Transcriptomic and proteomic analyses of core metabolism in Clostridium termitidis CT1112 during growth on α-cellulose, xylan, cellobiose and xylose

Abstract

Background: Clostridium termitidis CT1112 is an anaerobic, Gram-positive, mesophilic, spore-forming, cellulolytic bacterium, originally isolated from the gut of a wood feeding termite Nasusitermes lujae. It has the ability to hydrolyze both cellulose and hemicellulose, and ferment the degradation products to acetate, formate, ethanol, lactate, H2, and CO2. It is therefore ges in gene and gene product expression during growth of C. termitidis on cellobiose, xylose, xylan, and α-cellulose.

Results: Correlation of transcriptome and proteome data with growth and fermentation profiles identified putative carbon-catabolism pathways in C. termitidis. The majority of the proteins associated with central metabolism were detected in high abundance. While major differences were not observed in gene and gene-product expression for enzymes associated with metabolic pathways under the different substrate conditions, xylulokinase and xylose isomerase of the pentose phosphate pathway were found to be highly up-regulated on five carbon sugars compared to hexoses. In addition, genes and gene-products associated with a variety of cellulosome and non-cellulosome associated CAZymes were found to be differentially expressed. Specifically, genes for cellulosomal enzymes and components were highly expressed on α-cellulose, while xylanases and glucosidases were up-regulated on 5 carbon sugars with respect to cellobiose. Chitinase and cellobiophosphorylases were the predominant CAZymes expressed on cellobiose. In addition to growth on xylan, the simultaneous consumption of two important lignocellulose constituents, cellobiose and xylose was also demonstrated.

Conclusion: There are little changes in core-metabolic pathways under the different carbon sources compared. The most significant differences were found to be associated with the CAZymes, as well as specific up regulation of some key components of the pentose phosphate pathway in the presence of xylose and xylan. This study has enhanced our understanding of the physiology and metabolism of C. termitidis, and provides a foundation for future studies on metabolic engineering to optimize biofuel production from natural biomass.

Keywords: Biofuel; CAZymes; Clostridium termitidis; Metabolism; Quantitative proteomics; RNAseq.

Fig. 1

Growth curves for C. termitidis cultured on 2 g/L each of α-cellulose (a), cellobiose (b), xylan (c) and xylose (d). Arrows indicate the time points sampled for proteomics and transcriptomic analysis. Error bars too small to be visible

Fig. 2

Growth curve (a) and substrate consumption of C. termitidis cultured on 1 g/L each cellobiose and xylose (b), and 1 g/L cellobiose plus 1 g/L xylose (c). Reported values are averages of three biological replicates

Fig. 3

Normalized differential values (Znet-scores) of both the transcriptome (Rnet) and the proteome (Pnet) of glycolysis reactions in C. termitidis. TPI: triosephosphate isomerase; PGK: phosphoglycerate kinase; ND: not detected; XS: xylose; CB: cellobiose; AC: α-cellulose; XN: xylan. Locus tags in red indicate high abundance in comparison to paralogs

Fig. 4

Normalized differential values (Znet-scores) of both the transcriptome (Rnet) and the proteome (Pnet) of reactions involved in the non-oxidative pentose phosphate pathway in C. termitidis. In the absence of genes encoding transaldolase, we propose an alternative route for the production of important intermediates using ATP-dependent 6 phosphofructokinase and fructose-1,6-bisphosphate aldolase. Values in red indicate Znet scores of ≥1.65 (outermost 10 %), up regulated in the corresponding substrate with respect to cellobiose and values in green indicate Z-scores of ≤ −1.65 (outermost 10 %), down regulated in the corresponding substrate with respect to cellobiose. Values in black are the innermost 90 %. Locus tags in red indicate high abundance in comparison to paralogs. ND: not detected; XS: xylose; CB: cellobiose; AC: α-cellulose; XN: xylan

Fig. 5

Normalized differential values (Znet-scores) of both the transcriptome (Rnet) and the proteome (Pnet) of reactions involved in the conversion of phosphoenol pyruvate to pyruvate in C. termitidis. Values in red indicate Znet scores of ≥1.65 (outermost 10 %), up regulated in the corresponding substrate with respect to cellobiose and values in green indicate Z-scores of ≤ −1.65 (outermost 10 %), down regulated in the corresponding substrate with respect to cellobiose. Values in black are the innermost 90 %. ND: not detected; XS: xylose; CB: cellobiose; AC: α-cellulose; XN: xylan

Fig. 6

Normalized differential values (Znet-scores) of both the transcriptome (Rnet) and the proteome (Pnet) of reactions involved in the conversion of pyruvate to end products in C. termitidis. Values in red indicate Znet scores of ≥1.65 (outermost 10 %), up regulated in the corresponding substrate with respect to cellobiose and values in green indicate Z-scores of ≤ −1.65 (outermost 10 %), down regulated in the corresponding substrate with respect to cellobiose. Values in black are the innermost 90 %. Locus tags in red indicate high abundance in comparison to paralogs. ND: not detected; XS: xylose; CB: cellobiose; AC: α-cellulose; XN: xylan