EbsA is essential for both motility and biofilm formation in the filamentous cyanobacterium Nostoc punctiforme

Many cyanobacteria, both unicellular and filamentous, exhibit surface motility driven by type IV pili (T4P). While the component parts of the T4P machinery described in other prokaryotes are largely conserved in cyanobacteria, there are also several T4P proteins that appear to be unique to this phylum. One recently discovered component is EbsA, which has been characterized in two unicellular cyanobacteria. EbsA was found to form a complex with other T4P proteins and is essential for motility. Additionally, deletion of <i>ebsA</i> in one of these strains promoted the formation of biofilms. To expand the understanding of <i>ebsA</i> in cyanobacteria, its role in motility and biofilm formation were investigated in the model filamentous cyanobacterium <i>Nostoc punctiforme</i>. Expression of <i>ebsA</i> was strictly limited to hormogonia, the motile filaments of <i>N. punctiforme</i>. Deletion of <i>ebsA</i> did not affect hormogonium development but resulted in the loss of motility and the failure to accumulate surface pili or produce hormogonium polysaccharide (HPS), consistent with pervious observations in unicellular cyanobacteria. Protein-protein interaction studies indicated that EbsA directly interacts with PilB, and the localization of EbsA-GFP resembled that previously shown for both PilB and Hfq. Collectively, these results support the hypothesis that EbsA forms a complex along with PilB and Hfq that is essential for T4P extension. In contrast, rather than enhancing biofilm formation, deletion of both <i>ebsA</i> and <i>pilB</i> abolish biofilm formation in <i>N. punctiforme</i>, implying that distinct modalities for the relationship between motility, T4P function and biofilm formation may exist in different cyanobacteria.