Variation in the size of nascent RNA cleavage products as a function of transcript length and elongation competence.
RNA polymerase II arrested at specific template locations can be rescued by elongation factor SII via RNA cleavage. The size of the products removed from the 3'-end of the RNA varies. The release of single nucleotides, dinucleotides, and larger oligonucleotides has been detected by different workers. Dinucleotides tend to originate from SII-independent complexes and 7-14 base products from SII-dependent complexes (Izban, M. G., and Luse, D. S. (1993) J. Biol. Chem. 268, 12874-12885). Different modes of cleavage have also been recognized for bacterial transcription complexes and are thought to represent important structural differences between functionally distinct transcription intermediates. Using an elongation complex "walking" technique, we have observed factor-independent complexes as they approach and become arrested at an arrest site. Dinucleotides or 7-9-base (large) oligonucleotides were released from SII-independent or dependent complexes, respectively. The abrupt shift between the release of dinucleotide versus larger products accompanied the change from factor-dependent to factor-independent elongation, as described by others. However, not all factor-independent complexes showed cleavage in dinucleotide intervals since oligonucleotides 2-6 bases long were also liberated from elongation-competent complexes. These were all 5'-coterminal oligonucleotides indicating that a preferred phosphodiester bond is targeted for cleavage in a series of related complexes. This is consistent with recent models postulating a large product binding site that can hold RNA chains whose size increases as a function of chain polymerization. A specific transitional complex was identified that acquired the ability to cleave in a large increment one base insertion event prior to attaining the arrested configuration.
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Graduate Program in Biochemistry and Molecular Biology, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
Reines
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D000818:Animals; D001483:Base Sequence; D008099:Liver / Q000378:metabolism; D008969:Molecular Sequence Data; D009843:Oligoribonucleotides / Q000032:analysis* / Q000138:chemical synthesis / Q000378:metabolism; D012313:RNA / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D012319:RNA Polymerase II / Q000378:metabolism*; D051381:Rats; D012265:Ribonucleotides / Q000378:metabolism; D013698:Templates, Genetic; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic
D009843:Oligoribonucleotides; D012265:Ribonucleotides; D014157:Transcription Factors; D012313:RNA; D012319:RNA Polymerase II