Invitro selection of adenine-dependent ribozyme against Tpl2/Cot oncogene

Hairpin ribozymes possess the properties of RNA sequence-specific recognition and site-specific cleavage. These properties make them a powerful extension of the antisense approach for the inhibition of gene expression. From a randomized RNA pool of hairpin ribozymes, using the systematic evolution of ligands by exponential enrichment, we have obtained an adenine-dependent hairpin ribozyme, Tpl2/Cot (tumour progression locus2) ribozyme, which cleaves the Tpl2/Cot kinase mRNA sequence at nucleotides A225[darrG226 relative to the start codon of translation. This serine/threonine kinase activates the mitogen-activated protein kinase pathway implicated in cell proliferation in cancer. The selected 'Tpl2/Cot-YL ribozyme' efficiently cleaves its target sequence incis and intrans; furthermore, the ribozyme efficiently cleaves a longer target sequence of 54 nucleotides intrans, as well as the full-length mRNA.