Comparison of the effects of pachymic acid, moronic acid and hydrocortisone on the polysome loading of RNAs in lipopolysaccharide-treated THP-1 macrophages.

We have proposed that analysis of ribosome-loaded mRNAs (i.e., the translatome) is useful for elucidation of pharmacological effects of phytocompounds in immune cells, regarding the involvement of post-transcriptional regulation mechanisms. In the present study, we compared the effects of pachymic acid from Poria cocos fungus and moronic acid from propolis with those of hydrocortisone on the translatomes of THP-1 macrophages exposed to bacterial lipopolysaccharide (LPS) to find clues to their biological effects. Polysome-associated RNAs collected from cells treated for 3 h with LPS plus each of the compounds were analyzed by DNA microarray followed by analyses of pathways/gene ontologies (GO). Upregulated mRNAs in enriched pathways that were found to contain AUUUA (AU)-rich motifs were checked by real-time PCR, and expression of candidate RNA-binding proteins stabilizing/destabilizing such AU-rich mRNAs was checked by Western blotting. The numbers of upregulated and downregulated genes (fold-changes ± 2.0 versus vehicle-control) were, respectively, 209 and 125 for moronic acid, 23 and 2 for pachymic acid, and 214 and 59 for hydrocortisone treatment. Overlapping with hydrocortisone treatment for upregulation were 158 genes in moronic acid and 17 in pachymic acid treatment; of these, 16 overlapped within all treatments (C-X-C motif chemokine ligands, interferon-induced protein with tetratricopeptide repeats, etc.). Pathway analyses showed GO enrichments such as 'immune response', 'receptor binding', 'extracellular space' etc. The pachymic acid-upregulated mRNAs (highly overlapped with the other 2 treatments) showed the presence of signal peptides and AU-rich motifs, suggesting regulation by AU-rich element (ARE)-binding proteins. The expression of ARE-binding protein HuR/ELAV-1 was increased by the 3 compounds, and AUF1/hnRNP D was decreased by pachymic acid. These results suggested that pachymic acid and moronic acid effects may involve as yet unknown post-transcriptional modulation via ARE-binding proteins resembling that of glucocorticoids.