Effects of Reduced and Enhanced Glycogen Pools on Salt-Induced Sucrose Production in a Sucrose-Secreting Strain of Synechococcus elongatus PCC 7942

Abstract

Sucrose and glycogen syntheses in cyanobacteria share the common precursor glucose-1-phosphate. It is generally assumed that lowering glycogen synthesis could drive more carbon toward sucrose synthesis that can be induced by salt stress among cyanobacteria. By using a theophylline-dependent riboswitch system, the expression of , a key gene in glycogen synthesis, was downregulated in a quantitative manner in a sucrose-secreting strain of PCC 7942. We observed that the stepwise suppression of glycogen synthesis limited rather than stimulated sucrose production in the salt-stressed cells, suggesting that glycogen could serve as a carbon pool for the synthesis of sucrose. Accordingly, we generated glycogen-overproducing strains, but the increased glycogen pool alone did not stimulate sucrose production, indicating that alternative steps limit the carbon flux toward the synthesis of sucrose. Consistent with previous studies that showed that sucrose-phosphate synthase (SPS) catalyzes the rate-limiting step in sucrose synthesis, the combination of glycogen overproduction and overexpression resulted in increased sucrose production. Our results indicate that the glycogen and sucrose pools are closely linked in PCC 7942, and we propose that enhancing the glycogen pool could be a promising strategy for the improvement of sucrose production by cyanobacteria in the presence of a strong sucrose synthesis sink. Many cyanobacteria naturally synthesize and accumulate sucrose when stressed by NaCl, which provides novel possibilities for obtaining sugar feedstock by engineering of cyanobacteria. It has been assumed that glycogen synthesis competes with sucrose synthesis for the carbon flux. However, our results showed that the suppression of glycogen synthesis decreased rather than stimulated sucrose production in a sucrose-secreting strain of PCC 7942. This result suggests that glycogen could serve as a supportive rather than a competitive carbon pool for the synthesis of sucrose, providing new insights about the relation between glycogen synthesis and sucrose synthesis in cyanobacteria. This finding is also useful to guide metabolic engineering work to optimize the production of sucrose and possibly other products by cyanobacteria.

Enhanced production of sucrose in the fast-growing cyanobacterium Synechococcus elongatus UTEX 2973

“…3C), the sucrose titer is unlikely to be further improved by deletion of the glycogen pathway. In fact, a recent study reported that glycogen synthesis may serve as a carbon pool for sucrose production in cyanobacteria 24 . By using a theophylline-induced riboswitch system, reduced expression of glgC led to decreased sucrose titer in Synechococcus 7942 24 .…”

Synthetic Biology Toolkits for Metabolic Engineering of Cyanobacteria

“…By employing the tools developed for chromosomal expression of heterologous genes, the inventory of biofuels and biochemicals produced in cyanobacteria has been greatly expanded . For instance, Atsumi et al expressed four heterologous genes in NS1 and NS2 along with an extra copy of the endogenous rbcLS in NS4 (Table ), leading to a high yield of isobutyraldehyde directly from CO 2 .…”

The Synthetic Biology Toolkit for Photosynthetic Microorganisms

“…Promoters are key synthetic biology components in cyanobacteria (Camsund and Lindblad, 2014;Stensjö et al, 2018); a large selection is available, including constitutive, inducible, and repressible promoters and riboswitches Ohbayashi et al, 2016;Videau et al, 2016;Immethun and Moon, 2018;Qiao et al, 2018;Wegelius et al, 2018).A detailed description of the promoter landscape is provided in Box 1, and promoter characteristics are summarized in Table 1 for ease of reference. In addition, ribosome binding sites (Englund et al, 2016;Thiel et al, 2018) and riboregulators, which are RNA sequences that respond to signal nucleic acids to modify expression (Abe et al, 2014b), have been characterized.…”

“…Synechocystis sp PCC 6803 (Huang et al, 2010;Huang and Lindblad, 2013;Englund et al, 2016;Ferreira et al, 2018;Liu and Pakrasi, 2018) Synechococcus sp PCC 7002 (Markley et al, 2015;Ruffing et al, 2016;Zess et al, 2016) S. elongatus PCC 7942 Qiao et al, 2018) S. elongatus UTEX 2973 Nostoc sp PCC 712mix0 (Videau et al, 2016;Wegelius et al, 2018) Vectors and Sandardized Assembly…”