Abstract
Clostridium beijerinckii DG-8052, derived from NCIMB 8052, cannot produce solvent or form spores, a phenomenon known as degeneration. To explore the mechanisms of degeneration at the gene level, transcriptomic profiles of the wild-type 8052 and DG-8052 strains were compared. Expression of 5168 genes comprising 98.6% of the genome was assessed. Interestingly, 548 and 702 genes were significantly up-regulated in the acidogenesis and solventogenesis phases of DG-8052, respectively, and mainly responsible for the phosphotransferase system, sugar metabolic pathways, and chemotaxis; meanwhile, 699 and 797 genes were significantly down-regulated, respectively, and mainly responsible for sporulation, oxidoreduction, and solventogenesis. The functions of some altered genes, including 286 and 333 at the acidogenesis and solventogenesis phases, respectively, remain unknown. Dysregulation of the fermentation machinery was accompanied by lower transcription levels of glycolysis rate-limiting enzymes (pfk and pyk), and higher transcription of cell chemotaxis genes (cheA, cheB, cheR, cheW, and cheY), controlled mainly by σ54 at acidogenesis. Meanwhile, abnormal spore formation was associated with repressed spo0A, sigE, sigF, sigG, and sigK which are positively regulated by σ70, and correspondingly inhibited expression of CoA-transferase at the solventogenesis phase. These findings indicated that morphological and physiological changes in the degenerated Clostridium strain may be related to altered expression of sigma factors, providing valuable targets for strain development of Clostridium species.
Highlights
Solventogenic Clostridium species are unique microorganisms due to their natural ability to use a wide range of substrates as carbon sources to produce large amounts of bio-butanol, a process termed acetone-butanol-ethanol (ABE) fermentation (Lee et al, 2008)
While degeneration of Clostridium acetobutylicum ATCC 824 is caused by the loss of the mega-plasmid pSOL1 that harbors the sol operon expressing alcohol/aldehyde dehydrogenase and CoA transferase genes, responsible for acid re-assimilation (Cornillot et al, 1997); degeneration of C. saccharoperbutylacetonicum is caused by deficient formation of NADH from pyruvate (Hayashida and Yoshino, 1990)
10 up-regulated (Cbei_4123, Cbei_0411, Cbei_0311, Cbei_3120, Cbei_0441, Cbei_0331, Cbei_4824, Cbei_2740, Cbei_4871, and Cbei_3356) and 10 down-regulated (Cbei_3835, Cbei_1930, Cbei_0677, Cbei_2826, Cbei_2600, Cbei_1583, Cbei_1079, Cbei_0284, Cbei_3832, and Cbei_2261) genes were randomly selected for Quantitative Real-Time PCR (qRT-PCR)
Summary
Solventogenic Clostridium species are unique microorganisms due to their natural ability to use a wide range of substrates as carbon sources to produce large amounts of bio-butanol, a process termed acetone-butanol-ethanol (ABE) fermentation (Lee et al, 2008). ABE fermentation is a biphasic process in which solventogenic Clostridium species produce acetic and butyric acids. Transcriptome Analysis of Degenerated Clostridium beijerinckii intracellularly, and concomitantly release them into the fermentation broth during the exponential growth or acidogenic phase. This is followed by the solventogenic or stationary growth phase, when the acids are re-assimilated into cells and converted to ABE (Ezeji et al, 2010). Since Clostridium beijerinckii NCIMB 8052 has no mega-plasmid, with solventogenic genes located in the 6.7-Mbp single circular chromosomal DNA (Chen and Blaschek, 1999), this microorganism undergoes degeneration different from C. acetobutylicum ATCC 824. Studies applying proteomics and DNA microarrays have been carried out to generate industrially valuable Clostridia strains (Tomas et al, 2003; Alsaker and Papoutsakis, 2005; Wang et al, 2012, 2013; Han et al, 2013; Zhang and Ezeji, 2013); no genome wide transcriptomic analysis for strain degeneration has been reported
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