Reelin is a secreted glycoprotein whose function is regulated by proteolysis. One of the specific cleavage sites of Reelin, called C-t, is located approximately between the sixth and seventh Reelin repeat but its exact site was unknown. We here show that a metalloprotease present in the culture supernatant of cerebellar granular neurons (CGN) cleaves Reelin between Ala2688 and Asp2689. A Reelin mutant in which Asp2689 is replaced by Lys (Reelin-DK) is resistant to C-t cleavage by culture supernatant of CGN. From biochemical characteristics and the cleavage site preference, meprin α and meprin β were suggested candidate proteases and both were confirmed to cleave Reelin at the C-t site. Meprin α cleaved Reelin-DK but meprin β did not. Actinonin, a meprin α and meprin β inhibitor, did not inhibit the Reelin-cleaving activity of CGN and the amount of Reelin fragments in brains of meprin β knock-out mice was not significantly different from that of the wild-type, indicating that meprin β does not play a major role in Reelin cleavage under basal conditions. We propose that meprin α and meprin β join the modulators of Reelin signalling as they cleave Reelin at a specific site and are upregulated under specific pathological conditions.