Phylum-wide analysis of genes/proteins related to the last steps of assembly and export of extracellular polymeric substances (EPS) in cyanobacteria

Abstract

Many cyanobacteria produce extracellular polymeric substances (EPS) with particular characteristics (e.g. anionic nature and presence of sulfate) that make them suitable for industrial processes such as bioremediation of heavy metals or thickening, suspending or emulsifying agents. Nevertheless, their biosynthetic pathway(s) are still largely unknown, limiting their utilization. In this work, a phylum-wide analysis of genes/proteins putatively involved in the assembly and export of EPS in cyanobacteria was performed. Our results demonstrated that most strains harbor genes encoding proteins related to the three main pathways: Wzy-, ABC transporter-, and Synthase-dependent, but often not the complete set defining one pathway. Multiple gene copies are mainly correlated to larger genomes, and the strains with reduced genomes (e.g. the clade of marine unicellular Synechococcus and Prochlorococcus), seem to have lost most of the EPS-related genes. Overall, the distribution of the different genes/proteins within the cyanobacteria phylum raises the hypothesis that cyanobacterial EPS production may not strictly follow one of the pathways previously characterized. Moreover, for the proteins involved in EPS polymerization, amino acid patterns were defined and validated constituting a novel and robust tool to identify proteins with similar functions and giving a first insight to which polymer biosynthesis they are related to.

The role of the tyrosine kinase Wzc (Sll0923) and the phosphatase Wzb (Slr0328) in the production of extracellular polymeric substances (EPS) by Synechocystis PCC 6803

“…The first fully segregated mutants obtained, namely ∆ wzy (∆ sll0737 —encoding the polymerase), ∆ wzx (∆ sll5049 —flippase), ∆ kpsM (∆ slr2107 —ABC transporter component), and ∆ kpsM ∆ wzy (∆ slr2107 ∆ sll0737 ), did not show any obvious phenotype in terms of growth, total carbohydrates, RPS, and CPS content (Supporting Information Figures S1–S5). We also confirmed by RT‐PCR that all the putative copies of wzy ( sll0737 , slr0728 , slr1515 , sll5074, and slr1074 ), wzx ( sll5049 , slr0488 , slr0896, and slr1543 ), and kpsM ( slr2107 , slr0977, and sll0564 ) (see Pereira et al, ; Kopf et al, ) were transcribed under standard laboratory conditions (Supporting Information Figure S6). Subsequently, we generated ∆ wzc (∆ sll0923 ), ∆ wzb (∆ slr0328 ), ∆ wzc ∆ wzb (∆ sll0923 ∆ slr0328 ), and wzc Trunc mutants.…”

“…It was previously suggested that Wzc is a BY‐kinase (Mijakovic et al, ; Pereira et al, ). To demonstrate that this protein functions as a kinase, we investigated its ability to hydrolyze ATP.…”

“…The mechanisms involved in EPS production seem to be relatively conserved throughout bacteria, with the polymerization, assembly, and export of the polymers usually following one of three main mechanisms, namely the Wzy‐, ABC transporter‐, or synthase‐dependent pathways (Schmid, Sieber, & Rehm, ). However, in a phylum‐wide analysis, we showed that most cyanobacteria harbor gene‐encoding proteins related to the three pathways but often not the complete set defining a single pathway (Pereira, Mota, Vieira, Vieira, & Tamagnini, ), implying a more complex scenario than that observed for other bacteria. This complexity is also evident in the physical organization of the genes, with multiple copies scattered throughout the genomes, either isolated or in small clusters (Pereira et al, ).…”

“…However, in a phylum‐wide analysis, we showed that most cyanobacteria harbor gene‐encoding proteins related to the three pathways but often not the complete set defining a single pathway (Pereira, Mota, Vieira, Vieira, & Tamagnini, ), implying a more complex scenario than that observed for other bacteria. This complexity is also evident in the physical organization of the genes, with multiple copies scattered throughout the genomes, either isolated or in small clusters (Pereira et al, ). In particular, this analysis revealed that Synechocystis sp.…”

Mutations in Novel Lipopolysaccharide Biogenesis Genes Confer Resistance to Amoebal Grazing in Synechococcus elongatus

“…Whether this difference is due to an evolutionary adaptation to marine salinity, a general lack of sequence conservation in the transmembrane domains of ABC transporters (51), or the different sugar composition of the LPS of marine species (9) is unclear. Overall, these conservation data support the hypothesis that the cyanobacteria evolved to use the Wzm-Wzt pathway for O-antigen synthesis and the Wzx-Wzy flippase mechanism for generating exopolysaccharides (52,53). This strategy is in contrast to that of enteric bacteria, where the Wzx-Wzy mechanism is the dominant pathway for both O-antigen (2) and extracellular polysaccharide (54) syntheses.…”