No evidence of functional co-adaptation between clustered microRNAs

A significant fraction of microRNA loci are organized in genomic clusters. The origin and evolutionary dynamics of these clusters have been extensively studied, although different authors have come to different conclusions. In a recent paper, it has been suggested that microRNAs in the same clusters evolve to target overlapping sets of genes. The authors interpret this as functional co- adaptation between clustered microRNAs. Here I reanalyze their results and I show that the observed overlap is mostly due to two factors: similarity between two seed sequences of a pair of clustered microRNAs, and the expected high number of common targets between pairs of microRNAs that have a large number of targets each. After correcting for these factors, I observed that clustered microRNAs from different microRNA families do not share more targets than expected by chance. During an exchange of correspondence and manuscripts, the authors of the original report acknowledged that the permutation methods they performed was not the method they described in their original paper. Here I show that the new permutation test proposed is biased and leads to systematic errors of the first kind, which will explain why the p-values reported were extremely (and unrealistically) low. I also discuss how to investigate the evolutionary dynamics of clustered microRNAs and their targets. In conclusion, there is no evidence of widespread functional co-adaptation between clustered microRNAs.