INCREASED ACCOMMODATION OF NASCENT RNA IN A PRODUCT SITE ON RNA-POLYMERASE-II DURING ARREST

RNA polymerases encounter specific DNA sites at which RNA chain elonga tion takes place in the absence of enzyme translocation in a process c alled discontinuous elongation. For RNA polymerase II, at least some o f these sequences also provoke transcriptional arrest where renewed RN A polymerization requires elongation factor SII. Recent elongation mod els suggest the occupancy of a site within RNA polymerase that accommo dates nascent RNA during discontinuous elongation. Here we have probed the extent of nascent RNA extruded from RNA polymerase II as it appro aches, encounters, and departs an arrest site. Just upstream of an arr est site, 17-19 nucleotides of the RNA 3'-end are protected from exhau stive digestion by exogenous ribonuclease probes. As RNA is elongated to the arrest site, the enzyme does not translocate and the protected RNA becomes correspondingly larger, up to 27 nucleotides in length. Af ter the enzyme passes the arrest site, the protected RNA is again the 18-nucleotide species typical of an elongation-competent complex. Thes e findings identify an extended RNA product groove in arrested RNA pol ymerase II that is probably identical to that emptied during SII-activ ated RNA cleavage, a process required for the resumption of elongation . Unlike Escherichia coli RNA polymerase at a terminator, arrested RNA polymerase II does not release its RNA but can reestablish the normal elongation mode downstream of an arrest site. Discontinuous elongatio n probably represents a structural change that precedes, but may not b e sufficient for, arrest by RNA polymerase II.