We have been developing a unique system for the downregulation of a gene expression through cutting a specific mRNA by the long form of tRNA 3 0 -processing endoribonuclease (tRNase Z L ) under the direction of small-guide RNA (sgRNA). However, the efficacy of this system and the involvement of tRNase Z L in the living cells were not clear. Here we show, by targeting the exogenous luciferase gene, that the efficacy of the sgRNA/tRNase Z L method can become comparable to that of the RNA interference technology and that the gene silencing is owing to tRNase Z L directed by sgRNA not owing to a simple antisense effect. We also show that tRNase Z L together with sgRNA can downregulate expression of the endogenous human genes Bcl-2 and glycogen synthase kinase-3b by degrading their mRNAs in cell culture. Furthermore, we demonstrate that a gene expression in the livers of postnatal mice can be inhibited by an only seven-nucleotide sgRNA. These data suggest that sgRNA might be utilized as therapeutic agents to treat diseases such as cancers and AIDS.