4%) of 2410 evaluated genes showed ≥ 2 fold changes at 43°C, among which 39 were down-regulated and 54 upregulated. More extensive changes were recorded at 48°C, since 532 (22%) transcript levels showed ≥ 2 fold changes, with 232 genes being down-regulated and 300 up-regulated. The distributions of the responding genes based on COG functional categories are shown
on Additional file 1. Since several COG functional categories included a mixture of annotated and poorly functionally characterized Vorinostat concentration genes (e.g. transcription regulators), we listed all poorly characterized genes in the general function prediction only category (see also Additional file 2). To provide buy AP26113 some indication of basal gene activities under control conditions, we also provided (Additional file 3, 4 and 2) semi-quantitative estimates of normalized signal intensities recorded at 37°C, which were subdivided into four categories (see Methods).
Indeed, the highest-intensity signals (75th to 100th percentile) were well correlated with the most abundant transcript products of S. aureus predicted to be highly expressed from codon usage [34]. They also correlated quite well with the most abundant proteins revealed by S. aureus proteomic studies [35], in particular enzymes involved in DNA, RNA and protein transcription machineries, central metabolism and BMN 673 energy production. Conversely, the lowest intensity signals (25th percentile) recorded at 37°C were contributed by transcripts from poorly expressed genes, such as amino acid biosynthetic pathways known to be repressed by the presence of amino acids in the MHB medium [35]. Contribution of specific transcriptomic heat stress-responses As expected from previous studies of
heat-shock responses in gram-positive bacteria [13, 18, 19], all components of S. aureus HrcA and CtsR regulons [13] were strongly induced by up-shifts to both 43°C and 48°C (Additional file 3). Transcript levels of the genes regulated by CtsR only (ctsR, mcsA, mcsB, clpC, clpP, clpB) increased by ca. 3–5 fold at 43°C 4-Aminobutyrate aminotransferase and ca. 3–11 fold at 48°C. We also observed increased expression of genes simultaneously regulated by HrcA and CtsR (grpE, dnaK, dnaJ, prmA, groEL, groES) at both 43°C and 48°C heat-shock. At 48°C, several HSP transcripts were detected at saturating levels by the microarray setting and thus their increased expression was likely under-estimated. To circumvent this problem and also validate the microarray-determined, heat-induced changes, we tested up-regulation of HSP transcript levels by qRT-PCR. Indeed, several gene transcripts (ctsR, mcsA, mcsB, hrcA) whose levels were saturated in the microarray scanner after up-shift to 48°C were more highly increased (ca. 6–16-fold) when assayed by qRT-PCR (Additional file 3).