Uncoupling of RNAi from Active Translation in Mammalian Cells

Overview

Journal RNA

Specialty Molecular Biology

Date 2004 Dec 3

PMID 15574516

Citations 16

Authors

Shuo Gu

John J Rossi

Affiliations

Soon will be listed here.

Abstract

Small inhibitory RNAs (siRNAs) are produced from longer RNA duplexes by the RNAse III family member Dicer. The siRNAs function as sequence-specific guides for RNA cleavage or translational inhibition. The precise mechanism by which siRNAs direct the RNA-induced silencing complex (RISC) to find the complementary target mRNA remains a mystery. Some biochemical evidence connects RNAi with translation making attractive the hypothesis that RISC is coupled with the translational apparatus for scanning mRNAs. Such coupling would facilitate rapid alignment of the siRNA antisense with the complementary target sequence. To test this hypothesis we took advantage of a well-characterized translational switch afforded by the ferritin IRE-IRP to analyze RNAi mediated cleavage of a target mRNA in the presence and absence of translation. Our results demonstrate that neither active translation nor unidirectional scanning is required for siRNA mediated target degradation. Our findings demonstrate that nontranslated mRNAs are highly susceptible to RNAi, and blocking scanning from both the 5' and 3' ends of an mRNA does not impede RNAi. Interestingly, RNAi is about threefold more active in the absence of translation.

Citing Articles

Workability of mRNA Sequencing for Predicting Protein Abundance.

Ponomarenko E, Krasnov G, Kiseleva O, Kryukova P, Arzumanian V, Dolgalev G Genes (Basel). 2023; 14(11).

PMID: 38003008 PMC: 10671741. DOI: 10.3390/genes14112065.

Translation can affect the antisense activity of RNase H1-dependent oligonucleotides targeting mRNAs.

Liang X, Nichols J, Sun H, Crooke S Nucleic Acids Res. 2017; 46(1):293-313.

PMID: 29165591 PMC: 5758896. DOI: 10.1093/nar/gkx1174.

High-throughput screening of effective siRNAs using luciferase-linked chimeric mRNA.

Pang S, Pokomo L, Chen K, Kamata M, Mao S, Zhang H PLoS One. 2014; 9(5):e96445.

PMID: 24831610 PMC: 4022502. DOI: 10.1371/journal.pone.0096445.

Poly(amidoamine) dendrimer nanocarriers and their aerosol formulations for siRNA delivery to the lung epithelium.

Conti D, Brewer D, Grashik J, Avasarala S, da Rocha S Mol Pharm. 2014; 11(6):1808-22.

PMID: 24811243 PMC: 4051247. DOI: 10.1021/mp4006358.

siRNA has greatly elevated mismatch tolerance at 3'-UTR sites.

Wei N, Zhang L, Huang H, Chen Y, Zheng J, Zhou X PLoS One. 2012; 7(11):e49309.

PMID: 23145149 PMC: 3493533. DOI: 10.1371/journal.pone.0049309.

References

Scacheri P, Rozenblatt-Rosen O, Caplen N, Wolfsberg T, Umayam L, Lee J . Short interfering RNAs can induce unexpected and divergent changes in the levels of untargeted proteins in mammalian cells. Proc Natl Acad Sci U S A. 2004; 101(7):1892-7. PMC: 357023. DOI: 10.1073/pnas.0308698100. View

Lafontaine D, Tollervey D . Birth of the snoRNPs: the evolution of the modification-guide snoRNAs. Trends Biochem Sci. 1998; 23(10):383-8. DOI: 10.1016/s0968-0004(98)01260-2. View

Todorov I, Smalko P, Galkin A . [Polysomal apparatus as a reflection of function interactions between translation and transcription processes during protein synthesis inhibited by cycloheximide]. Biokhimiia. 1977; 42(12):2149-59. View

