RNASE-L INHIBITOR (RLI) ANTISENSE CONSTRUCTIONS BLOCK PARTIALLY THE DOWN-REGULATION OF THE 2-5A RNASE-L PATHWAY IN ENCEPHALOMYOCARDITIS-VIRUS-(EMCV)-INFECTED CELLS/

The interferon-(IFN)-inducible 2',5'-oligoadenylate (2-5A)/endoribonuc lease L (RNase L) pathway plays a major role in the antiviral and anti proliferative effects of IFN. The 2-5A/RNase L pathway appears to be r egulated by the cell-growth status or viral infection. Viruses, and pi cornaviruses in particular, have evolved strategies to escape the 2-5A /RNase L-pathway-associated antiviral activity. We have recently clone d a cDNA coding for RLI, a RNase-L-specific protein inhibitor. Its reg ulated expression by viral infection could provide a new strategy to m odulate the 2-5A/RNase L pathway. Since RNase L had been shown to be d own regulated upon encephalomyocarditis (EMCV) infection, we stably tr ansfected HeLa cells with a RLI antisense cDNA expressing vector. Four independant clones named VAS1, VAS2, VAS3 and VAS4 and one clone tran sfected with the empty vector (VV) as control, were analyzed. The leve l of RLI was decreased by 20% for VAS1, 25% for VAS2, 75% for VAS3 and 50% for VAS4. The inactivation of RNase L observed during EMCV infect ion was decreased in these clones as compared to control HeLa cells. H ere again the results vary between the four clones. The maximum inhibi tion of RNase L (90%) was observed in control cells and in VAS1 while 48% inhibition was observed in VAS3 and 25 % in VAS3. The reversal in RNase L inhibition thus reflects closely the resulting RLI level, in k eeping with a major role of RLI in EMCV-induced down regulation of 2-5 A-binding activity of RNase L. Moreover, cells expressing a low level of RLI (VAS3 and VAS 4) are partially resistant to EMCV infection.