Structural basis of transcription antitermination by Q{lambda}: NusA induces refolding of Q{lambda} to form nozzle for RNA polymerase exit channel

Lambdoid bacteriophage Q proteins are transcription antipausing and antitermination factors that enable RNA polymerase (RNAP) to read through pause and termination sites. Q proteins load onto RNAP engaged in promoter-proximal pausing at a Q binding element (QBE) and adjacent sigma-dependent pause element to yield a Q-loading complex, and translocate with RNAP as a pausing-deficient, termination-deficient Q-loaded complex. In previous work, we showed that the Q protein from bacteriophage 21 (Q21) functions by forming a "nozzle" that narrows and extends the RNAP RNA-exit channel, preventing formation of pause and termination RNA hairpins. Here, we report atomic structures of four states on the pathway of antitermination by the Q protein from bacteriophage {lambda} (Q{lambda}), a Q protein that shows no sequence similarity to Q21 and that, unlike Q21, requires the transcription elongation factor NusA for efficient antipausing and antitermination. We report structures of Q{lambda}, the Q{lambda}-QBE complex, the NusA-free "pre-engaged" Q{lambda}-loading complex, and the NusA-containing "engaged" Q{lambda}-loading complex. The results show that Q{lambda}, like Q21, forms a nozzle that narrows and extends the RNAP RNA-exit channel, preventing formation of RNA hairpins. However, the results show that Q{lambda} has no three-dimensional structural similarity to Q21, employs a different mechanism of QBE recognition than Q21, and employs a more complex process for loading onto RNAP than Q21, involving recruitment of Q{lambda} to form a "pre-engaged" loading complex, followed by NusA-facilitated refolding of Q{lambda} to form an "engaged" loading complex. The results establish Q{lambda} and Q21 are not structural homologs and are solely functional analogs. SIGNIFICANCE STATEMENTBacteriophage Q proteins are textbook examples of regulators of gene expression that function at the level of transcription antitermination. Here, we report structures defining the mechanism of antitermination by the Q protein of bacteriophage {lambda} (Q{lambda}). The results show Q{lambda} forms a "nozzle" that narrows and extends the RNA polymerase RNA-exit channel, precluding the formation of terminator RNA hairpins. The results show Q{lambda} exhibits no structural similarity to the Q protein of bacteriophage 21 (Q21), employs a different mechanism for DNA binding than Q21, and employs a more complex process of loading onto RNA polymerase than Q21. We conclude Q{lambda} and Q21 are not structural homologs and are solely functional analogs, akin to a bird wing and a bat wing.