Fig. 3

Each diet group and age elicits its own miRNA expression profile but miR-98-3p is overexpressed in all CR and LA diets and may affect HAT and HDAC activity. a Heatmap of the 175 DE miRNAs across all diets and ages; created in R using the heatmap3 package. mir-98-3p is highlighted by a green box. Red = up-regulated, and blue = down-regulated, log2 fold change reported for statistically significant DE genes. White means no statistically significant change in expression. b qPCR confirmation of RNA-seq results, showing miR98-3p differential expression in all CR and LA supplemented diets. Orange bars are RNA-seq log2 FC and blue bars are qPCR FC (corrected to provide the true FC in the sample by Ratio – 1 if the sample was up-regulated and by (–1/Ratio) + 1 for down-regulated samples). The horizontal dotted line represents the 1 FC cutoff. Errors bars represent the standard error calculated for log2 FC as follows: (Standard error/Mean) × log2e. Snord96a, Snord95 and Snord68 were used as reference genes. c The difference in HDAC and HAT activity was calculated as a ratio of the transfection (TF) control and then transformed into true FC (by Ratio – 1 if the sample was up-regulated and by (–1/Ratio) + 1 for down-regulated samples). A cutoff of 1 FC was used, represented by the dotted line. The figure shows that the miR-98-3p inhibitor significantly (p < 0.001, two-tailed t-test) decreases HAT activity when compared with HDAC activity. A miR-98-3p mimic appears not to affect HDAC/HAT balance (no statistically significant difference). LA reduces HDAC activity (p < 0.001) and the untreated cells show a slight increase in both HAT and HDAC activity, indicating that transfection might affect HDAC and HAT activity but not HDAC/HAT balance (no statistically significant difference). The error bars represent the standard deviation. Experiments were performed independently three times with n = 3 wells per treatment