We reasoned that integrating differential expression profiles from both microRNAs and mRNAs would provide greater insight into the perturbed gene networks associated with withdrawal/protracted withdrawal. Our microRNA-target association procedure allowed for both perfect and imperfect sequence matching between microRNA-mRNA pairs. This undoubtedly influenced our results since a microRNA’s function is determined in part by the extent of sequence complementarity with its target. In general, a perfect sequence match between microRNA and target is believed to cause mRNA degradation while an imperfect match is believed to result in translational repression [45]. Thus, utilizing only perfect-match microRNA-mRNA pairs would have omitted non-degraded mRNA targets from our analysis, potentially resulting in more precise associations. However, the relationship between microRNA-mRNA sequence complementarity and microRNA function is not absolute; and limiting our analysis exclusively to perfectly matched pairs would likely result in a significant loss of information. Thus, we opted to include both perfect and imperfect sequence matches, relying on dual analyses to ensure reliable results. Two different strategies were used to reveal relationships between DE microRNAs and their targets: discrete time-based analysis and a hierarchical cluster-based analysis.
