Sequence specific cleavage of the HIV‐1 coreceptor CCR5 gene by a hammer‐head ribozyme and a DNA‐enzyme: inhibition of the coreceptor function by DNA‐enzyme

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Abstract
The chemokine receptor CCR5 is used as a major coreceptor for fusion and entry by non‐syncytia inducing macrophage tropic isolates of HIV‐1, which is mainly involved in transmission. Individuals who are homozygous for the Δ32 allele of CCR5 are usually resistant to HIV‐1 infection and continue to lead a normal healthy life. Thus this gene is dispensable and is, therefore, an attractive target in the host cell for interfering specifically with the virus‐host interaction. With the aim to develop a specific antiviral approach at the molecular level, we have synthesized a hammer‐head ribozyme and a DNA‐enzyme. Both ribozyme and DNA‐enzyme cleaved the CCR5 RNA in a sequence specific manner. This cleavage was protein independent but Mg2+ dependent. The extent of cleavage increased with increasing concentration of magnesium chloride. DNA‐enzyme was more effective in cleaving a full length (1376 bases) in vitro generated transcript of CCR5 gene. In this communication, we show that the DNA‐enzyme when introduced into a mammalian cell, results in decreased CD4‐CCR5‐gp160 mediated fusion of cell membranes. Potential applications of these trans acting molecules are discussed.