genes exceeds 103,000 and that the ancient tetraploid character of maize continues to degrade by fractionation to the present day...

5-methylcytosine (5mC) is a modified base often described as necessary for the proper regulation of genes and transposons and for the maintenance of genome integrity in plants. However, the extent of this dogma is limited by the current phylogenetic sampling of land plant species diversity. Here, we report that a monocot plant, Spirodela polyrhiza...

Background: Plants can transmit somatic mutations and epimutations to offspring, which in turn can affect fitness. Knowledge of the rate at which these variations arise is necessary to understand how plant development contributes to local adaption in an ecoevolutionary context, particularly in long-lived perennials. Results: Here, we generate a...

Stochastic changes in DNA methylation (i.e., spontaneous epimutations) contribute to methylome diversity in plants. Here, we describe AlphaBeta, a computational method for estimating the precise rate of such stochastic events using pedigree-based DNA methylation data as input. We demonstrate how AlphaBeta can be employed to study transgenerationall...

Madin-Darby canine kidney II (MDCKII) cells are widely used to study epithelial morphogenesis. To better understand this process, we performed time course RNA-seq analysis of MDCKII 3D cystogenesis, along with polarized 2D cells for comparison. Our study reveals a biphasic change in the transcriptome that occurs after the first cell cycle and coinc...

Cis-regulatory elements (CREs) encode the genomic blueprints for coordinating spatiotemporal gene expression programs underlying highly specialized cell functions. To identify CREs underlying cell-type specification and developmental transitions, we implemented single-cell sequencing of Assay for Transposase Accessible Chromatin in an atlas of Zea...

Significance Crop genomes can be very large, with many repetitive elements and pseudogenes. Distilling a genome down to the relatively small fraction of regions that are functionally valuable for trait variation can be like looking for needles in a haystack. The location of these regions is often not obvious, and current detection technologies are...

While DNA methylation is an important chromatin modification in many groups of organisms, the function of DNA methylation within the insects is unclear. The taxonomic distribution of DNA methyltransferase genes in insects is highly variable, as is the presence of methylated genomes. In the large milkweed bug, Oncopeltus fasciatus, we have shown the...

The genomic sequences of crops continue to be produced at a frenetic pace. However, it remains challenging to develop complete annotations of functional genes and regulatory elements in these genomes. Here, we explore the potential to use DNA methylation profiles to develop more complete annotations. Using leaf tissue in maize, we define ∼100,000 u...

Transposable elements (TEs) have the potential to create regulatory variation both through disruption of existing DNA regulatory elements and through creation of novel DNA regulatory elements. In a species with a large genome, such as maize, the many TEs interspersed with genes creates opportunities for significant allelic variation due to TE prese...

Cotton is an important crop that has made significant gains in production over the last century. Emerging pests such as the reniform nematode have threatened cotton production. The rare African diploid species Gossypium longicalyx is a wild species that has been used as an important source of reniform nematode immunity. While mapping and breeding e...

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Base J, β-D-glucosyl-hydroxymethyluracil, is a modification of thymine DNA base involved in RNA Polymerase (Pol) II transcription termination in kinetoplastid protozoa. Little is understood regarding how specific thymine residues are targeted for J-modification or the mechanism of J regulated transcription termination. To identify proteins involved...

In plants and mammals, DNA methylation plays a critical role in transcriptional silencing by delineating heterochromatin from transcriptionally active euchromatin. A homeostatic balance between heterochromatin and euchromatin is essential to genomic stability. This is evident in many diseases and mutants for heterochromatin maintenance, which are c...

DNA methylation is an important chromatin modification that can stably alter gene expression in cells and maintain genome integrity in plants and vertebrates. The function of DNA methylation outside of these well-studied systems, however, is unclear. Insects, in particular, represent an understudied group. Variation in the level of DNA methylation...

Cotton is an important crop that has made significant gains in production over the last century. Emerging pests such as the reniform nematode have threatened cotton production. The rare African diploid species Gossypium longicalyx is a wild species that has been used as an important source of reniform nematode immunity. While mapping and breeding e...

Background Plants can transmit somatic mutations and epimutations to offspring, which in turn can affect fitness. Knowledge of the rate at which these variations arise is necessary to understand how plant development contributes to local adaption in an eco-evolutionary context, particularly in long-lived perennials. Results Here, we generated a ne...

Introduction: Heritable changes in cytosine methylation can arise stochastically in plant genomes independently of DNA sequence alterations. These so-called "spontaneous epimutations" appear to be a byproduct of imperfect DNA methylation maintenance during mitotic or meitotic cell divisions. Accurate estimates of the rate and spectrum of these stoc...

Chromatin accessibility and modification is a hallmark of regulatory DNA, the study of which led to the discovery of cis-regulatory elements (CREs). Here, we characterize chromatin accessibility, histone modifications and sequence conservation in 13 plant species. We identified thousands of putative CREs and revealed that distal CREs are prevalent...

