The higher order folding process of the catalytic RNA derived from the self-splicing intron of Tetrahymena thermophila was monitored with the use of Fe(II)-EDTA-induced free radical chemistry. The overall tertiary structure of the RNA molecule forms cooperatively with the uptake of at least three magnesium ions. Local folding transitions display different metal ion dependencies, suggesting that the RNA tertiary structure assembles through a specific folding intermediate before the catalytic core is formed. Enzymatic activity, assayed with an RNA substrate that is complementary to the catalytic RNA active site, coincides with the cooperative structural transition. The higher order RNA foldings produced by Mg(II), Ca(II), and Sr(II) are similar; however, only the Mg(II)-stabilized RNA is catalytically active. Thus, these results directly demonstrate that divalent metal ions participate in general folding of the ribozyme tertiary structure, and further indicate a more specific involvement of Mg(II) in catalysis.

Although the murine retrovirus SL3-3 is highly leukaemogenic, in both the structure of its genome and in its properties of replication in tissue culture it closely resembles the nonleukaemogenic retrovirus Akv (refs 3, 4). An earlier investigation of the properties of recombinant SL3-3-Akv viruses localized the major determinant of leukaemogenicity outside the env gene, in a region of the viral genome that includes the gag gene and the noncoding long terminal repeat (LTR). To localize the determinant of SL3-3's leukaemogenicity more precisely we have now construced a recombinant provirus containing the LTR of SL3-3 and the coding region of Akv. The leukaemogenicity of these recombinants demonstrates that the determinant of leukaemogenicity lies within the SL3-3 LTR. Nucleotide sequencing of the LTRs of SL3-3 and Akv shows that they differ by a set of changes in the region thought to contain a transcriptional enhancer element. We suggest that enhancer region sequences are the major determinants of leukaemogenicity in these viruses.

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