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| Author | : Kathryn Marjorie Jones |
| Publisher | : |
| Total Pages | : 342 |
| Release | : 2001 |
| Genre | : Anabaena |
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| Author | : Krithika Kumar |
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| Total Pages | : |
| Release | : 2012 |
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Many multicellular cyanobacteria produce specialized nitrogenfixing heterocysts. During diazotrophic growth of Anabaena (Nostoc) sp. strain PCC 7120, a regulated developmental pattern of single heterocysts separated by about 10 to 20 photosynthetic vegetative cells is maintained along filaments. Heterocyst structure and metabolic activity function together to accommodate oxygensensitive nitrogen fixation, catalyzed by nitrogenase. In this work, we show that the promoter of the nifHDK genes that encode nitrogenase, lies upstream from the intergenic region between nifH and nifU. Excision of the fdxN element is required for transcription of the nifHDK genes. Fluorescence microscopy of reporter strain PnifHD-gfp, in the chromosomal nif locus indicated that expression of nifHDK is blocked in mutants that are unable to excise the fdxN element after nitrogen deprivation. We proposed that a promoter upstream of the element, likely PnifB, is required for transcription of the nifHDK genes. Indeed, the PnifHD-gfp reporter at an ectopic site did not show GFP fluorescence. A PnifB-gfp reporter was expressed specifically in heterocysts indicating that a promoter for the nifB gene lies in the intergenic region upstream of nifB. A stem loop structure located in the intergenic region between nifH and nifU may act as a processing site for production of nifHDK transcripts. We also provide evidence that DevH, a transcriptional regulator, is involved in regulating the nifBfdxNnifSUHDK genes. DevH is a protein belonging to the cAMP receptor protein (CRP) family of proteins that are widespread in bacteria and regulate genes in response to a gamut of physiological conditions. We show that DevH binds specifically to the nifB upstream region but not to the immediate upstream region of nifH. We predict that DevH binds to an NtcAlike binding site upstream of nifB and functions as an activator of the nifB-fdxN-nifSUHDK genes. Finally, we show that sigE, which is expressed at 16 hours after nitrogen deprivation, is required for normal expression of some heterocyst specific genes, including nifHDK. A sigE mutant shows delayed and reduced expression of nifHDK and some middle and late genes. We hypothesize that DevH in concert with SigE upregulates the expression of nifHDK in heterocysts after nitrogen deprivation.
| Author | : Pritty B. Borthakur |
| Publisher | : |
| Total Pages | : 194 |
| Release | : 2008 |
| Genre | : Anabaena |
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The goal of this research was to understand regulation of heterocyst differentiation in Anabaena sp. strain PCC 7120 by characterizing regulatory genes for heterocyst formation and their mutants. Anabaena is a filamentous cyanobacterium that forms specialized cells for nitrogen fixation, called heterocysts, which differentiate from vegetative cells at intervals of 10--12 cells. Two genes, patS and hetN, are known to suppress the differentiation of vegetative cells into heterocysts for establishing a de novo pattern and maintaining a pattern of heterocysts along the filament. A mutant, UHM100, was created to study the function of both genes by deleting patS and making expression of hetN conditional. This study has established that PatS and HetN are members of two separate heterocyst suppression pathways. In absence of nitrogen, inactivation of both patS and hetN increases heterocyst differentiation to nearly 100%, giving rise to a phenotype called 'multiple contiguous heterocysts' (Mch). UHM100 has an Mch phenotype even in the presence of combined nitrogen, which usually suppresses heterocyst differentiation. In absence of both patS and hetN, the expression of hetR, a master regulator of heterocyst differentiation, was observed in ~55% cells and was asynchronous. The distribution of heterocysts next to a vegetative cell in UHM 100 was found to be nonrandom. These results suggest that besides PatS and HetN, there are other factors that influence pattern formation in Anabaena PCC 7120. A heterocyst-deficient (Hef) spontaneous mutant, NSM6, was isolated from UHM 100. A novel gene, alr9018, from the Anabaena Epsilon plasmid complemented NSM6 and restored the Mch phenotype of this mutant. Transconjugants of Anabaena PCC 7120 containing the cloned alr9018 gene fixed 50% more N2 than PCC 7120, suggesting that multiple copies of alr9018 enhance heterocyst development. This is the first report showing that the Epsilon plasmid of Anabaena PCC 7120 contains genes involved in heterocyst differentiation. Expression of alr9018 was observed in both vegetative cells and heterocysts. Similar to alr9018, hetR could also restore the Mch phenotype in NSM6, suggesting functional similarity between a1r9018 and hetR. The Alr9018 protein contains an NTPase domain, which is a characteristic of proteins involved in signal transduction.
| Author | : Britt Lee Flaherty |
| Publisher | : |
| Total Pages | : 125 |
| Release | : 2012 |
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| ISBN | : 9781267832580 |
Fixed nitrogen is a limited resource for growth in the environment and the fixation of atmospheric nitrogen is vital to nutrient cycling and growth. Cyanobacteria are a group of photosynthetic bacteria that evolved 4.5 billion years ago to harvest sunlight as energy. Cyanobacteria have since evolved a wide array of metabolic capabilities over time, including the ability to fix atmospheric nitrogen. Anabaena sp. strain PCC 7120, hereafter Anabaena, is a species of cyanobacteria that fixes atmospheric N2 into ammonia by forming specialized nitrogen-fixing cells called heterocysts. Heterocysts form only in the absence of a source of fixed nitrogen and are evenly spaced along a filament of Anabaena cells. We studied the gene expression networks that regulate heterocyst development through deep sequencing, employing both RNA-seq on a nitrogen-deprived culture as well as ChIP-seq on key transcription factors involved in heterocyst development. Deep sequencing gave us a global view of gene expression in response to nitrogen deprivation in Anabaena. Our RNA-seq work identified new genes involved in heterocyst development, mapped operon structure and transcript length, and discovered abundant antisense transcription in the genome. In particular, we identified antisense transcription in the coding region of the gene nblA, which codes for a small peptide that triggers the proteolysis of the photosynthetic machinery in response to nutrient stress. Furthermore, we used ChIP-seq to identify the regulon of two transcription factors, HetR and DevH, in response to nitrogen deprivation. Our work on HetR, a transcription factor with known roles in regulating heterocyst development, identified many new HetR targets, including genes involved in HetR's role during nitrogen deprivation and during vegetative cell growth. Our work on DevH, a transcription factor required for forming the heterocyst-specific cell wall, also identified new DevH targets, including many genes involved in cell wall formation and transcriptional regulators. This work adds to our understanding of transcriptional networks that regulate heterocyst development in Anabaena. Furthermore, our study provides insight into gene structure and transcriptional regulation in cyanobacteria as a whole.
