Abstract
ABSTRACTA complex of highly conserved proteins consisting of NusB, NusE, NusA, and NusG is required for robust expression of rRNA in Escherichia coli. This complex is proposed to prevent Rho-dependent transcription termination by a process known as “antitermination.” The mechanism of this antitermination in rRNA is poorly understood but requires association of NusB and NusE with a specific RNA sequence in rRNA known as BoxA. Here, we identify a novel member of the rRNA antitermination machinery: the inositol monophosphatase SuhB. We show that SuhB associates with elongating RNA polymerase (RNAP) at rRNA in a NusB-dependent manner. Although we show that SuhB is required for BoxA-mediated antitermination in a reporter system, our data indicate that the major function of the NusB/E/A/G/SuhB complex is not to prevent Rho-dependent termination of rRNA but rather to promote correct rRNA maturation. This occurs through formation of a SuhB-mediated loop between NusB/E/BoxA and RNAP/NusA/G. Thus, we have reassigned the function of these proteins at rRNA and identified another key player in this complex.
Highlights
A complex of highly conserved proteins consisting of NusB, NusE, NusA, and NusG is required for robust expression of rRNA in Escherichia coli
High levels of N obviate the requirement for NusB, NusE, and BoxA [16], suggesting that the role of BoxA-NusB/E complex is to stabilize the antitermination complex [17]. In addition to their role in the life cycle of bacteriophage, Nus factors are required in many bacterial species for proper expression of rRNA [18]. rRNA loci in Escherichia coli, and many other species, contain two copies of boxA a short distance upstream of the 16S and 23S rRNA genes
The mechanism by which Nus factors prevent Rho-dependent termination is unclear, it has been suggested that a NusE-NusG interaction can prevent association of Rho with RNA polymerase (RNAP)-associated NusG, a critical requirement for Rhodependent termination [31]
Summary
A complex of highly conserved proteins consisting of NusB, NusE, NusA, and NusG is required for robust expression of rRNA in Escherichia coli. BoxA serves as a binding site for a complex of NusB and NusE [14], and BoxB serves as a binding site for N [15] These proteins, together with the RNAP-associated elongation factors NusA and NusG, modify the RNAP such that it is resistant to both Rho-dependent and intrinsic termination [5, 15]. Rho-dependent termination of a reporter construct is inhibited by insertion of sequence from the leader region of rRNA [19] This antitermination activity has been localized to a DNA segment containing a boxA sequence [20] and requires functional NusB and NusG [21]. The mechanism by which Nus factors prevent Rho-dependent termination is unclear, it has been suggested that a NusE-NusG interaction can prevent association of Rho with RNAP-associated NusG, a critical requirement for Rhodependent termination [31]
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