Human RNase L tunes gene expression by selectively destabilizing the microRNA-regulated transcriptome

Abstract

Double-stranded RNA (dsRNA) activates the innate immune system of mammalian cells and triggers intracellular RNA decay by the pseudokinase and endoribonuclease RNase L. RNase L protects from pathogens and regulates cell growth and differentiation by destabilizing largely unknown mammalian RNA targets. We developed an approach for transcriptome-wide profiling of RNase L activity in human cells and identified hundreds of direct RNA targets and nontargets. We show that this RNase L-dependent decay selectively affects transcripts regulated by microRNA (miR)-17/miR-29/miR-200 and other miRs that function as suppressors of mammalian cell adhesion and proliferation. RNase L mimics the effects of these miRs and acts as a suppressor of proliferation and adhesion in mammalian cells. Our data suggest that RNase L-dependent decay serves to establish an antiproliferative state via destabilization of the miRregulated transcriptome.R Nase L is a mammalian endoribonuclease regulated by the action of dsRNA and IFNs α/β/λ, which induce the intracellular synthesis of a specific RNase L activator, 2-5A (1). RNA cleavage is thought to account for all biological functions of the RNase L·2-5A complex, including innate immunity during the IFN response (2, 3), and regulation of cell cycle (4), proliferation (5), adipocyte differentiation (6), and apoptosis (7). RNase L inhibits translation by site-specific cleavage of 18S and 28S rRNA (8) and activates transcription and the inflammasome NLRP3 by releasing signaling RNA fragments (2, 9, 10). These mechanisms complement or operate in parallel with posttranscriptional gene control via regulated decay of some mRNAs, including myogenic regulatory factor MyoD (11), components of IFN signaling ISG43 and ISG15 (12), translation-inhibiting kinase PKR (13), cathepsin E gastric protease (3), 3′-UTRbinding protein HuR (4), as well as ribosomal and mitochondrial protein-encoding mRNAs (14-16). Although the repression of these transcripts depends on RNase L, it remains unknown how many of them are cleaved physically and how many are downregulated indirectly-for example, via transcription.A recent RNA-sequencing (RNA-seq) study described some of the direct targets of RNase L (8). Cleavages were reported for 18S rRNA and U6 snRNA, however the experiment was designed to detect predominantly ribosomal reads and the direct impact of RNase L on mRNAs was not defined. Structural and biochemical studies found that RNase L cleaves RNA at the consensus sequence UN^N (N = A, U, G, or C;^is the cleavage location) (17-19). The UN^N motifs are abundant in all mammalian RNAs, suggesting that RNase L may degrade every mRNA it encounters, which surprisingly contrasts regulation of RNase L by the highly specific stimuli dsRNA, IFNs, and 2-5A.Mammalian cells contain a transmembrane RNase L homolog, a kinase/RNase Ire1, which drives the unfolded protein response and regulated Ire1-dependent decay (RIDD) (18,20). The cleavage consensus sequence of Ire1 (UĜC) is similarly relaxed, however Ire1 targets only specific mRNAs....

RNase L Suppresses Androgen Receptor Signaling, Cell Migration and Matrix Metalloproteinase Activity in Prostate Cancer Cells

“…Our results, consistent with other published data [21,22], shows that prostate cancer cells depleted of RNase L show greater migration in wound healing and transwell migration assays in response to fibronectin and serum (Figure 5). In reconstitution experiments, over-expression of RNase L suppressed cell migration compared to both endogenous levels and knockdown cells while activation of RNase L, which requires RNase L dimerization, inhibited cell migration (Figure 6).…”

“…RNase L also inhibits cell attachment to ECM substrates and cell spreading on fibronectin. An earlier report showed that activity of RNase L downregulates transcripts involved in cell adhesion both transcriptionally and post-transcriptionally [22]. In contrast, we observed the suppressive effect of RNase L on cell migration in cells expressing endogenous RNase L in the absence of activation or with mutants that lacked enzyme activity.…”

“…The cleaved RNA products amplify interferon production [18], activate inflammasome [19] and promote a switch from autophagy to apoptosis [20]. Recent reports show that RNase L negatively regulates cell migration and downregulates messenger RNAs (mRNAs) for cell adhesion [21,22]. While these established roles of RNase L may contribute to tumor development, they do not provide understanding of how mutations in RNase L predispose to prostate cancer.…”

miRNA in Cancer Pathology

“…RNase L protects from pathogens and controls cell growth and differentiation by disrupting largely unidentified mammalian RNA targets. Rath et al (2015) revealed that RNase L-dependent decay selectively disturbs transcripts controlled by microRNA (miR)-17/miR-29/miR-200 and other miRs that function as suppressors of mammalian cell adhesion and multiplication. RNase L acts as a suppressor of proliferation and adhesion in mammalian cells.…”

“…RNase L acts as a suppressor of proliferation and adhesion in mammalian cells. Rath et al (2015) concluded that RNase L-dependent decay starts an anti-proliferative state via disruption of the miR-controlled transcriptome.…”