An Extracytoplasmic Function Sigma Factor Controls Cellulose Utilization by Regulating the Expression of an Outer Membrane Protein in Cytophaga hutchinsonii

Abstract

The common soil cellulolytic bacterium known as Cytophaga hutchinsonii makes use of a unique but poorly understood strategy in order to utilize cellulose. While several genes have been identified as being an active part of the utilization of cellulose, the mechanism(s) by which C. hutchinsonii both (i) senses its environment and (ii) regulates the expression of those genes are not as yet known. In this study, we identified and characterized the gene CHU_3097 encoding an extracytoplasmic function (ECF) σ factor (σ^cel1), the disruption of which compromised C. hutchinsonii cellulose assimilation to a large degree. The σ^cel1 and its putative partner anti-σ^cel1, encoded by the CHU_3096 gene found immediately downstream from CHU_3097, copurified in vitro The σ^cel1 was discovered to be associated with inner membrane when cells were cultured on glucose and yet was partially released from the membrane in response to cellulose. This release was found to occur on glucose when the anti-σ^cel1 was absent. Transcriptome analyses found a σ^cel1-regulated, cellulose-responsive gene regulon, within which an outer membrane protein encoding the gene CHU_1276, essential for cellulose utilization, was discovered to be significantly downregulated by CHU_3097 disruption. The expression of CHU_1276 almost fully restored cellulose utilization to the CHU_3097 mutant, demonstrating that CHU_1276 represents a critical regulatory target of σ^cel1 In this way, our study provided insights into the role of an ECF σ factor in coordinating the cellulolytic response of C. hutchinsoniiIMPORTANCE The common cellulolytic bacterium Cytophaga hutchinsonii uses a unique but poorly understood strategy in order to make use of cellulose. Throughout the process of cellulosic biomass breakdown, outer membrane proteins are thought to play key roles; this is evidenced by CHU_1276, which is required for the utilization of cellulose. However, the regulatory mechanism of its expression is not yet known. We found and characterized an extracytoplasmic function σ factor that is involved in coordinating the cellulolytic response of C. hutchinsonii by directly regulating the expression of CHU_1276 This study makes a contribution to our understanding of the regulatory mechanism used by C. hutchinsonii in order to adjust its genetic programs and so deal with novel environmental cues.

Keywords: Cytophaga hutchinsonii; ECF σ factor; anti-σ factor; cellulose degradation; outer membrane protein.