On the nature of 5' termini in nuclear pre-mRNA of Ehrlich carcinoma cells.

Abstract 5′ terminal nucleosides of nuclear pre-mRNA of Ehrlich ascites carcinoma cells were analyzed by a combination of different chromatographic methods and phosphatase treatment. The heavy nuclear pre-mRNA contains mainly unblocked triphosphorylated nucleosides at the 5′ end, although some capped 5′ ends could also be found. In this respect, it differs from cytoplasmic poly(A) + mRNA which contains blocked 5′ termini and no triphosphorylated ends. The 5′ terminal nucleotides in pre-mRNA are pppGp and pppAp (in a ratio of 3:2). The determination of pppNp content in poly(A) + , poly(U) + , and poly(A) − (U) − fragments of RNA has been used as an approach to establish the topography of pre-mRNA. We also established that the technique for isolation of triphosphorylated 5′ terminal fragments of RNA based on hydroxyapatite chromatography (Bajszar, Samarina, and Georgiev, 1974) is still valid in the presence of blocked oligonucleotides. The latter do not interfere with fragments containing free triphosphate groups. Using this technique, we showed that a small but significant portion of triphosphorylated 5′ end fragments of ∼100 nucleotides in length contain oligo(U) sequences reacting with poly(A)-Sepharose.