Identification of RNase L-Dependent, 3'-End-Modified, Viral Small RNAs in Sindbis Virus-Infected Mammalian Cells

Girardi, Erika; Chane-Woon-Ming, Béatrice; Messmer, Mélanie; Kaukinen, Pasi; Pfeffer, Sébastien

3= on the last ribose is needed for the periodate reaction. ␤-elimination of the oxidized RNA results in an RNA product one or two nucleotides shorter. On the contrary, the presence of modifications on the ribose of the 3= terminal residue renders the RNA insensitive to periodate oxidation and impairs the chain scission (31) . For a control for the reaction, we measured its effect on the unmodified cellular miRNA miR-16. As expected, the electrophoretic mobility of the miRNA strongly changed after treatment (Fig. 3B) . In contrast, SvsRNA-1 mobility was found to be insensitive to periodate oxidation and ␤-elimination (Fig. 3B) . This indicated the presence of a modification on the terminal ribose, most likely in the 2= position, which is perfectly compatible with our cloning protocol. Interestingly, it also seems that the intermediate RNA products are insensitive to periodate oxidation and are consequently modified (see Fig. S7 in the supplemental material). Moreover, we were also able to assess the existence of a modified 3= end for other SvsRNAs by Northern blotting (Fig. S8) .