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J. Biol. Chem., Vol. 276, Issue 41, 38201-38209, October 12, 2001

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The Anti-initial Transcribed Sequence, a Portable Sequence That Impedes Promoter Escape, Requires ⁷⁰ for Function^*

Cathleen L. Chan and Carol A. Gross^§

From the Departments of Stomatology and of Microbiology and Immunology, University of California, San Francisco, California 94143

The anti-sequence, a portable element extending from +1 to +15 of the transcript, is sufficient to prevent promoter escape from a variety of strong ⁷⁰ promoters. We show here that this sequence does not function with even the strongest ³² promoter. Moreover, a particular class of substitutions in ⁷⁰ that disrupt interaction between Region 2.2 of ⁷⁰ and a coiled-coiled motif in the '-subunit of RNA polymerase antagonizes the function of the anti-element. This same group of mutants prevents Q-mediated anti-termination at the P_R' promoter. At this promoter, interaction of ⁷⁰ with the non-template strand of the initial transcribed sequence (ITS) is required to promote the pause prerequisite for anti-termination. These mutants prevent pausing because they are defective in this recognition event. By analogy, we suggest that interaction of ⁷⁰ with the non-template strand of the anti-ITS is required for function of this portable element, thus explaining why neither ³² nor the Region 2.2 ⁷⁰ mutants mediate anti-function. Support for the analogy with the P_R' promoter comes from preliminary experiments suggesting that the anti-ITS, like the P_R' ITS, is bipartite.

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