The cellular prion protein (PrP ^C ) is best known for its misfolded disease-causing conformer, PrP ^Sc . Because the availability of PrP ^C is often limiting for prion propagation, understanding its regulation may point to possible therapeutic targets. We sought to determine to what extent the human microRNAome is involved in modulating PrP ^C levels through direct or indirect pathways. We probed PrP ^C protein levels in cells subjected to a genome-wide library encompassing 2019 miRNA mimics using a robust time-resolved fluorescence-resonance screening assay. Screening was performed in three human neuroectodermal cell lines: U-251 MG, CHP-212 and SH-SY5Y. The three screens yielded 17 overlapping high-confidence miRNA mimic hits, 13 of which were found to regulate PrP ^C biosynthesis directly via binding to the PRNP 3'UTR, thereby inducing transcript degradation. The four remaining hits (miR-124-3p, 192-3p, 299-5p and 376b-3p) did not bind either the 3'UTR or CDS of PRNP, and were therefore deemed indirect regulators of PrP ^C . Our results show that multiple miRNAs regulate PrP ^C levels both directly and indirectly. These findings may have profound implications for prion disease pathogenesis and potentially also for their therapy. Furthermore, the possible role of PrP ^C as a mediator of Aβ toxicity suggests that its regulation by miRNAs may also impinge on Alzheimer's disease.