Motivation: MicroRNAs (miRNAs) play critical roles in gene regulation. Although it is well known that multiple miRNAs may work as miRNA modules to synergistically regulate common target mRNAs, the understanding of miRNA modules is still in its infancy.

Results: We employed the recently generated high throughput experimental data to study miRNA modules. We predicted 181 miRNA modules and 306 potential miRNA modules. We observed that the target sites of these predicted modules were in general weaker compared with those not bound by miRNA modules. We also discovered that miRNAs in predicted modules preferred to bind unconventional target sites rather than canonical sites. Surprisingly, contrary to a previous study, we found that most adjacent miRNA target sites from the same miRNA modules were not within the range of 10–130 nucleotides. Interestingly, the distance of target sites bound by miRNAs in the same modules was shorter when miRNA modules bound unconventional instead of canonical sites. Our study shed new light on miRNA binding and miRNA target sites, which will likely advance our understanding of miRNA regulation.

Availability and implementation: The software miRModule can be freely downloaded at http://hulab.ucf.edu/research/projects/miRNA/miRModule.

Supplementary information: Supplementary data are available at Bioinformatics online.

Contact: haihu@cs.ucf.edu or xiaoman@mail.ucf.edu.