RNA Folding in Transcription Elongation Complex IMPLICATION FOR TRANSCRIPTION TERMINATION

Intrinsic transcription termination signal in DNA consists of a short inverted repeat followed by a T-rich stretch. Transcription of this sequence by RNA polymerase (RNAP) results in formation of a "termination hairpin" (TH) in the nascent RNA and in rapid dissociation of the transcription elongation complex (EC) at termination points located 7-8 nt downstream of the base of TH stem. RNAP envelops 15 nt of the RNA following RNA growing 3'-end, suggesting that folding of the TH is impeded by a tight protein environment when RNAP reaches the termination points. To monitor TH folding under this constraint, we halted Escherichia coli ECs at various distances downstream from a TH and treated them with single-strand specific RNase T1. The EC interfered with TH formation when halted at 6, 7, and 8, but not 9, nt downstream from the base of the potential stem. Thus, immediately before termination, the downstream arm of the TH is protected from complementary interactions with the upstream arm. This protection makes TH folding extremely sensitive to the sequence context, because the upstream arm easily engages in competing interactions with the rest of the nascent RNA. We demonstrate that by de-synchronizing TH formation and transcription of the termination points, this subtle competition significantly affects the efficiency of transcription termination. This finding can explain previous puzzling observations that sequences far upstream of the TH or point mutations in the terminator that preserve TH stability affect termination. These results can help understand other time sensitive co-transcriptional processes in pro- and eukaryotes.

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