Information de reference pour ce titreAccession Number: | 00006178-200533060-00041.
|
Author: | Zhou, Hongxia 1; Xia, Xu Gang 1; Xu, Zuoshang 1,2,3,*
|
Institution: | (1)Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605, USA (2)Department of Cell Biology, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605, USA (3)Neuroscience Program, University of Massachusetts Medical School, 364 Plantation Street, Worcester, MA 01605, USA
|
Title: | An RNA polymerase II construct synthesizes short-hairpin RNA with a quantitative indicator and mediates highly efficient RNAi.[Miscellaneous Article]
|
Source: | Nucleic Acids Research. 33(6):e62, 2005.
|
Abstract: | RNA interference (RNAi) mediates gene silencing in many eukaryotes and has been widely used to investigate gene functions. A common method to induce sustained RNAi is introducing plasmids that synthesize short hairpin RNAs (shRNAs) using Pol III promoters. While these promoters synthesize shRNAs and elicit RNAi efficiently, they lack cell specificity. Monitoring shRNA expression levels in individual cells by Pol III promoters is also difficult. An alternative way to deliver RNAi is to use Pol II-directed synthesis of shRNA. Previous efforts in developing a Pol II system have been sparse and the results were conflicting, and the usefulness of those Pol II vectors has been limited due to low efficacy. Here we demonstrate a new Pol II system that directs efficient shRNA synthesis and mediates strong RNAi at levels that are comparable with the commonly used Pol III systems. In addition, this system synthesizes a marker protein under control of the same promoter as the shRNA, thus providing an unequivocal indicator, not only to the cells that express the shRNA, but also to the levels of the shRNA expression. This system may be adapted for in vivo shRNA expression and gene silencing.
(C) Copyright Oxford University Press 2005.
|
References: | 1. Zamore, P.D., Tuschl, T., Sharp, P.A., Bartel, D.P. (2000) RNAi: double-stranded RNA directs the ATP-dependent cleavage of mRNA at 21 to 23 nucleotide intervals. Cell, 101, 25-33
2. Elbashir, S.M., Lendeckel, W., Tuschl, T. (2001) RNA interference is mediated by 21- and 22-nucleotide RNAs. Genes Dev., 15, 188-200
3. Bernstein, E., Caudy, A.A., Hammond, S.M., Hannon, G.J. (2001) Role for a bidentate ribonuclease in the initiation step of RNA interference. Nature, 409, 363-366
4. Tomari, Y., Matranga, C., Haley, B., Martinez, N., Zamore, P.D. (2004) A protein sensor for siRNA asymmetry. Science, 306, 1377-1380
5. Liu, Q., Rand, T.A., Kalidas, S., Du, F., Kim, H.-E., Smith, D.P., Wang, X. (2003) R2D2, a bridge between the initiation and effector steps of the Drosophila RNAi pathway. Science, 301, 1921-1925
6. Liu, J., Carmell, M.A., Rivas, F.V., Marsden, C.G., Thomson, J.M., Song, J.-J., Hammond, S.M., Joshua-Tor, L., Hannon, G.J. (2004) Argonaute2 is the catalytic engine of mammalian RNAi. Science, 305, 1437-1441
7. Hammond, S.M., Bernstein, E., Beach, D., Hannon, G.J. (2000) An RNA-directed nuclease mediates post-transcriptional gene silencing in Drosophila cells. Nature, 404, 293-296
8. Elbashir, S.M., Harborth, J., Lendeckel, W., Yalcin, A., Weber, K., Tuschl, T. (2001) Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature, 411, 494-498
9. Shi, Y., Sawada, J.-I., Sui, G., Affar, E.B., Whetstine, J.R., Lan, F., Ogawa, H., Po-Shan Luke, M., Nakatani, Y., Shi, Y. (2003) Coordinated histone modifications mediated by a CtBP co-repressor complex. Nature, 422, 735-738
10. Hutvagner, G. and Zamore, P.D. (2002) A microRNA in a multiple-turnover RNAi enzyme complex. Science, 297, 2056-2060
11. Xia, H., Mao, Q., Paulson, H.L., Davidson, B.L. (2002) siRNA-mediated gene silencing in vitro and in vivo. Nat. Biotechnol., 20, 1006-1010
12. Tiscornia, G., Singer, O., Ikawa, M., Verma, I.M. (2003) A general method for gene knockdown in mice by using lentiviral vectors expressing small interfering RNA. Proc. Natl Acad. Sci. USA, 100, 1844-1848
13. Hasuwa, H., Kaseda, K., Einarsdottir, T., Okabe, M. (2002) Small interfering RNA and gene silencing in transgenic mice and rats. FEBS Lett., 532, 227-230
