Thesis

Characterization of eleven genes putatively associated with akinete development in Nostoc punctiforme

The cyanobacterium, Nostoc punctiforme is capable of differentiating into various cell types that allow adaptation and survival under various environmental threats. It has proven to be a unique model system for investigating genetic regulation leading to cyanobacterial cell differentiation. Heterocysts are a nitrogen fixing cells that form every 15-20 cells within a filament in response to a deficiency in combined nitrogen. Akinetes are another morphologically distinct cell type that differentiate from vegetative cells under low-light intensities and limiting phosphate or potassium. They enable survival of environmental extremes such as cold or desiccation and are thought to allow persistence of the species from season to season. A previous DNA microarray experiment identified a set of eleven genes with homology to known transcriptional regulators that exhibited 1.5 to 2-fold increase in expression during akinete formation. The aim of this project was to verify the microarray results using transcriptional reporter strains. The pSUN119 transcriptional GFP reporter plasmid was fused with promoters of each gene of interest and electroporated into N. punctiforme. Under akinete inducing conditions, 8 out of 11 reporter strains showed GFP fluorescence under epifluorescence microscopy indicating transcriptional up-regulation during akinesis. Of the remaining 3 genes, two showed increased expression in heterocysts while the third had no upregulation in either cell types. Additionally, three mutant strains were created by gene deletion. So far the phenotype of one mutant has been identified; ΔNpF2889 is deficient in heterocyst formation and is unable to fix N2.

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