Single-cell RNA-sequencing analysis of Arabidopsis roots
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ABSTRACT:
ORGANISM(S): Arabidopsis thaliana
PROVIDER: GSE123013 | GEO | 2019/02/05
REPOSITORIES: GEO
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Project description:We provide an atlas of gene and protein expression in Arabidopsis root hair cells, generated by paired-end RNA-seq and LC/MS-MS analysis from protoplasts that carry the root hair-specific pEXP7-GFP reporter construct. In total, transcripts from 23,234 genes were detected in root hairs; those related to cell wall biosynthesis and translation differed most dramatically in abundance when compared to non-GFP root protoplasts.
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Project description:A single cell RNA sequencing atlas of the Arabidopsis root distinguishes cells both by developmental fate and time, revealing defining expression features that depict a complex cascade of developmental progressions from stem cell through differentiation supported by mirroring waves of transcription factor expression. Methods: mRNA profiles of 6-day-old wild-type (WT) and shortroot-knockout Arabidopsis thaliana roots were generated by deep sequencing of single cell (all) and bulk RNA libraries (wild type only), in duplicate (bulk & wild-type single cell) and singlicate (shr-3), using Illumina NextSeq. The sequence reads that passed quality filters were analyzed at the transcript level. Single Cell libraries were processed and analysed using Cell Ranger, STAR, Seurat. Bulk libraries were processed using Trimmomatic, STAR, HTSeq and DEseq2. Results: For Single Cell - Using an optimized data analysis workflow, we mapped ~87k sequence reads per wild-type cell to the Arabidopsis genome (TAIR10) and identified a median of 4,276 genes and 14,758 transcripts per cell. In total, transcripts for 16,975 genes were detected (RPM ≥1) from wild-type cells. After correction for read depth, this represents ~90% of genes detected by bulk RNA-seq of protoplasted root tissue. Bulk RNA-seq data identified genes induced by protplasting of the Arabidopsis root which could be discounted from single cell analysis. The global gene expression profiles of pooled scRNA-seq and bulk RNA-seq are highly correlated (r = 0.9) indicating that plant scRNA-seq is highly sensitive. Conclusions: Our high-resolution single cell RNA sequencing atlas of the Arabidopsis root captures precise temporal information for all major cell types, revealing new regulators and defining features foreach. Developmental trajectories derived from pseudotime analysis depict a finely resolved cascade of developmental progressions between stem cell and final differentiation supported by mirroring waves of transcription factor expression.
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Project description:Background: Single-cell reconstruction of gene regulatory programs provides an important tool to understand the cellular phenotypic variation in complex tissues and their response to endogenous and environmental stimuli. While the single-cell transcriptomes of several plant organs have been elucidated, the underlying chromatin landscapes remain largely unknown. Results: To comprehensively delineate chromatin accessibility during root development of an important crop, we applied single-cell ATAC-seq to 46,758 cells from rice root tips under normal and heat stress conditions. Our data revealed cell-type-specific accessibility variance across most of the major cell types and allowed us to identify sets of transcription factors which associate with accessible chromatin regions (ACRs). Using root hair differentiation as a model, we demonstrate that chromatin dynamics and gene expression dynamics during cell type differentiation correlate in pseudotime analyses. In addition to developmental trajectories, we describe chromatin responses to heat, and identify cell type specific accessibility changes to this key environmental stimulus. Conclusions: Our work provides a framework for the integrative analysis of regulatory dynamics in an important plant organ at single-cell resolution.
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Project description:Background: Single-cell reconstruction of gene regulatory programs provides an important tool to understand the cellular phenotypic variation in complex tissues and their response to endogenous and environmental stimuli. While the single-cell transcriptomes of several plant organs have been elucidated, the underlying chromatin landscapes remain largely unknown. Results: To comprehensively delineate chromatin accessibility during root development of an important crop, we applied single-cell ATAC-seq to 46,758 cells from rice root tips under normal and heat stress conditions. Our data revealed cell-type-specific accessibility variance across most of the major cell types and allowed us to identify sets of transcription factors which associate with accessible chromatin regions (ACRs). Using root hair differentiation as a model, we demonstrate that chromatin dynamics and gene expression dynamics during cell type differentiation correlate in pseudotime analyses. In addition to developmental trajectories, we describe chromatin responses to heat, and identify cell type specific accessibility changes to this key environmental stimulus. Conclusions: Our work provides a framework for the integrative analysis of regulatory dynamics in an important plant organ at single-cell resolution.
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Project description:Purpose: The root hair is a model for understanding evolution of individual cell differentiation programs in plants. We compare the expression of the genes that participate in root hair development between Arabidopsis and other vascular plants to assess the conservation/diversification of the root hair development programs in vascular plants. Methods: We used RNA-Seq, in triplicates, to measure the genome-wide transcription activity of the root-hair cells isolated by Fluorescence-activated cell sorting (FACS) in Arabidopsis (COBL9::GFP transgeneic line, AtRH) and rice (EXPA30::GFP transgenic line, OsRH). We also generated RNA-Seq data, in triplicates, on the Arabidopsis rhd6 WER::GFP and WT WER::GFP by FACS to identify the RHD6-regulating root hair morphogenesis genes (AtRHM). For Arabidopsis, rice, tomato, soybean, cucumber and maize, we used RNA-seq, in triplicates, to measure genome-wide transcription activity of root hair cells filtered by sieves after stirred in liquid nitrogen (HAIR genes). Each sample was trimmed to retain high-quality reads, mapped to the reference genome by TopHat, and quantified by Cufflinks. The number of raw reads of Arabidopsis rhd6 WER::GFP and WT WER::GFP sample was counted by HTSeq and analyzed by edgeR to identify the differentially expressed genes. Results: We defined the root-hair transcriptome in diverse vascular plant species and analyzed the relative conservation/divergence in the expression of a large set of gene families.
2017-05-11 | GSE85516 | GEO
Project description:In order to identify the possible protein targets of cyanide in the regulation of root hair, we have carried out a single cell proteomic approach to characterize the protein atlas in wild type root hair cells compared to the cas-c1 mutant cells. Root hair specific cells were obtained from isolated protoplast from root tissues of Arabidopsis wt-pCOBL9:GFP and cas-c1-pCOBL9:GFP lines. To analyze the effect of the CAS-C1 mutation, we isolated 1 x 106 root hair cells by Fluorescence-activated cell sorting (FACS) from three independent replicate of wt-pCOBL9:GFP and cas-c1-pCOBL9:GFP roots. The extracted proteins from each sample were trypsin-digested and the digested peptides were analyzed by liquid chromatography-high resolution mass spectrometry (LC-MS/MS) for protein identification. In wild type samples, we have identified 3829 unique proteins at a false discovery rate below 1% (FDR<1%), that represent almost 10% of the total Arabidopsis proteome. In protein extract of the cas-c1-pCOBL9:GFP roots samples, we have identified 3972 proteins of which 3515 were commons in both cell types, 310 were only identified y wild type and 457 only in cas-c1 mutant (Fig. 5).
2024-01-26 | PXD028991 | Pride