(a) RT-qPCR quantifying the relative mRNA expression levels
of a panel of epidermal differentiation genes in scrambled control (CTLi) and
BRD4 (BRD4i) siRNA treated keratinocytes after three days of differentiation.
Two separate siRNAs (siRNA 1 and siRNA 2) targeting different regions of BRD4
mRNA were used (siRNA 1 n=4, siRNA 2 n=3). Statistics: t-test, ***p <
0.001. (b) RT-qPCR quantifying the relative mRNA expression levels
of LOR, BRD4, and FLG in CTLi and BRD4i day 5
regenerated human epidermis. n=5 (c) Immunofluorescent staining of
late differentiation markers LOR (green) and FLG (red) in CTLi and BRD4i day 5
regenerated human epidermis. Merged image includes Hoechst staining of nuclei.
n=3. White Scale bar = 20μm. (d) Heatmap generated for
replicate (n=2) RPKM normalized RNASequencing data from CTLi and BRD4i
keratinocytes differentiated for 3 days. The expression of genes significantly
increased (red) or decreased (blue) is shown. Differential expression was
determined with FDR ≤ 0.05 and fold change ≥ 2 vs. CTLi. Graphs
are displayed in log2 scale. (e) Gene ontology (GO) term enrichment
for the 1,791 genes significantly decreased in BRD4 knockdown cells.
(f) Gene ontology (GO) term enrichment for the 840 genes
significantly increased in expression in BRD4 knockdown cells.