Fig. 5

Gata2b+/− result in the impaired erythroid differentiation. (A) Coverage plots showing the gata2b chromatin accessibility in ‘erythroid progenitors 1’ cluster comparing wild type (WT) and gata2b+/−. The location of the gata2b gene block is shown in blue. The normalized peak signal range is 0–40. Highlighted genomic regions within grey blocks indicate the location of two putative enhancers, enhancer 1 (E1) and enhancer 2 (E2). (B) The dot plot displays the log2 fold-change values of differentially activated TF motifs in ‘erythroid progenitors 1’ cluster between WT and gata2b+/− kidney marrow (KM) in snATAC-seq. The transcription factor (TF) motifs are ranked in increasing order based on log2 fold-change values. Dotted lines indicate the threshold for significance, with absolute log2 fold-change values ≥0.1, the default parameter in function FindMarkers. The blue and red colours represent TF motifs that are statistically significant (p value <0.05) with absolute log2 fold-change values ≥0.1. (C) Uniform manifold approximation and projection (UMAP) analysis showing the subclusters of ‘erythroid progenitors 1’ cluster in 12 months postfertilization (mpf) scRNA-seq. UMAP atlas was separated between WT and gata2b+/− cells. (D) Cluster distribution quantification of ‘erythroid progenitors 1’ cluster in 12 mpf scRNA-seq between WT and gata2b+/− KM, with significantly differentially distributed clusters represented in pink. A statistical significance threshold of the false discovery rate <0.05 and absolute log2 fold change >1 were applied to determine the significance of the observed differences. (E) Pseudotime trajectory of ‘erythroid progenitors 1’ cluster in 12 mpf scRNA-seq, separated between WT and gata2b+/− cells. Dots indicating cells labelled by pseudotime scores. (F) Ridge chart showing the ‘erythroid progenitors 1’ cell distribution along with pseudotime trajectory in 12 mpf scRNA-seq, separated between WT and gata2b+/− cells. Ridge curves indicate the cell density after ordering by pseudotime score. Box chart showing the statistical pseudotime score analysis between WT and gata2b+/− ‘erythroid progenitors 1’ cells; t test was used to calculate p value. (G) Violin plot displaying the quantification and statistics of mki67 (left panel) and cdk1 (right panel) mRNA levels in ‘erythroid progenitors 1’ cluster between WT and gata2b+/− in 12 mpf scRNA-seq. t Test was used to calculate p value. (H) UMAP analysis showing the subclusters of ‘erythroid progenitors 1’ cluster in 18 mpf scRNA-seq. UMAP atlas was separated between WT and gata2b+/− cells. (I) Cluster distribution quantification of ‘erythroid progenitors 1’ cluster in 18 mpf scRNA-seq between WT and gata2b+/− KM, with significantly differentially distributed clusters represented in pink. The statistical significance threshold is the same as (D). (J) Pseudotime trajectory of ‘erythroid progenitors 1’ cluster in 18 mpf scRNA-seq. The left panel shows the cells labelled by pseudotime scores, separated between WT and gata2b+/− cells. The right panel shows the cells labelled by subclusters as shown in (H). (K) Ridge chart showing the ‘erythroid progenitors 1’ cell distribution along with pseudotime trajectory in 18 mpf scRNA-seq, separating between WT and gata2b+/− cells. Box chart showing the statistical pseudotime score analysis between 18mpf WT and gata2b+/− ‘erythroid progenitors 1’ cells, and t test was used to calculate p value. (L) Violin plot displaying the quantification and statistics of klf1 (left panel) and gata1a (right panel) mRNA level in ‘erythroid progenitors 1’ cluster between WT and gata2b+/− in 18 mpf scRNA-seq. t Test was used to calculate p value. (M) Down-regulated pathways in 18 mpf gata2b+/− ‘erythroid progenitors 1’ cluster. Dots are coloured by adjusted p value, and dot size indicates the enriched gene number. (N) Violin plot displaying the quantification and statistics of DNA repair score in ‘erythroid progenitors 1’ cluster between WT and gata2b+/− in 12 mpf (left panel) and 18 mpf (right panel) scRNA-seq. t Test was used to calculate p value. ns, not significant; ***p < 0.001; ****p < 0.0001.