Clostridium thermocellum is a gram positive, anaerobic,
thermophilic soil bacterium that secretes a high molecular weight protein
complex, the cellulosome, which is capable of hydrolyzing crystalline
cellulose. C. thermocellum attracts much interest because of its ability
to both hydrolyze cellulose and to produce ethanol and thus can be part of a consolidated
bio-processing for bioethanol production. Recently, we identified a set of σ
and anti- σ factors that play a role in the regulation of the cellulosomal
genes in C. thermocellum. These alternative σ-factors are located upstream
to a gene encoding to a trans-membrane protein (RsgI) with an intracellular anti
- σ domain at its N-terminus and an extracellular polysaccharide-related
function module at its C-terminus. This
arrangement provides a novel regulatory mechanism in which the expression of
the cellulosomal genes can be controlled by the composition of the extracellular
polysaccharides. In this study, this regulatory mechanism was directly
challenged by deleting one of the rsgI genes, rsgI6, and the
transcription level of the suspected genes was measured by real – time RT PCR.
In total, 4 genes, including sigI6, exhibited
increased transcript level in the rsgI6 deletion mutant compared to its
parental strain. The results from this analysis enabled us to determine the
consensus promoter sequence recognized by the alternative σ factor, SigI6. The rsgI6 – deleted strain also exhibited
higher activity on two synthetic substrates: para-nitrophenyl-
β-D-xylopyranoside (pNPX) and para-nitrophenyl- β-D-xylobioside (pNPX2),
consistent with the up regulation of the SigI6-regulated genes.
