Search

σ factors are single subunit general transcription factors that reversibly bind core RNA polymerase and mediate gene-specific transcription in bacteria. Previously, an atypical two-subunit σ factor was identified that activates transcription from a group of related promoters in Bacillus subtilis. Both of the subunits, named SigO and RsoA, share primary sequence similarity with the canonical σ70 family of σ factors and interact with each other and with RNA polymerase subunits. Here we show that the σ70 region 2.3-like segment of RsoA is unexpectedly sufficient for interaction with the amino-terminus of SigO and the β' subunit. A mutational analysis of RsoA identified aromatic residues conserved amongst all RsoA homologues, and often amongst canonical σ factors, that are particularly important for the SigO-RsoA interaction. In a canonical σ factor, region 2.3 amino acids bind non-template strand DNA, trapping the promoter in a single-stranded state required for initiation of transcription. Accordingly, we speculate that RsoA region 2.3 protein-binding activity likely arose from a motif that, at least in its ancestral protein, participated in DNA-binding interactions.

Extracytoplasmic function (ECF) σ factors constitute a major component of the physicochemical sensory apparatus in bacteria. Most ECF σ factors are co-expressed with a negative regulator called an anti-σ factor that binds to its cognate σ factor and sequesters it from productive association with core RNA polymerase (RNAP). Anti-σ factors constitute an important element of signal transduction pathways that mediate an appropriate transcriptional response to changing environmental conditions. The Bacillus subtilis genome encodes seven canonical ECF σ factors and six of these are co-expressed with experimentally verified anti-σ factors. B. subtilis also expresses an ECF-like atypical two-subunit σ factor composed of subunits SigO and RsoA that becomes active after exposure to certain cell-wall-acting antibiotics and to growth under acidic conditions. This work describes the identification and preliminary characterization of a protein (RsiO, formerly YvrL) that constitutes the anti-σ factor cognate to SigO-RsoA. Synthesis of RsiO represses SigO-RsoA-dependent transcription initiation by binding the N-terminus of SigO under neutral (pH 7) conditions. Reconstitution of the SigO-RsoA-RsiO regulatory system into a heterologous host reveals that the imposition of acid stress (pH 5.4) abolishes the ability of RsiO to repress SigO-RsoA-dependent transcription and this correlates with loss of RsiO binding affinity for SigO. A current model for RsiO function indicates that RsiO responds, either directly or indirectly, to increased extracytoplasmic hydrogen ion concentration and becomes inactivated. This results in the release of SigO into the cytoplasm, where it productively associates with RsoA and core RNAP to initiate transcription from target promoters in the cell.

The sigma-like factor YvrI and coregulator YvrHa activate transcription from a small set of conserved promoters in Bacillus subtilis. We report here that these two proteins independently contribute sigma-region 2 and sigma-region 4 functions to a holoenzyme-promoter DNA complex. YvrI binds RNA polymerase (RNAP) through a region 4 interaction with the beta-subunit flap domain and mediates specific promoter recognition but cannot initiate DNA melting at the -10 promoter element. Conversely, YvrHa possesses sequence similarity to a conserved core-binding motif in sigma-region 2 and binds to the N-terminal coiled-coil element in the RNAP beta'-subunit previously implicated in interaction with region 2 of sigma-factors. YvrHa plays an essential role in stabilizing the open complex and interacts specifically with the N-terminus of YvrI. Based on these results, we propose that YvrHa is situated in the transcription complex proximal to the -10 element of the promoter, whereas YvrI is responsible for -35 region recognition. This system presents an unusual example of a two-subunit bacterial sigma-factor.

YvrI is a recently identified alternative sigma factor in Bacillus subtilis that requires the coactivator YvrHa to activate transcription. Previously, a strain engineered to overproduce YvrI was found to overproduce oxalate decarboxylase (OxdC), and further analysis identified three YvrI-activated promoters preceding the yvrI-yvrHa, yvrJ, and oxdC-yvrL operons. Independently, proteome analyses identified OxdC as a highly abundant, cell wall-associated protein that accumulated under acidic growth conditions. We show here that the accumulation of OxdC in the cell wall proteome under acidic growth conditions is absolutely dependent on YvrI and is correlated with enhanced transcription of both the yvrI-yvrHa and the oxdC-yvrL operons. Conversely, OxdC accumulates to a high level even under nonacidic growth conditions in cells lacking YvrL, a negative regulator of YvrI/YvrHa-dependent transcription. These results indicate that YvrI and its associated coregulators YvrHa and YvrL are required for the regulation of OxdC expression by acid stress. The high-level accumulation of OxdC depends, in part, on a strong oxdC promoter. A regulatory sequence with similarity to an upstream promoter element (UP) was identified upstream of the oxdC promoter and is required for high-level promoter activity. Conservation of the YvrI/YvrHa/YvrL regulatory system among related species allowed us to deduce an expanded consensus sequence for the compositionally unusual promoters recognized by this new sigma factor.

We have investigated the function of a cell envelope stress-inducible gene, yvrI, which encodes a 22.5 kDa protein that includes a predicted sigma(70) region 4 domain, but lacks an apparent region 2 domain. YvrI interacts with RNA polymerase and overexpression of YvrI results in induction of OxdC, an oxalate decarboxylase maximally expressed under low-pH conditions. We have used microarray-based analyses to define the YvrI regulon. YvrI is required for the transcription of three operons (oxdC-yvrL, yvrJ and yvrI-yvrHa) each of which is preceded by a highly similar promoter sequence. Activation of these promoters requires both YvrI and the product of the second gene in the yvrI-yvrHa operon, YvrHa. YvrI and YvrHa together allow recognition of the oxdC promoter, stimulate DNA melting and activate transcription by core RNA polymerase. Together, these results suggest that YvrI is a previously unrecognized sigma factor in Bacillus subtilis and that the 9.5 kDa YvrHa protein acts as a required co-activator of transcription. A yvrL deletion results in the upregulation of YvrI activity suggesting that YvrL is a negative regulator of YvrI-dependent transcription, possibly functioning as an anti-sigma factor.

Copyright of Canadian Journal of Microbiology is the property of Canadian Science Publishing and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

The σ-like factor Yvrl and coregulator YvrHa activate transcription from a small set of conserved promoters in Bacillus subtilis. We report here that these two proteins independently contribute σ-region 2 and σ-region 4 functions to a holoenzyme-promoter DNA complex. Yvrl binds RNA polymerase (RNAP) through a region 4 interaction with the β-subunit flap domain and mediates specific promoter recognition but cannot initiate DNA melting at the -10 promoter element. Conversely, YvrHa possesses sequence similarity to a conserved core-binding motif in σ-region 2 and binds to the N-terminal coiled-coil element in the RNAP β-subunit previously implicated in interaction with region 2 of σ-factors. YvrHa plays an essential role in stabilizing the open complex and interacts specifically with the N-terminus of Yvrl. Based on these results, we propose that YvrHa is situated in the transcription complex proximal to the -10 element of the promoter, whereas Yvrl is responsible for -35 region recognition. This system presents an unusual example of a two-subunit bacterial σ-factor. [ABSTRACT FROM AUTHOR]