Septum-Localized Protein Required for Filament Integrity and Diazotrophy in the Heterocyst-Forming Cyanobacterium Anabaena sp. Strain PCC 7120

Abstract

Heterocysts, formed when filamentous cyanobacteria, such as Anabaena sp. strain PCC 7120, are grown in the absence of combined nitrogen, are cells that are specialized in fixing atmospheric nitrogen (N 2 ) under oxic conditions and that transfer fixed nitrogen to the vegetative cells of the filament. Anabaena sp. mutants whose sepJ gene (open reading frame alr2338 of the Anabaena sp. genome) was affected showed filament fragmentation and arrested heterocyst differentiation at an early stage. In a sepJ insertional mutant, a layer similar to a heterocyst polysaccharide layer was formed, but the heterocyst-specific glycolipids were not synthesized. The sepJ mutant did not exhibit nitrogenase activity even when assayed under anoxic conditions. In contrast to proheterocysts produced in the wild type, those produced in the sepJ mutant still divided. SepJ is a multidomain protein whose N-terminal region is predicted to be periplasmic and whose C-terminal domain resembles an export permease. Using a green fluorescent protein translationally fused to the carboxyl terminus of SepJ, we observed that in mature heterocysts and vegetative cells, the protein is localized at the intercellular septa, and when cell division starts, it is localized in a ring whose position is similar to that of a Z ring. SepJ is a novel composite protein needed for filament integrity, proper heterocyst development, and diazotrophic growth.

Requirement of Fra proteins for communication channels between cells in the filamentous nitrogen-fixing cyanobacteriumAnabaenasp. PCC 7120

“…Very few gold particles were observed in cells grown under N+ conditions (Fig. S2), even in septa where FraG-GFP had been localized by confocal microscopy (26). The specificity of the gold particles in the septum could not be confirmed in comparison with gold particles detected in the background, probably due to the low expression level of FraG.…”

“…fraG deletion prevents heterocyst differentiation and glycolipid layer formation, whereas the deletion of either fraC or fraD allows heterocyst differentiation, but the heterocysts formed show an aberrant neck and do not fix nitrogen (23,25). Using GFP tags, FraC, FraD, and FraG proteins were shown to be located in the septum between cells (23,26). FraD was further localized to the septum by immunogold labeling using an antibody raised Significance Cellular communication along the filaments of heterocystforming, nitrogen-fixing cyanobacteria has been discussed for at least 50 y but how this might be accomplished is not fully understood.…”

“…fraC encodes a 179-aa protein with three predicted transmembrane segments; fraD encodes a 343-aa protein with five predicted transmembrane segments and a coiled-coil domain; and fraG (also called sepJ) encodes a 751-aa protein predicted to have an N-terminal coiled-coil domain, an internal linker domain, and a C-terminal permease-like domain with either 10 transmembrane segments (22) or 9 or 11 transmembrane segments (26). fraG deletion prevents heterocyst differentiation and glycolipid layer formation, whereas the deletion of either fraC or fraD allows heterocyst differentiation, but the heterocysts formed show an aberrant neck and do not fix nitrogen (23,25).…”

The pknH gene restrictively expressed in heterocysts is required for diazotrophic growth in the cyanobacterium Anabaena sp. strain PCC 7120

“…Proteins constituting the cell-cell joining structure have been identified over the past few years (Flores et al, 2007;Mariscal et al, 2011;Merino-Puerto et al, 2011). Since PknH is predicted to be a cytosolic protein by the secondary structure prediction system SOSUI (Hirokawa et al, 1998), PknH itself is unlikely to constitute the cellcell joining structure.…”

“…In fragmentation mutants of Anabaena PCC 7120, heterocyst differentiation is impaired (Flores et al, 2007;Jang et al, 2007;Merino-Puerto et al, 2010;Nayar et al, 2007). Fig.…”