CORRELATION OF ACTIVITY WITH STABILITY OF CHEMICALLY-MODIFIED RIBOZYMES IN NUCLEI SUSPENSION

To examine hammerhead ribozyme activity in the nuclear environment, we have used nuclei isolated from HTLV-I tax transformed fibroblasts to evaluate ribozymes targeted against HTLV-I tax RNA. The ribozyme activ ity in nuclei suspension was strongly dependent on the resistance of t he particular ribozyme to endogenous nucleases, A ribozyme containing exclusively 2'-deoxynucleotides in its stems cleaved target RNA by its catalytic activity in the absence of proteins and caused degradation in their presence by induction of nuclear RNase H activity. A ribozyme containing 2'-amino- and 2'-fluoropyrimidine nucleosides in combinati on with terminal phosphorothioate linkages was significantly more stab le in nuclei suspension and also exhibited a more than threefold highe r cleavage efficacy than its unmodified counterpart. The increased res istance against nuclease degradation is mainly due to terminal phospho rothioate linkages, suggesting that both 5' and 3'-exonucleases are pr imarily responsible for the nuclear degradation of oligonucleotides.