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The regulatory landscape of Arabidopsis thaliana roots at single-cell resolution.

ABSTRACT: The scarcity of accessible sites that are dynamic or cell type-specific in plants may be due in part to tissue heterogeneity in bulk studies. To assess the effects of tissue heterogeneity, we apply single-cell ATAC-seq to Arabidopsis thaliana roots and identify thousands of differentially accessible sites, sufficient to resolve all major cell types of the root. We find that the entirety of a cell's regulatory landscape and its transcriptome independently capture cell type identity. We leverage this shared information on cell identity to integrate accessibility and transcriptome data to characterize developmental progression, endoreduplication and cell division. We further use the combined data to characterize cell type-specific motif enrichments of transcription factor families and link the expression of family members to changing accessibility at specific loci, resolving direct and indirect effects that shape expression. Our approach provides an analytical framework to infer the gene regulatory networks that execute plant development.

SUBMITTER: Dorrity MW

PROVIDER: S-EPMC8184767 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundThere is a growing interest in the role of chromatin in acquiring and maintaining cell identity. Despite the ever-growing availability of genome-wide gene expression data, understanding how transcription programs are established and regulated to define cell identity remains a puzzle. An important mechanism of gene regulation is the binding of transcription factors (TFs) to specific DNA sequence motifs across the genome. However, these sequences are hindered by the packaging of DNA to chromatin. Thus, the accessibility of these loci for TF binding is highly regulated and determines where and when TFs bind. We present a workflow for measuring chromatin accessibility in Arabidopsis thaliana and define organ-specific regulatory sites and binding motifs of TFs at these sites.ResultsWe coupled the recently described isolation of nuclei tagged in specific cell types (INTACT) and assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) as a genome-wide strategy to uncover accessible regulatory sites in Arabidopsis based on their accessibility to nuclease digestion. By applying this pipeline in Arabidopsis roots, we revealed 41,419 accessible sites, of which approximately half are found in gene promoters and contain the H3K4me3 active histone mark. The root-unique accessible sites from this group are enriched for root processes. Interestingly, most of the root-unique accessible sites are found in nongenic regions but are correlated with root-specific expression of distant genes. Importantly, these gene-distant sites are enriched for binding motifs of TFs important for root development as well as motifs for TFs that may play a role as novel transcriptional regulators in roots, suggesting that these accessible loci are functional novel gene-distant regulatory elements.ConclusionsBy coupling INTACT with ATAC-seq methods, we present a feasible pipeline to profile accessible chromatin in plants. We also introduce a rapid measure of the experiment quality. We find that chromatin accessibility at promoter regions is strongly associated with transcription and active histone marks. However, root-specific chromatin accessibility is primarily found at intergenic regions, suggesting their predominance in defining organ identity possibly via long-range chromatin interactions. This workflow can be rapidly applied to study the regulatory landscape in other cell types, plant species and conditions.

Project description:Gene duplication is a key evolutionary phenomenon, prevalent in all organisms but particularly so in plants, where whole genome duplication (WGD; polyploidy) is a major force in genome evolution. Much effort has been expended in attempting to understand the evolution of duplicate genes, addressing such questions as why some paralog pairs rapidly return to single copy status whereas, in other pairs, both paralogs are retained and may diverge in expression pattern or function. The effect of a gene - its site of expression and thus the initial locus of its function - occurs at the level of a cell comprising a single cell type at a given state of the cell's development. Using Arabidopsis thaliana single cell transcriptomic data we categorized patterns of expression for 11,470 duplicate gene pairs across 36 cell clusters comprising nine cell types and their developmental states. Among these 11,470 pairs, 10,187 (88.8%) had at least one copy expressed in at least one of the 36 cell clusters. Pairs produced by WGD more often had both paralogs expressed in root cells than did pairs produced by small scale duplications. Three quarters of gene pairs expressed in the 36 cell clusters (7,608/10,187) showed extreme expression bias in at least one cluster, including 352 cases of reciprocal bias, a pattern consistent with expression subfunctionalization. More than twice as many pairs showed reciprocal expression bias between cell states than between cell types or between roots and leaves. A group of 33 gene pairs with reciprocal expression bias showed evidence of concerted divergence of gene networks in stele vs. epidermis. Pairs with both paralogs expressed without bias were less likely to have paralogs with divergent mutant phenotypes; such bias-free pairs showed evidence of preservation by maintenance of dosage balance. Overall, we found considerable evidence of shifts in gene expression following duplication, including in >80% of pairs encoding 7,653 genes expressed ubiquitously in all root cell types and states for which we inferred the polarity of change.

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The regulatory landscape of Arabidopsis thaliana roots at single-cell resolution.

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