To characterize the impact of butyrate exposure on the
mast cell transcriptome, we measured gene expression profiles of two
independent primary human mast cell cultures upon 24 h of 5mM butyrate
treatment using RNA-Sequencing (RNA-Seq). Across all genes detected, 551
were upregulated by butyrate whereas 864 genes were downregulated
(FDR<0.05; Fig. 1A,B). Correlation of gene expression values
between the two biological replicates was high, both before and after
butyrate treatment (R²>0.97, Supplementary Fig. 1A,B).
Pathway enrichment analyses indicated that downregulated genes were
mainly associated with leukocyte and mast cell activation (Fig. 1C,D,
upper panels). Indeed, expression of genes coding for proteins involved
in FcεRI- and MRGPRX2-mediated mast cell activation was
significantly downregulated (including BTK, SYK, LAT and MRGPRX2,
Supplementary Table 1), supporting our earlier findings that butyrate
inhibits mast cell activation induced via IgE and substance P (an
MRGPRX2 ligand). Upregulated genes were enriched in more diverse
biological pathways (Fig. 1C,D, lower panels), including genes involved
in responses to metal ions (i.e., MT1 family proteins), cellular
responses to external stimuli and Ras/Rab GTPase signaling (Fig. 1C,D,
lower panels, Supplementary Table 1).
The downregulated genes - in particular canonical mast
cell genes - showed substantially higher median basal expression values
than unaffected or upregulated genes (Fig. 1E). This is in line with aprevious study in cancer cells demonstrating that HDAC inhibitors are
more likely to repress highly expressed
genes 26. Conversely, upregulated genes
displayed slightly lower basal expression levels as compared to
unaffected genes (Fig. 1E). Nevertheless, high basal expression levels
were not solely predictive for responsiveness to butyrate treatment, as
the 50 most highly expressed genes did not respond to butyrate treatment
(Fig. 1F).
Together, these results show that butyrate, in primary
human mast cells, selectively downregulates gene expression associated
with leukocyte/mast cell activation and upregulates a more diverse group
of genes. Basal expression levels alone only partially
predict responsiveness to butyrate treatment, indicating a more
selective mechanism of action through which butyrate exerts it
effects.