Translational Control of the SigR-Directed Oxidative Stress Response in Streptomyces via IF3-Mediated Repression of a Noncanonical GTC Start Codon

• Published in: mBio 2017-06, Vol.8 (3)
• Main Authors: Feeney, Morgan A, Chandra, Govind, Findlay, Kim C, Paget, Mark S B, Buttner, Mark J
• Format: Article
• Language: English
• Subjects: Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Codon, Initiator; Gene Expression Regulation, Bacterial; Operon; Oxidative Stress; Prokaryotic Initiation Factor-3 - genetics; Prokaryotic Initiation Factor-3 - metabolism; Regulon; Sigma Factor - chemistry; Sigma Factor - metabolism; Streptomyces - genetics; Streptomyces - physiology; Transcription Factors - chemistry; Transcription Factors - genetics; Transcription Factors - metabolism
• ISSN: 2161-2129; 2150-7511
• DOI: 10.1128/mBio.00815-17

The major oxidative stress response in is controlled by the sigma factor SigR and its cognate antisigma factor RsrA, and SigR activity is tightly controlled through multiple mechanisms at both the transcriptional and posttranslational levels. Here we show that has a highly unusual GTC start codon and that this leads to another level of SigR regulation, in which SigR translation is repressed by translation initiation factor 3 (IF3). Changing the GTC to a canonical start codon causes SigR to be overproduced relative to RsrA, resulting in unregulated and constitutive expression of the SigR regulon. Similarly, introducing IF3* mutations that impair its ability to repress SigR translation has the same effect. Thus, the noncanonical GTC start codon and its repression by IF3 are critical for the correct and proper functioning of the oxidative stress regulatory system. and are cotranscribed and translationally coupled, and it had therefore been assumed that SigR and RsrA are produced in stoichiometric amounts. Here we show that RsrA can be transcribed and translated independently of SigR, present evidence that RsrA is normally produced in excess of SigR, and describe the factors that determine SigR-RsrA stoichiometry. In all sigma factor-antisigma factor regulatory switches, the relative abundance of the two proteins is critical to the proper functioning of the system. Many sigma-antisigma operons are cotranscribed and translationally coupled, leading to a generic assumption that the sigma and antisigma factors are produced in a fixed 1:1 ratio. In the case of - , we show instead that the antisigma factor is produced in excess over the sigma factor, providing a buffer to prevent spurious release of sigma activity. This excess arises in part because has an extremely rare noncanonical GTC start codon, and as a result, SigR translation initiation is repressed by IF3. This finding highlights the potential significance of noncanonical start codons, very few of which have been characterized experimentally. It also emphasizes the limitations of predicting start codons using bioinformatic approaches, which rely heavily on the assumption that ATG, GTG, and TTG are the only permissible start codons.