Fig. 2
miRNA, gene targets, and ontologies associated with maternal separation and enrichment. a two sizeable groups of miRNAs altered by MS and Enr localized onto chromosomes 6 and 10. Direction of fold change in MS relative to controls, MS + Enr relative to controls, and MS + Enr relatve to MS are represented as circles, diamonds, and triangles, respectively (blue indicates upregulation and green, downregulation). b miRNA gene target network shows that 8 significantly altered miRNAs target many stress-related genes and share many targets. c MetascapeGO revealed functions most associated with significant miRNAs’ gene targets (Top 3: rno05200-“pathways in cancer”, rno04014-“Ras signaling pathway”, and GO:0010720-“postive regulation of cell development”; more significant bars are darker colored). d Metascape also mapped similar ontologies based on common member genes (top map) and significance (bottom map). There is a high level of overlap in membership of these gene targets in Metascape ontologies. e Changes in miRNA gene target expression was tested using qPCR. Bars represent M ± SEM and significance were determined by independent samples t-test (n = 24 animals per group) and one-way ANOVA (for GRIN2B). GABRA1, MAPK6, and MMP19 were significantly upregulated by MS (t(46)> 2.035, p < 0.048) and MAPK6 and MMP19 were partially recovered by Enr. GRIN2B was the only gene affected by the interaction between sex and group (F(2, 72)=3.703, p = 0.030). f CPG islands were identified near miR-promoter region using the UCSC genome browser; the blue arrow shows miR-212 to the left and miR-132 on the right. The row marked in green shows a CPG island. g methylation in the 3’UTR of miR-219-1 was significantly increased by MS (t(30)=2.29, p = 0.029); due to limited tissue availability, methylation was tested in n = 8 animals per group (compared to n = 12 for gene and miRNA expression). For miR-207, methylation in two distinct regions of one CPG island was tested. Significance: *p < 0.05. Abbreviations: MS-maternal separation, C-controls, Enr-environmental enrichment, UTR-untranslated region, SEM-standard error of the mean