A CHIMERIC HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 (HIV-1) MINIMAL REV RESPONSE ELEMENT-RIBOZYME MOLECULE EXHIBITS DUAL ANTIVIRAL FUNCTION AND INHIBITS CELL-CELL TRANSMISSION OF HIV-1

We have previously shown that hairpin ribozymes targeting the human im munodeficiency virus (HIV) genome can effectively inhibit virus replic ation in a variety of primary and cultured hematopoietic cells, To fur ther increase antiviral potency and minimize the chance of viral resis tance, we have now cloned the stem-loop II sequences of the HIV type 1 Rev response element into ribozyme transcription cassettes, Fusion RN A molecules were shown to function both as RNA decoys and ribozymes. S table Molt-1/8 cell lines expressing fusion RNA of stem-loop II and a ribozyme directed at the HIV type 1 U5 sequence (MSLMJT) or its disabl ed counterpart (MSLdMJT) were generated, The expression of fusion RNA was persistent for at least 6 months without apparent cytotoxicity. Wh en virus inhibition was examined after the cocultivation of transduced cells with chronically infected Jurkat cells, much greater protection was observed in MSLMJT cells than in MSLdMJT or MMJT (expressing only the ribozyme) cells, Furthermore, to specifically compare the ribozym e activities in various transduced cells, we determined the quantitati ve levels of proviral DNA in the first round of virus replication (7 h after infection with HXB2). By competitive PCR, the proviral DNA leve ls in MSLMJT and MMJT cells were found to be reduced to 1/7 and 1/3, r espectively, compared with those in MSLdMJT and MdMJT cells, These res ults suggest not only that the greater inhibition afforded by this fus ion RNA was due to its function both as decoy and ribozyme but also th at the ribozyme activity may be facilitated.