Figure 3.

 U5L and U5S arise from distinct intermediates generated by endonucleolytic cleavage. (A) The long and the short forms of U5 arise from different intermediates. Gel-purified σ, λ, and U5L species were incubated in extracts for the amount of time indicated and fractionated on a 6% acrylamide gel. Legends as in Figure 1B. (B) Time course of 3′ processing of internally labeled or 3′-labeled precursors. Legends as in Fig. 1B. In the mock incubation points (Mk), samples were incubated for 15 min at 30°C with extract, buffer, and 25 mm EDTA. The sums of σ′ (60 nucleotides) plus σ (270 nucleotides), or λ′ (90 nucleotides) plus λ (240 nucleotides) are about the same as the total length of pre-U5 (332 nucleotides), supporting the interpretation that the short unstable RNAs are the downstream products of endonucleolytic cleavages that generate the intermediates.