A terD domain-encoding gene (SCO2368) is involved in calcium homeostasis and participates in calcium regulation of a DosR-like regulon in Streptomyces coelicolor

Abstract

Although Streptomyces coelicolor is not resistant to tellurite, it possesses several TerD domain-encoding (tdd) genes of unknown function. To elucidate the function of tdd8, the transcriptomes of S. coelicolor strain M145 and of a tdd8 deletion mutant derivative (the Δtdd8 strain) were compared. Several orthologs of Mycobacterium tuberculosis genes involved in dormancy survival were upregulated in the deletion mutant at the visual onset of prodiginine production. These genes are organized in a putative redox stress response cluster comprising two large loci. A binding motif similar to the dormancy survival regulator (DosR) binding site of M. tuberculosis has been identified in the upstream sequences of most genes in these loci. A predicted role for these genes in the redox stress response is supported by the low NAD(+)/NADH ratio in the Δtdd8 strain. This S. coelicolor gene cluster was shown to be induced by hypoxia and NO stress. While the tdd8 deletion mutant (the Δtdd8 strain) was unable to maintain calcium homeostasis in a calcium-depleted medium, the addition of Ca(2+) in Δtdd8 culture medium reduced the expression of several genes of the redox stress response cluster. The results shown in this work are consistent with Tdd8 playing a significant role in calcium homeostasis and redox stress adaptation.

FIG 1

Growth curve of Streptomyces coelicolor strains in R5− medium represented as the optical density at 450 nm (O.D._{450 nm}) of subsamples diluted 10 times. The arrows indicate the visual onset of Red production.

FIG 2

Expression levels (±standard deviations [SD]) of five genes determined by RT-qPCR and calculated relative to gyrA expression (ΔC_T method), in the wild (squares) and Δtdd8 (circles) strains, as a function of the developmental stage.

FIG 3

Representation of the two putative Streptomyces coelicolor redox stress response loci. Genes upregulated in the Δtdd8 strain at the onset of Red production are presented as colored arrows. The red dots indicate the presence of a DosR-like binding motif in intergenic regions. The consensus sequence motif is shown with the size of the base representative of its degree of conservation in the 19 input sequences analyzed. PPK2, polyphosphate kinase 2; CRP, cyclic AMP receptor protein; FNR, fumarate and nitrate reduction regulator; PPDK, pyruvate phosphate dikinase.

FIG 4

Intracellular free Ca²⁺ concentration (+SD) in Streptomyces coelicolor strains determined under various growth conditions. Bacteria were cultivated in R5− medium supplemented or not supplemented with CaCl₂ (a) and R5− medium supplemented or not supplemented with the NO donor sodium nitroprusside (SNP) (b). Data with the same letter are not statistically significantly different (LSD test, P < 0.05).

FIG 5

Relative levels of expression (ΔC_T + SD, determined by RT-qPCR) of genes of the redox stress response cluster and developmental genes in Streptomyces coelicolor M145 and Δtdd8 strains in R5− (black bars) and R5+ (white bars) culture media. Values accompanied by the same letter do not statistically significantly differ (LSD test, P < 0.05).