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Arabidopsis Cell Atlas: a comprehensive single-cell transcriptomic atlas of Arabidopsis
Arabidopsis Cell Atlas: a comprehensive single-cell transcriptomic atlas of Arabidopsis

To clarify cell-type composition of all major organs of Arabidopsis and created a cell atlas, we use single nucleus RNA sequencing technology to reveal the cell type composition of 17 tissues across different developmental stages of the life cycle. We identified 47 cell types and clarified single-cell hierarchy for many tissues that previously have not been well characterized.

This atlas serves as a valuable resource for the entire Arabidopsis community and contributes a comprehensive reference for future investigations of non-model plant species at single cell resolution.

数据量: 158
更新时间: 2022-09-05

To clarify cell-type composition of all major organs of Arabidopsis and created a cell atlas, we use single nucleus RNA sequencing technology to reveal the cell type composition of 17 tissues across different developmental stages of the life cycle. We identified 47 cell types and clarified single-cell hierarchy for many tissues that previously have not been well characterized.

This atlas serves as a valuable resource for the entire Arabidopsis community and contributes a comprehensive reference for future investigations of non-model plant species at single cell resolution.

1.Background

Plant research is ushering in a new era of single-cell transcriptomics. However, a comprehensive single-cell transcriptomic landscape of the whole organism has not been achieved in any plant species.

Here we use single nucleus RNA sequencing to clarify cell-type composition of all major organs of Arabidopsis and created a cell atlas which includes over 150k cells from 17 tissues across different developmental stages of the life cycle. We identified over 47 cell types and uncovered single-cell hierarchy for many tissues that previously have not been well characterized. Comparative analyses across multiple organs revealed consensus and specific cell types.

We also constructed the differentiation trajectory using five successive developmental stages of reproductive organs from flower to silique, allowed the understanding of gene expression dynamic of tissue-specific cell type. Finally, we characterized cellular processes across different leaf ages. Our results revealed progressive accumulation of senescence related changes which are highly synchronized among major cell types, highlighting the importance of multi-cell-type coordination in critical physiological process although cellular heterogeneity. We also developed an Ageing-index to quantify the senescent state at single cell resolution. Facilitated by this index, weighted gene co-expression network analysis identified hundreds of novel hub genes that will elucidate the process of senescence.

This atlas serves as a valuable resource for the entire Arabidopsis community and contributes a comprehensive reference for future investigations of non-model plant species at single cell resolution.

Fig. 1 | A comprehensive cell atlas of Arabidopsis.
a. 17 tissues sampled (left) and numbers of cells/genes for each tissue captured (right).
b. t-Distributed stochastic neighbor embedding (t-SNE) analysis of Arabidopsis cell atlas data included eight organs. Forty-seven main cell type clusters are labeled in t-SNE map.
c. Tissue types are labeled in the t-SNE map of Arabidopsis cell atlas data, which are indicated as the color of spots in a.
d. 47 cell types identified in 17 individual tissue and the atlas.

2. Data description

2.1 Raw data

​ In total, 17 samples were collected at indicated day post-germination for all tissues including seedling, cotyledon, hypocotyl, root, rosette leaf, stem, lateral leaf, flower, and silique. Then we used an in-house nuclei isolation protocol (see method) and DNBelab C Series Single-Cell Library Prep Set (MGI, 1000021082) for snRNA data generation as previously described (Han et al., 2022). The concentration of DNA library was measured by Qubit (Invitrogen). Libraries were sequenced by DNBSEQ-T7RS.

2.2 Expression matrix

​ The raw sequencing reads were filtered and demultiplexed by PISA (Version 1.1.0) (https://github.com/shiquan/PISA), and aligned to the TAIR10 reference genome using STAR (version 2.7.4a) (Dobin et al., 2013) with default parameters. Besides standard data filtering, we used a more rigorous criteria with unique molecular identifier (UMI) counts > 1000 to obtain high-quality nuclei for downstream analyses. A total of 153,959 nuclei passed our quality control, with median genes between 1043–1947 and median UMIs between 1694–3927 per nucleus. Cells from each of the 17 tissues were clustered using Seurat and cells of representative tissues for all collected organs were integrated to generate a comprehensive transcriptome atlas.

