ROLE OF ESCHERICHIA-COLI RNA-POLYMERASE ALPHA-SUBUNIT IN MODULATION OF PAUSING, TERMINATION AND ANTI-TERMINATION BY THE TRANSCRIPTION ELONGATION-FACTOR NUSA

The alpha subunit (alpha) of RNA polymerase (RNAP) is critical for ass embly of polymerase and positive control of transcription initiation i n Escherichia coil, Here, we report that alpha also plays a role in tr anscription elongation, and this involves a direct interaction between alpha and NusA factor. During in vitro transcription without NusA, al pha interacts with the nascent RNA, as revealed by photocrosslinking, When NusA is present, RNA crosslinks to NusA rather than to alpha. We show that this NusA-RNA interaction is diminished during transcription with an RNAP mutant that lacks the C-terminus of alpha beyond amino a cid 235, including the so-called alpha CTD, The absence of alpha CTD a lso affects NusA's ability to enhance transcription pausing, terminati on at intrinsic terminators and anti-termination by the phage lambda Q anti-terminator, but not antitermination by the lambda N anti-termina tor. NusA functions are not recovered even when transcription with mut ant RNAP is done with excess NusA, a condition which does restore NusA -RNA crosslinking, By affinity chromatography, we show that NusA inter acts directly with alpha, and also with beta and beta', but not with m utant alpha. Hence, alpha-NusA interaction is vital for the control of transcript elongation and termination.