Abstract
Ribozymes are naturally-occurring catalytic RNAs from the viroid world and are being engineered in the laboratory to perform sequence-specific cleavage of a desired mRNA target. Since their Nobel Prize-winning discovery, there has been considerable interest in the utility of ribozymes as gene therapeutic agents to silence disease-causing genes. This technology is not perfect, but extensive efforts to improve upon natural design of ribozymes have enabled these RNA molecules to perform various tasks. In this chapter, we highlight the construction of two types of ribozymes: conventional and hybrid hammerhead ribozymes. The hybrid ribozyme described here is an improved version of the basic hammerhead motif with the following features: (a) the use of the RNA polymerase III (polIII) tRNAVal promoter to achieve a high level of transcription, (b) 5′ linkage to the cloverleaf-shaped tRNAVal to enhance intracellular stability and cytoplasmic transport, and (c) a 3′ end poly-(A) tail to act as a “molecular anchor” for endogenous RNA helicases endowing the ribozyme ability to disentangle higher-order structures of the target mRNA. Randomized hybrid ribozyme libraries have been used successfully for revelation of gene functions involved in metastasis, invasion, differentiation, apoptosis, endoplasmic reticulum stress and may be extended to gene functions involved in innate or induced cellular senescence of human cells.
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Deocaris, C.C., Kaul, S.C., Wadhwa, R. (2007). Use of Ribozymes in Cellular Aging Research. In: Tollefsbol, T.O. (eds) Biological Aging. Methods in Molecular Biology™, vol 371. Humana Press. https://doi.org/10.1007/978-1-59745-361-5_16
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DOI: https://doi.org/10.1007/978-1-59745-361-5_16
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