Biased inheritance of the protein PatN frees vegetative cells to initiate patterned heterocyst differentiation.

Heterocysts, cells specialized for nitrogen fixation in certain filamentous cyanobacteria, appear singly in a nonrandom spacing pattern along the chain of vegetative cells. A two-stage, biased initiation and competitive resolution model has been proposed to explain the establishment of this spacing pattern. There is substantial evidence that competitive resolution of a subset of cells initiating differentiation occurs by interactions between a self-enhancing activator protein, HetR, and a diffusible pentapeptide inhibitor PatS-5 (RGSGR). Results presented here show that the absence of a unique membrane protein, PatN, in Nostoc punctiforme strain ATCC 29133 leads to a threefold increase in heterocyst frequency and a fourfold decrease in the vegetative cell interval between heterocysts. A PatN-GFP translational fusion shows a pattern of biased inheritance in daughter vegetative cells of ammonium-grown cultures. Inactivation of another heterocyst patterning gene, patA, is epistatic to inactivation of patN, and transcription of patA increases in a patN-deletion strain, implying that patN may function by modulating levels of patA. The presence of PatN is hypothesized to decrease the competency of a vegetative cell to initiate heterocyst differentiation, and the cellular concentration of PatN is dependent on cell division that results in cells transiently depleted of PatN. We suggest that biased inheritance of cell-fate determinants is a phylogenetic domain-spanning paradigm in the development of biological patterns.

DNA binding properties of the HrmR protein of Nostoc punctiforme responsible for transcriptional regulation of genes involved in the differentiation of hormogonia.

Nostoc punctiforme is an example of a filamentous cyanobacterium that is capable of differentiating non-growing cells that constitute gliding filaments termed hormogonia. These gliding filaments serve in short distance dispersal and as infective units in establishing a symbiosis with plants, such as the bryophyte Anthoceros punctatus. Mutants of N. punctiforme exist which show elevated levels of initial infection of A. punctatus as a consequence of repeated cycles of hormogonium differentiation. Such mutations occur within the hrmA and hrmU genes. Further characterization of the hrm locus revealed several genes with an organizational and predicted protein sequence similarity to genes of heterotrophic bacteria that are involved in hexuronic acid metabolism. Genes in the N. punctiforme locus are transcribed in response to the presence of a plant extract containing hormogonium-repressing factors. A predicted transcriptional repressor encoded in the locus, HrmR, was shown herein to be a specific DNA binding protein that regulates the transcription of its own gene and that of hrmE, a nearby gene. The ability of HrmR to bind DNA was abolished upon addition of either galacturonate or lysate from specifically induced N. punctiforme cells, implying that the in vivo HrmR binding activity is modulated via an internal compound, most likely a sugar molecule.

Establishment of a functional symbiosis between the cyanobacterium Nostoc punctiforme and the bryophyte Anthoceros punctatus requires genes involved in nitrogen control and initiation of heterocyst differentiation.

Three mutant strains (ntcA, hetR, hetF) of the cyanobacterium Nostoc punctiforme unable to differentiate heterocysts were characterized and examined for their ability to form a symbiotic association with the bryophyte Anthoceros punctatus. Previously unknown characteristics of the N. punctiforme hetR mutant include differentiation of chilling-resistant akinetes, while vegetative cells of the ntcA mutant randomly lysed, yielding short filaments, following ammonium deprivation. Strains with mutations in hetF and hetR infected A. punctatus with similar frequency to that of wild-type N. punctiforme but did not support growth of the plant partner. These results confirm that the infection of A. punctatus by hormogonia leading to the establishment of an association is physiologically uncoupled from the development of a functional diazotrophic association. They also indicate that heterocyst regulatory elements downstream from HetR and HetF are required in both free-living and symbiotic heterocyst differentiation and nitrogenase expression. A strain with a mutation in the global nitrogen regulator ntcA did not infect A. punctatus despite its ability to differentiate hormogonia at a low frequency. When complemented with one or more copies of ntcA, the mutant strain infected A. punctatus at a similar frequency as the wild-type and supported growth of the plant partner in the absence of combined nitrogen. These results established a connection between the presence of a functional copy of ntcA and the magnitude of hormogonium differentiation, and the behaviour of the formed hormogonia.