Gonzalez A, Jimenez A, Vazquez D, Davies J, Schindler D . Studies on the mode of action of hygromycin B, an inhibitor of translocation in eukaryotes. Biochim Biophys Acta. 1978; 521(2):459-69. DOI: 10.1016/0005-2787(78)90287-3. View

Hentze M, Caughman S, Rouault T, Barriocanal J, Dancis A, Harford J . Identification of the iron-responsive element for the translational regulation of human ferritin mRNA. Science. 1987; 238(4833):1570-3. DOI: 10.1126/science.3685996. View

Theil E . Regulation of ferritin and transferrin receptor mRNAs. J Biol Chem. 1990; 265(9):4771-4. View

Merrick W . Mechanism and regulation of eukaryotic protein synthesis. Microbiol Rev. 1992; 56(2):291-315. PMC: 372869. DOI: 10.1128/mr.56.2.291-315.1992. View

Hamilton A, Baulcombe D . A species of small antisense RNA in posttranscriptional gene silencing in plants. Science. 1999; 286(5441):950-2. DOI: 10.1126/science.286.5441.950. View

Hammond S, Bernstein E, Beach D, Hannon G . An RNA-directed nuclease mediates post-transcriptional gene silencing in Drosophila cells. Nature. 2000; 404(6775):293-6. DOI: 10.1038/35005107. View

10.

Zamore P, Tuschl T, Sharp P, Bartel D . RNAi: double-stranded RNA directs the ATP-dependent cleavage of mRNA at 21 to 23 nucleotide intervals. Cell. 2000; 101(1):25-33. DOI: 10.1016/S0092-8674(00)80620-0. View

11.

Elbashir S, Harborth J, Lendeckel W, Yalcin A, Weber K, Tuschl T . Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature. 2001; 411(6836):494-8. DOI: 10.1038/35078107. View

12.

Hammond S, Boettcher S, Caudy A, Kobayashi R, Hannon G . Argonaute2, a link between genetic and biochemical analyses of RNAi. Science. 2001; 293(5532):1146-50. DOI: 10.1126/science.1064023. View

13.

Jacque J, Triques K, Stevenson M . Modulation of HIV-1 replication by RNA interference. Nature. 2002; 418(6896):435-8. PMC: 9524216. DOI: 10.1038/nature00896. View

14.

Kennerdell J, Yamaguchi S, Carthew R . RNAi is activated during Drosophila oocyte maturation in a manner dependent on aubergine and spindle-E. Genes Dev. 2002; 16(15):1884-9. PMC: 186417. DOI: 10.1101/gad.990802. View

15.

Zeng Y, Cullen B . RNA interference in human cells is restricted to the cytoplasm. RNA. 2002; 8(7):855-60. PMC: 1370302. DOI: 10.1017/s1355838202020071. View

16.

Hu W, Myers C, Kilzer J, Pfaff S, Bushman F . Inhibition of retroviral pathogenesis by RNA interference. Curr Biol. 2002; 12(15):1301-11. DOI: 10.1016/s0960-9822(02)00975-2. View

17.

Martinez J, Patkaniowska A, Urlaub H, Luhrmann R, Tuschl T . Single-stranded antisense siRNAs guide target RNA cleavage in RNAi. Cell. 2002; 110(5):563-74. DOI: 10.1016/s0092-8674(02)00908-x. View

18.

McManus M, Sharp P . Gene silencing in mammals by small interfering RNAs. Nat Rev Genet. 2002; 3(10):737-47. DOI: 10.1038/nrg908. View

19.

Caudy A, Myers M, Hannon G, Hammond S . Fragile X-related protein and VIG associate with the RNA interference machinery. Genes Dev. 2002; 16(19):2491-6. PMC: 187452. DOI: 10.1101/gad.1025202. View

20.

Ishizuka A, Siomi M, Siomi H . A Drosophila fragile X protein interacts with components of RNAi and ribosomal proteins. Genes Dev. 2002; 16(19):2497-508. PMC: 187455. DOI: 10.1101/gad.1022002. View