Genetic mapping studies on crops suggest that agronomic traits can be controlled by gene–distal intergenic loci. Despite the biological importance and the potential agronomic utility of these loci, they remain virtually uncharacterized in all crop species to date. Here, we provide genetic, epigenomic and functional molecular evidence to support the...

DNA methylation is an important chromatin modification that can stably alter gene expression in cells and maintain genome integrity in plants and vertebrates. The function of DNA methylation outside of these well-studied systems, however, is unclear. Insects, in particular, represent an understudied group. Variation in the level of DNA methylation...

Duplication and divergence of primary pathway genes underlie the evolution of plant specialized metabolism; however, mechanisms partitioning parallel hormone and defence pathways are often speculative. For example, the primary pathway intermediate ent-kaurene is essential for gibberellin biosynthesis and is also a proposed precursor for maize antib...

DNA methylation and epigenetic silencing play important roles in the regulation of transposable elements (TEs) in many eukaryotic genomes. A majority of the maize genome is derived from TEs that can be classified into different orders and families based on their mechanism of transposition and sequence similarity, respectively. TEs themselves are hi...

Somatic embryogenesis is an important tissue culture technique that sometimes leads to phenotypic variation via genetic and/or epigenetic changes. To understand the genomic and epigenomic impacts of somatic embryogenesis, we characterized soybean (Glycine max) epigenomes sampled from embryos at 10 different stages ranging from 6 weeks to 13 years o...

The central regulator of the ethylene (ET) signaling pathway, which controls a plethora of developmental programs and responses to environmental cues in plants, is ETHYLENE-INSENSITIVE2 (EIN2). Here we identify a chromatin-dependent regulatory mechanism at EIN2 requiring two genes: ETHYLENE-INSENSITIVE6 (EIN6), which is a H3K27me3 demethylase also...

Recent pangenome studies have revealed a large fraction of the gene content within a species exhibits presence-absence variation (PAV). However, coding regions alone provide an incomplete assessment of functional genomic sequence variation at the species level. Little to no attention has been paid to noncoding regulatory regions in pangenome studie...

In many plant species, a subset of transcribed genes are characterized by strictly CG-context DNA methylation, referred to as gene body methylation (gbM). The mechanisms that establish gbM are unclear, yet flowering plant species naturally without gbM lack the DNA methyltransferase, CMT3, which maintains CHG (H = A, C, or T) and not CG methylation...

A major barrier to both metabolic engineering and fundamental biological studies is the lack of genetic tools in most microorganisms. One example is Clostridium thermocellum ATCC 27405T, where genetic tools are not available to help validate decades of hypotheses. A significant barrier to DNA transformation is restriction–modification systems, whic...

In flowering plants, gene body methylation (gbM) is associated with a subset of constitutively expressed genes. It has been proposed that gbM modulates gene expression. Here, we show that there are no consistent and direct differences to expression following the loss of gbM. By comparing expression of gbM genes in Arabidopsis thaliana accessions to...

The generation of thousands of fungal genomes is leading to a better understanding of genes and genomic organization within the kingdom. However, the epigenome, which includes DNA and chromatin modifications, remains poorly investigated in fungi. Large comparative studies in animals and plants have deepened our understanding of epigenomic variation...

AUXIN RESPONSE FACTORS (ARFs) are plant-specific transcription factors (TFs) that couple perception of the hormone auxin to gene expression programs essential to all land plants. As with many large TF families, a key question is whether individual members determine developmental specificity by binding distinct target genes. We use DAP-seq to genera...

Fireflies and their luminous courtships have inspired centuries of scientific study. Today firefly luciferase is widely used in biotechnology, but the evolutionary origin of bioluminescence within beetles remains unclear. To shed light on this long-standing question, we sequenced the genomes of two firefly species that diverged over 100 million-yea...

Contents I. II. III. IV. References SUMMARY: Heritable gains or losses of cytosine methylation can arise stochastically in plant genomes independently of DNA sequence changes. These so-called 'spontaneous epimutations' appear to be a byproduct of imperfect DNA methylation maintenance and epigenome reinforcement events that occur in specialized cell...

The function of cytosine (DNA) methylation in insects remains unknown. Using RNA interference we provide evidence for the functional role of the maintenance DNA methyltransferase 1 ( Dnmt1 ) in Oncopeltus fasciatus , a hemimetabolous insect. Individuals depleted for Dnmt1 , and subsequently DNA methylation, failed to reproduce. Depletion of DNA met...

High-throughput short-read sequencing has revolutionized how transcriptomes are quantified and annotated. However, while Illumina short-read sequencers can be used to analyze entire transcriptomes down to the level of individual splicing events with great accuracy, they fall short of analyzing how these individual events are combined into complete...