DIRECT STRUCTURAL EVIDENCE FOR FORMATION OF A STEM-LOOP STRUCTURE INVOLVED IN RIBOSOMAL FRAMESHIFTING IN HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1

Programmed ribosomal frameshifting in viral messenger RNA occurs in re sponse to neighboring sequence elements consisting of: a frameshift si te, a spacer, and a downstream enhancer sequence. In human immunodefic iency virus type I (HIV-1) mRNA, this sequence element has a potential to form either a stem-loop or a pseudoknot structure. Based on many m utational studies, the stem-loop structure has been proposed for the d ownstream enhancer region of the HIV-1 mRNA. This stimulatory stem-loo p structure is separated from the shift site by a spacer of seven nucl eotides. In contrast, a recent report has proposed an alternative mode l in which the bases in the spacer sequence form a pseudoknot structur e as the downstream enhancer sequence [Du et al., Biochemistry 35 (199 6) 4187-4198.]. Using UV melting and enzymatic mapping analyses, we ha ve investigated the conformation of the sequence region involved in ri bosomal frameshifting in HIV-1. Our S-1, V-1, and T-1 endonuclease map pings, together with UV melting analysis, clearly indicate that this s equence element of the HIV-1 mRNA frameshift site forms a stem-loop st ructure, not a pseudoknot structure. This finding further supports the stem-loop structure proposed by many mutational studies for the downs tream enhancer sequence of the HIV-1 mRNA. (C) 1998 Elsevier Science B .V.