Fig 1.
A) Cells with trisomy 21 have increased acute heat shock activated HSF1 transcription factor function as determined by both PRO-seq and ATAC-seq analysis. B) Several trisomy 21 cellular changes may contribute to an increased response to heat shock.
Fig 2.
Individuals with trisomy 21 have elevated levels of some heat shock-regulated genes under normal conditions.
All Data from the Human Trisome project [36]. A) Several heat shock genes (HSPA8, DNAJA1, HSPH1, HSPA1A, SERPINH1) are differentially expressed (RNA-seq) in some blood cell lineages in individuals with trisomy 21 (purple) compared to disomic controls (green). B) HSF1 is different, but often down, in trisomy 21. Multiple clinical samples shown: whole blood (W), white blood cells (WBC), monocytes (M), bulk T cells (T), and skin fibroblasts (SF). Significance key: * ≤ 0.1, ** ≤ 0.01. C) Clinical blood sample proteomics data [37] shows elevated levels for some heat shock induced genes (HSPA1A, HSPB1, and HSPA8) in people with trisomy 21.
Fig 3.
Cells with trisomy 21 have increased chromatin accessibility near heat shock response elements compared to disomic controls.
A) Conceptual diagram of the conditions analyzed. Disomic cells (green) and trisomic cells (purple) at control (37°, light color) or mild heat shock (42°, dark color). B) ATAC-seq data traces of two heat shock regulated genes: HSPB1 (chr7:76301841–76308320) and SERPINH1 (chr11:75561294–75582876). One representative replicate is shown. Transcription factor enrichment analysis (TFEA) enrichment score (y-axis) in the C) disomic cell line and D) trisomic cell line ATAC-seq data [30]. Grey: non-significant TFs, colored: GC-corrected p-adjusted value of p<1x10-10. E) Scatter plot comparing heat shock-induced changes in transcription factor activity (E-values from TFEA) between disomic cells (x-axis) and trisomic cells (y-axis). Red: significant in both comparisons, Purple: significant in trisomy only, Green: significant in disomy only. All TFEA analyses (C-E) utilize both replicates. F) Averaged signal of ATAC-seq data for 1 kilobase region around active HSF1 motifs. One representative replicate is shown.
Fig 4.
A mild heat shock treatment induces more robust transcriptional changes in the trisomic cell line compared to disomic control.
A) Total normalized read count corrected PRO-seq gene traces at two heat shock regulated genes (Hspb1, Serpinh1) in the four cell types/conditions. One representative replicate is shown. Number of genes B) increasing or C) decreasing in response to heat shock (by DESeq2) in PRO-seq in one or both cell lines. TFEA enrichment score (y-axis) in the D) disomic cell line and E) trisomic cell line PRO-seq data [30]. Grey: non-significant TFs, colored: GC-corrected p-adjusted value of p<1x10-10. F) Scatter plot comparing heat shock induced changes in transcription factor activity (E-values from TFEA) between disomic cells (x-axis) and trisomic cells (y-axis). Red: significant in both comparisons, Purple: significant in trisomy only, Green: significant in disomy only. Panels B-F utilize both replicates. G) Average metaplot of PRO-seq data surrounding (± 500 bp) lymphoblastoid-active HSF1 motifs (at zero). One representative replicate is shown.
Fig 5.
Single Cell RNA-seq indicates that changes in heat shock-induced gene expression in trisomy 21 cells are population-wide.
A) Violin plots of percent of total normalized scRNA-seq gene counts per cell for two genes (CCT8, SUMO3) present on chromosome 21 and non-responsive to heat shock. B) Heatmap of the Z-scores of chromosome 21 genes showing increased reads in trisomy 21 cells. C) Violin plots of two heat shock responsive genes (HSPD1, HSP90AA1) not encoded on chromosome 21; the y-axis is the levels across more than 500 cells in each sample via scRNA-seq. D) Heatmap of the Z-scores of heat shock genes shows a general up-regulation in the expression of heat shock genes, rather than a few cells with extreme heat shock phenotypes. Violin plots showing the E) sum and F) median of Z-scores of all heat shock genes for more than 500 cells in each sample. The genes showed must be present in at least 75% of cells. All panels: control (light color) and heat shock (dark color) of disomic (green) and trisomic (purple) lymphoblastoid cells.