Phosphosugar stress in Bacillus subtilis: Intracellular accumulation of mannose 6-phosphate derepresed the glcR-phoC operon from repression by GlcR

Bacillus subtilis phosphorylates sugars during or after their transport into the cell.Perturbation in the conversion of intracellular phosphosugars to the central carbon metabolitesand accumulation of phosphosugars can impose stress on the cells. In this study, we investigated the effect of phosphosugar stress on B. subtilis. Preliminary experiments indicated that the non-matabolizable analogs of glucose were unable to impose stress on B. subtilis. In contrast, deletion of manA encoding mannose 6-phosphate isomerase (responsible for conversion of mannose 6-phosphate to fructose 6-phosphate) resulted in growth arrest and bulged cell shape in the medium containing mannose. Besides, an operon encoding a repressor (GlcR) and a haloic acid dehalogenase (HAD)-like phosphatase (PhoC; previously YwpJ) were upregulated. Integration of the PglcR-lacZ cassette into different mutational backgrounds indicated that PglcR is induced when (i) a manA-deficient strain is cultured with mannose or (ii) when glcR is deleted.GlcR represses the transcription of glcR-phoC bybinding to the A-type core elements of PglcR. Electrophoretic mobility shift assay showed no interaction between mannose 6-phosphate (or other phosphosugars) and the GlcR-PglcR DNA complex. PhoCwas an acid phosphatase mainly able to dephosphorylate glycerol 3-phosphate and ribose 5- phosphate. Mannose 6-phosphatewas only weakly dephosphorylated by PhoC. Since deletion of glcR and phoCalone or in combination had no effect on the cell duringphosphosugar stress,it is assumed that the derepression of glcR-phoC is a side effect of phosphosugar stress in B. subtilis. We selected two Bacillus Subtilis Strain KMO and KM283. Cells were grwon in triplicate on different sugars and total RNA was isolated followd by cDNA preparation as described in protocol section, and subsequently Miroarray Array data and analysis was done. Pleaes note that the values from multiple probes were combined and normalized further using TMM (Trimmed Media Mean) and provided in the Series supplementary file, Count.txt. For downstream analysis the Locus_tags (e.g. BSU00010, BSU00020 etc.) of Bacillus_subtilis_subsp_subtilis_str_168 (ASM904v1) are used, derived from GEO platform ORF column.