MAPPING OF MULTIPLE RNA-BINDING SITES OF HUMAN T-CELL LYMPHOTROPIC VIRUS TYPE-I REX PROTEIN WITHIN 5'- AND 3'-REX RESPONSE ELEMENTS

Interaction between the human T-cell lymphotropic virus type I Rex pro tein and viral transcripts in the nucleus is essential to the cytoplas mic appearance of unspliced and singly spliced viral RNA. Rex has been shown to mediate its function through direct interaction with a highl y ordered secondary structure in the 3'-untranslated region of all hum an T-cell lymphotropic virus type I mRNAs termed the Rex response elem ent (3'-RxRE). Part of the 3'-RxRE sequence is also present in the 5'- end of viral transcripts (5'-RxRE), and we demonstrate that Rex binds to this RNA with essentially the same affinity and specificity as to t he 3'-RxRE. We have analyzed the secondary structures and binding site s of Rex within the 5'- and 5'-RxREs by enzymatic probing and chemical modification interference and show that multiple Rex molecules bind w ithin a stem-loop, which is similarly structured in the two RxREs. Our experiments confirm the presence of a previously characterized Rex bi nding site but also identify a common motif within an extended region that comprises an additional Rex binding site. This suggests that Rex oligomerizes on the RxREs similarly to what has been observed for bind ing of the human immunodeficiency virus type 1 Rev protein to the Rev response element.