Figure 5.

In vitro processing of HAC1^u to HAC1ⁱ is not affected by mRTA or mutant forms. A, incubation of in vitro transcribed Cy5-UTP–labeled 600-nt HAC1^u RNA transcript with increasing amounts of IRE1 results in processing of HAC1^u into 181- and 167-nt 5′ and 3′ exons and a 252-nt intron. B, HAC1^u processing in the presence of increasing amounts of mRTA (top), R193A/R235A (middle), and G212E (bottom). HAC1^u transcript was preincubated with RTA for 30 min prior to the addition of 0.05 μg of recombinant IRE1 and then incubated for an additional 30 min.