Control of transcription at pause and termination sites is common in bacteria. Many transcriptional pause and termination events are thought to occur in response to formation of an RNA hairpin in the nascent transcript. Some mutations in the 13-subunit of Escherichia coli RNA polymerase that confer resistance to the transcription inhibitor rifampicin also alter the response to transcriptional pause and termination signals. Here, we report isolation of termination-altering mutations that do not confer rifampicin resistance and show that such mutations occur predominantly in limited regions of the 13-subunit polypeptide. One region is between amino acid residues 500 and 575, which encompasses the locations of almost all known rifampicin-resistance mutations. Many termination-altering mutations also occur in two other regions: between amino acid residues 740 and 840 and near the carboxyl terminus of the I~-subunit (amino acid residues 1225-1342). Amino acid sequences in these three regions of the 13-subunit are conserved between prokaryotic and eukaryotic 13-subunit homologs. Several mutations that alter transcription termination in vitro affect amino acid residues that are identical in prokaryotic and eukaryotic RNA polymerase 13-subunit homologs, suggesting that they alter an important function common to multisubunit RNA polymerases. We propose that these three regions of the 13-subunit may contact the nascent RNA transcript, the RNA-DNA heteroduplex, or the DNA template in the transcription complex and that mutations in these regions alter transcription pausing and termination by affecting these contacts.

A central enigma of transcriptional regulation is how the normally efficient transcription elongation complex stops at pause and termination signals. One possibility, raised by the discovery that RNA polymerase sometimes contracts its DNA footprint, is that discontinuous movements contribute to recognizing these signals. We report that E. coli RNA polymerase responds to sequences immediately downstream and upstream from the his leader pause site by changing neither its downstream DNA contact nor its upstream RNA contact for 8 bp preceding the pause. This compressed complex isomerizes to a paused conformation by an approximately 10 bp jump of its downstream DNA contact and simultaneous extrusion of an RNA hairpin that stabilizes the paused conformation. We suggest pausing and termination could be alternative outcomes of a similar isomerization that depend on the strength of contacts to 3'-proximal RNA remaining after the jump.

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