14. Xia, H., Mao, Q., Eliason, S.L., Harper, S.Q., Martins, I.H., Orr, H.T., Paulson, H.L., Yang, L., Kotin, R.M., Davidson, B.L. (2004) RNAi suppresses polyglutamine-induced neurodegeneration in a model of spinocerebellar ataxia. Nat. Med., 10, 816-820
15. Kawasaki, H. and Taira, K. (2003) Short hairpin type of dsRNAs that are controlled by tRNA(Val) promoter significantly induce RNAi-mediated gene silencing in the cytoplasm of human cells. Nucleic Acids Res., 31, 700-707
16. Bridge, A.J., Pebernard, S., Ducraux, A., Nicoulaz, A.L., Iggo, R. (2003) Induction of an interferon response by RNAi vectors in mammalian cells. Nature Genet., 34, 263-264
17. Jackson, A.L., Bartz, S.R., Schelter, J., Kobayashi, S.V., Burchard, J., Mao, M., Li, B., Cavet, G., Linsley, P.S. (2003) Expression profiling reveals off-target gene regulation by RNAi. Nat. Biotechnol., 21, 635-637
18. Lagos-Quintana, M., Rauhut, R., Meyer, J., Borkhardt, A., Tuschl, T. (2003) New microRNAs from mouse and human. RNA, 9, 175-179
19. Lee, Y., Kim, M., Han, J., Yeom, K.H., Lee, S., Baek, S.H., Kim, V.N. (2004) MicroRNA genes are transcribed by RNA polymerase II. EMBO J., 23, 4051-4060
20. Zeng, Y. and Cullen, B.R. (2003) Sequence requirements for micro RNA processing and function in human cells. RNA, 9, 112-123
21. Miyagishi, M., Sumimoto, H., Miyoshi, H., Kawakami, Y., Taira, K. (2004) Optimization of an siRNA-expression system with an improved hairpin and its significant suppressive effects in mammalian cells. J. Gene Med., 6, 715-723
22. Ling, X. and Li, F. (2004) Silencing of antiapoptotic survivin gene by multiple approaches of RNA interference technology. Biotechniques, 36, 450-454 456-460
23. Gou, D., Narasaraju, T., Chintagari, N.R., Jin, N., Wang, P., Liu, L. (2004) Gene silencing in alveolar type II cells using cell-specific promoter in vitro and in vivo. Nucleic Acids Res., 32, e134
24. Lagos-Quintana, M., Rauhut, R., Lendeckel, W., Tuschl, T. (2001) Identification of novel genes coding for small expressed RNAs. Science, 294, 853-858
25. Li, X., Makela, S., Streng, T., Santti, R., Poutanen, M. (2003) Phenotype characteristics of transgenic male mice expressing human aromatase under ubiquitin C promoter. J. Steroid Biochem. Mol. Biol., 86, 469-476
26. Xia, X.G., Zhou, H., Ding, H., Affar el, B., Shi, Y., Xu, Z. (2003) An enhanced U6 promoter for synthesis of short hairpin RNA. Nucleic Acids Res., 31, e100
27. Cashman, N.R., Durham, H.D., Blusztajn, J.K., Oda, K., Tabira, T., Shaw, I.T., Dahrouge, S., Antel, J.P. (1992) Neuroblastoma x spinal cord (NSC) hybrid cell lines resemble developing motor neurons. Dev. Dyn., 194, 209-221
28. Yang, S., Tutton, S., Pierce, E., Yoon, K. (2001) Specific double-stranded RNA interference in undifferentiated mouse embryonic stem cells. Mol. Cell. Biol., 21, 7807-7816
29. Zeng, Y., Wagner, E.J., Cullen, B.R. (2002) Both natural and designed micro RNAs can inhibit the expression of cognate mRNAs when expressed in human cells. Mol. Cell, 9, 1327-1333
30. Lee, Y., Ahn, C., Han, J., Choi, H., Kim, J., Yim, J., Lee, J., Provost, P., Radmark, O., Kim, S., et al. (2003) The nuclear RNase III Drosha initiates microRNA processing. Nature, 425, 415-419
31. Lee, Y., Jeon, K., Lee, J.T., Kim, S., Kim, V.N. (2002) MicroRNA maturation: stepwise processing and subcellular localization. EMBO J., 21, 4663-4670
32. Lois, C., Hong, E.J., Pease, S., Brown, E.J., Baltimore, D. (2002) Germline transmission and tissue-specific expression of transgenes delivered by lentiviral vectors. Science, 295, 868-872
33. Lee, M., Hyun, D.-H., Halliwell, B., Jenner, P. (2001) Effect of overexpression of wild-type and mutant Cu/Zn-superoxide dismutases on oxidative stress and cell death induced by hydrogen peroxide, 4-hydroxynonenal or serum deprivation: potentiation of injury by ALS-related mutant superoxide dismutases and protection by Bcl-2. J. Neurochem., 78, 209-220
|
Language: | English.
|
Document Type: | METHODS.
|
Journal Subset: | Clinical Medicine. Life Sciences.
|
ISSN: | 0305-1048
|
NLM Journal Code: | o8l, 0411011
|
Annotation(s) | |
|
|