ABSTRACT: Staphylococcus aureus is a major human pathogen that causes infection in a wide variety of sites within the human body. Its ability to adapt to the human host and to produce a successful infection requires precise orchestration of gene expression. While DNA-dependent RNA polymerase (RNAP) is generally well characterized, the roles of several small accessory subunits within the complex have yet to be fully explored. This is particularly true for the omega (? or RpoZ) subunit, which has been extensively studied in Gram-negative bacteria but largely neglected in Gram-positive counterparts. In Escherichia coli, it has been shown that ppGpp binding, and thus control of the stringent response, is facilitated by ?. Interestingly, key residues that facilitate ppGpp binding by ? are not conserved in S. aureus, and consequently, survival under starvation conditions is unaffected by rpoZ deletion. Further to this, ?-lacking strains of S. aureus display structural changes in the RNAP complex, which result from increased degradation and misfolding of the ?' subunit, alterations in ? and ? factor abundance, and a general dissociation of RNAP in the absence of ?. Through RNA sequencing analysis we detected a variety of transcriptional changes in the rpoZ-deficient strain, presumably as a response to the negative effects of ? depletion on the transcription machinery. These transcriptional changes translated to an impaired ability of the rpoZ mutant to resist stress and to fully form a biofilm. Collectively, our data underline, for the first time, the importance of ? for RNAP stability, function, and cellular physiology in S. aureus IMPORTANCE: In order for bacteria to adjust to changing environments, such as within the host, the transcriptional process must be tightly controlled. Transcription is carried out by DNA-dependent RNA polymerase (RNAP). In addition to its major subunits (?2??') a fifth, smaller subunit, ?, is present in all forms of life. Although this small subunit is well studied in eukaryotes and Gram-negative bacteria, only limited information is available for Gram-positive and pathogenic species. In this study, we investigated the structural and functional importance of ?, revealing key roles in subunit folding/stability, complex assembly, and maintenance of transcriptional integrity. Collectively, our data underline, for the first time, the importance of ? for RNAP function and cellular harmony in S. aureus.
