Long-Range RNA Interaction of Two Sequence Elements Required for Endonucleolytic Cleavage of Human Insulin-Like Growth Factor II mRNAs

Abstract

Human insulin-like growth factor II (IGF-II) mRNAs are subject to site-specific endonucleolytic cleavage in the 3′ untranslated region, leading to an unstable 5′ cleavage product containing the IGF-II coding region and a very stable 3′ cleavage product of 1.8 kb. This endonucleolytic cleavage is most probably the first and rate-limiting step in degradation of IGF-II mRNAs. Two sequence elements within the 3′ untranslated region are required for cleavage: element I, located approximately 2 kb upstream of the cleavage site, and element II, encompassing the cleavage site itself. We have identified a stable double-stranded RNA stem structure (DG 5 2100 kcal/mol [2418.4 kJ/mol]) that can be formed between element I and a region downstream of the cleavage site in element II. This structure is conserved among human, rat, and mouse mRNAs. Detailed analysis of the requirements for cleavage shows that the relative position of the elements is not essential for cleavage. Furthermore, the distance between the coding region and the cleavage site does not affect the cleavage reaction. Mutational analysis of the long-range RNA-RNA interaction shows that not only the double-stranded character but also the sequence of the stable RNA stem is important for cleavage.