Abstract
BackgroundSigma factors are one of the components of RNA polymerase holoenzymes, and an essential factor of transcription initiation in bacteria. Corynebacterium glutamicum possesses seven genes coding for sigma factors, most of which have been studied to some detail; however, the role of SigD in transcriptional regulation in C. glutamicum has been mostly unknown.ResultsIn this work, pleiotropic effects of sigD overexpression at the level of phenotype, transcripts, proteins and metabolites were investigated. Overexpression of sigD decreased the growth rate of C. glutamicum cultures, and induced several physiological effects such as reduced culture foaming, turbid supernatant and cell aggregation. Upon overexpression of sigD, the level of Cmt1 (corynomycolyl transferase) in the supernatant was notably enhanced, and carbohydrate-containing compounds were excreted to the supernatant. The real-time PCR analysis revealed that sigD overexpression increased the expression of genes related to corynomycolic acid synthesis (fadD2, pks), genes encoding corynomycolyl transferases (cop1, cmt1, cmt2, cmt3), L, D-transpeptidase (lppS), a subunit of the major cell wall channel (porH), and the envelope lipid regulation factor (elrF). Furthermore, overexpression of sigD resulted in trehalose dicorynomycolate accumulation in the cell envelope.ConclusionsThis study demonstrated that SigD regulates the synthesis of corynomycolate and related compounds, and expanded the knowledge of regulatory functions of sigma factors in C. glutamicum.
Highlights
Sigma factors are one of the components of RNA polymerase holoenzymes, and an essential factor of transcription initiation in bacteria
The physiological functions of SigA, SigB, SigC, SigE, SigH and SigM have been studied to some extent [11]; the regulation and physiological roles of SigD in C. glutamicum have not yet been revealed. sigD gene is well conserved among corynebacteria, and 17 out of 19 examined Corynebacterium species possess sigD genes [11]
Deletion and overexpression of sigD influenced the maximum growth rate The sigD deletion mutant (ΔsigD) and the sigD overexpressing strain were cultivated in CGXII medium containing 222 mM of glucose as carbon source. sigD was overexpressed from the plasmid pVWEx1-sigD using an Isopropyl β-D-1-thiogalactopyranoside (IPTG) inducible promoter and different IPTG concentrations (0, 10, 50, 250 or 1000 μM)
Summary
Sigma factors are one of the components of RNA polymerase holoenzymes, and an essential factor of transcription initiation in bacteria. Corynebacterium glutamicum possesses seven genes coding for sigma factors, most of which have been studied to some detail; the role of SigD in transcriptional regulation in C. glutamicum has been mostly unknown. Sigma factors are a component of bacterial RNA polymerase holoenzymes essential for promoter recognition and transcription initiation [1]. Corynebacterium glutamicum was first isolated as an organism secreting high amounts of L-glutamate [7]. C. glutamicum ATCC 13032 has seven sigma factor genes in its chromosome, sigA, sigB, sigC, sigD, sigE, sigH and sigM [9, 10]. It is assumed that SigD plays a substantial role in transcriptional regulation and subsequent adaptation of C. glutamicum to changing environments
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