Modulation of Gene Expression by Human Cytosolic tRNase ZL through 5′-Half-tRNA

Abstract

A long form (tRNase ZL) of tRNA 3′ processing endoribonuclease (tRNase Z, or 3′ tRNase) can cleave any target RNA at any desired site under the direction of artificial small guide RNA (sgRNA) that mimics a 5′-half portion of tRNA. Based on this enzymatic property, a gene silencing technology has been developed, in which a specific mRNA level can be downregulated by introducing into cells a synthetic 5′-half-tRNA that is designed to form a pre-tRNA-like complex with a part of the mRNA. Recently 5′-half-tRNA fragments have been reported to exist stably in various types of cells, although little is know about their physiological roles. We were curious to know if endogenous 5′-half-tRNA works as sgRNA for tRNase ZL in the cells. Here we show that human cytosolic tRNase ZL modulates gene expression through 5′-half-tRNA. We found that 5′-half-tRNAGlu, which co-immunoprecipitates with tRNase ZL, exists predominantly in the cytoplasm, functions as sgRNA in vitro, and downregulates the level of a luciferase mRNA containing its target sequence in human kidney 293 cells. We also demonstrated that the PPM1F mRNA is one of the genuine targets of tRNase ZL guided by 5′-half-tRNAGlu. Furthermore, the DNA microarray data suggested that tRNase ZL is likely to be involved in the p53 signaling pathway and apoptosis.

Human cytosolic tRNase ZL can downregulate gene expression through miRNA

“…Recently, we have also shown that human cytosolic tRNase Z L modulates gene expression through cellular 5 0 -half-tRNA in the cells [18]. The present results together with this observation imply that other types of small cellular ncRNA would also mediate regulation of gene expression by tRNase Z L .…”

“…This observation astonished us since 5 0 -half-tRNA is the very RNA that we have been using as tailor-made sgRNA for TRUE gene silencing. We have also shown that the 5 0 -half-tRNA Glu indeed works as sgRNA in vitro and that human tRNase Z L modulates gene expression through the 5 0 -half-tRNA Glu [18]. Furthermore, we have demonstrated that the human PPM1F mRNA is one of the genuine targets of tRNase Z L guided by the 5 0 -half-tRNA Glu .…”

“…The 293 and HeLa cells were cultured as described previously [18]. The 293 or HeLa cells were transfected with plasmids or with plasmids and RNA using Lipofectamine 2000 (Invitrogen).…”

“…Real-time PCR for luciferase mRNA quantitation and 3 0 -RACE Real-time PCR for luciferase mRNA quantitation and 3 0 -RACE were performed as described previously [18].…”

A load of small RNAs in the sperm — how many bits of hereditary information?

“…Moreover, these fragments can induce formation of stress granules and cytoplasmic foci with untranslated mRNPs [9]. Elbarbary and coworkers [10] have studied gene expression modulated by human 5′-half-tRNA coupled with tRNase Z L or tRNase Zl activity. 5′-half-tRNA acts as a small guide RNA (sgRNA) for tRNase Z L , and efficiently leads to the suppression of gene expression of a specific mRNA containing complementary binding sequence of sgRNA.…”