3. Results

3.1 Cotyledon at DAG-6

Supplementary fig. 1 | snRNA-seq profiling of cotyledon (DAG-6)

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of cotyledon (DAG-6) and manually annotation of cell types.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.2 Hypocotyl at DAG-6

Supplementary fig. 2 | snRNA-seq profiling of hypocotyl (DAG-6)

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of hypocotyl (DAG-6) and manually annotation of cell types. Cell clusters giving “Unknown” labs means that their cell type annotations are not able to be defined due to lack of cell type specific markers.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.3 Root at DAG-6

Supplementary fig. 3 | snRNA-seq profiling of root (DAG-6)

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of root (DAG-6) and manually annotation of cell types. Cell clusters giving “Unknown” labs means that their cell type annotations are not able to be defined due to lack of cell type specific markers.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.4 Root at DAG-11

Supplementary fig. 4 | snRNA-seq profiling of root (DAG-11)

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of root (DAG-11) and manually annotation of cell types. Cell clusters giving “Unknown” labs means that their cell type annotations are not able to be defined due to lack of cell type specific markers.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.5 Stem at stage-5

Supplementary fig. 5 | snRNA-seq profiling of stem (Stage-5)

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of stem (Stage-5) and manually annotation of cell types. Cell clusters giving “Unknown” labs means that their cell type annotations are not able to be defined due to lack of cell type specific markers.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.6 Rosette at stage 1

Supplementary fig. 6 | 6. snRNA-seq profiling of rosette at stage 1

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of rosette at stage 1 and manually annotation of cell types.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.7 Rosette at stage 2

Supplementary fig. 7 | snRNA-seq profiling of rosette at stage 2

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of rosette at stage 2 and manually annotation of cell types.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.8 Rosette at stage 3

Supplementary fig. 8 | snRNA-seq profiling of rosette at stage 3

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of rosette at stage 3 and manually annotation of cell types.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.9 Rosette at stage 4

Supplementary fig. 9 | snRNA-seq profiling of rosette at stage 4

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of rosette at stage 4 and manually annotation of cell types.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.10 Rosette at stage 5

Supplementary fig. 10 | snRNA-seq profiling of rosette at stage 5

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of rosette at stage 5 and manually annotation of cell types.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.11 Rosette at stage 6

Supplementary fig. 11 | snRNA-seq profiling of rosette at stage 6

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of rosette at stage 6 and manually annotation of cell types.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.12 Cauline at Stage 5

Supplementary fig. 12 | snRNA-seq profiling of cauline (Stage 5)

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of cauline at Stage 5 and manually annotation of cell types.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.13 Early flower

Supplementary fig. 13 | snRNA-seq profiling of early flower

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of early flower and manually annotation of cell types. Cell clusters giving “Unknown” labs means that their cell type annotations are not able to be defined due to lack of cell type specific markers.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.14 Middle flower

Supplementary fig. 14 | snRNA-seq profiling of middle flower

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of middle flower and manually annotation of cell types. Cell clusters giving “Unknown” labs means that their cell type annotations are not able to be defined due to lack of cell type specific markers.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.15 Late flower

Supplementary fig. 15 | snRNA-seq profiling of late flower

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of late flower and manually annotation of cell types. Cell clusters giving “Unknown” labs means that their cell type annotations are not able to be defined due to lack of cell type specific markers.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.16 Early silique

Supplementary fig. 16 | snRNA-seq profiling of early silique

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of early silique and manually annotation of cell types. Cell clusters giving “Unknown” labs means that their cell type annotations are not able to be defined due to lack of cell type specific markers.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.

3.17 Middle silique

Supplementary fig. 17 | snRNA-seq profiling of middle silique

a. Uniform Manifold Approximation and Projection (UMAP) plot showing the cell clustering of middle silique and manually annotation of cell types. Cell clusters giving “Unknown” labs means that their cell type annotations are not able to be defined due to lack of cell type specific markers.

b. Dot plot showing the expression of marker genes in the indicated cell types. The color key from light blue to dark blue indicates low to high gene expression levels. The size of the dot indicates the ratio of cells with expression compared to cells in that cluster.