| Author | : Rodrigo Andres Mella Herrera |
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| Total Pages | : |
| Release | : 2010 |
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Many multicellular cyanobacteria produce specialized nitrogen-fixing heterocysts. During diazotrophic growth of the model organism Anabaena (Nostoc) sp. strain PCC 7120, a regulated developmental pattern of single heterocysts separated by about 10 to 20 photosynthetic vegetative cells is maintained along filaments. Heterocyst structure and metabolic activity function to accommodate the oxygen-sensitive process of nitrogen fixation. This dissertation focuses on my research on heterocyst development, including morphogenesis, transport of molecules between cells in a filament, differential gene expression, and pattern formation. We using microarray experiments we found that conR (all0187) gene is necessary for normal septum-formation of vegetative cells, diazotrophic grow, and heterocyst morphogenesis. In our studies we characterized the expression of sigma factors genes in Anabaena PCC 7120 during heterocyst differentiation, and we found that the expression of sigC, sigG and sigE is localized primarily in heterocysts. Expression studies using sigE mutant showed that nifH is under the control of this specific sigma factor.
| Author | : Todd Alan Black |
| Publisher | : |
| Total Pages | : 346 |
| Release | : 1993 |
| Genre | : Anabaena |
| ISBN | : |
| Author | : |
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| Total Pages | : |
| Release | : 2004 |
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Anabaena sp. strain PCC 7120 is a cyanobacterium that carries out photosynthesis in a manner similar to plants and is capable of nitrogen fixation. This organism has developed a necessary spatial separation of the incompatible processes of photosynthesis and nitrogen fixation, as nitrogen fixation is sensitive to oxygen that is produced during photosynthesis. A differentiated cell type, called a heterocyst, is formed when Anabaena is in an environment lacking nitrogen, and these cells are the sites of nitrogen fixation. Heterocyst formation occurs about every tenth cell along a filament of photosynthetic vegetative cells after 24-36 hours of nitrogen starvation. A screen for sequences up-regulated at the transcript level during heterocyst development in Anabaena identified adjacent loci alr4311 and all4312. The sequence of alr4311 suggests it encodes the ATP-binding protein of an ABC transporter complex, while that of all4312 suggests it encodes the response regulator of a two-component regulatory system. Phylogenetic analysis of the predicted protein sequences of alr4311 and all4312 indicated that both of these proteins have orthologs in Nostoc punctiforme and Anabaena variabilis, two filamentous, diazotrophic cyanobacteria. Additionally, alr4311 appears to be most similar to ABC transporters involved in the import of cobalt, while all4312 was most similar to uncharacterized response regulators. The transcripts of alr4311 and all4312 are expressed at low levels in vegetative cells, and increase in abundance after nitrogen starvation and the induction of heterocyst development. Northern analysis and real-time RT-PCR showed that expression of alr4311 and all4312 are induced as early as 3 hours after initiation of differentiation, and expression levels of both genes remain elevated through the first 24 hours of development. Expression of both of these genes was blocked in an ntcA mutant, and significantly decreased in a hetR mutant. alr4311 was shown to be part of an.
| Author | : Yuping Cai |
| Publisher | : |
| Total Pages | : 372 |
| Release | : 1991 |
| Genre | : Anabaena |
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| Author | : Rebecca Elizabeth Thayer |
| Publisher | : |
| Total Pages | : 72 |
| Release | : 2006 |
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Keywords: all4312, alr4311, Anabaena, cyanobacteria, heterocyst development.
| Author | : Maria Ramona Aldea |
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| Total Pages | : |
| Release | : 2010 |
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The regulatory mechanisms that govern heterocyst development in Anabaena sp. strain PCC 7120 have been continuously refined over the last two decades. In this work, we show that three of the sigma factor genes present in the Anabaena sp. strain PCC 7120 genome are developmentally regulated. Time-lapse microscopy of gfp reporter strains indicated that expression of sigC, sigG, and sigE is upregulated specifically in differentiating cells at 4 h, 9 h and 16 h, respectively, after induction of heterocyst development. We proposed that the sigma factors encoded by these genes are involved in regulation of heterocyst-specific genes whose expression is relatively coincident with that of sigC, sigG, or sigE. Indeed, inactivation of the sigC gene caused delayed and reduced expression of genes required for the early stages of heterocyst development, and caused delayed development. Inactivation of the sigE gene caused a considerable drop in expression of nifH, a late gene required for nitrogen fixation. We also provide evidence that c-di-GMP, a novel bacterial second messenger, is involved in regulating heterocyst development. The all2874 gene encodes a bona fide diguanylate cyclase, which synthesizes c-di-GMP, and the gene's inactivation resulted in a decreased tendency to form heterocysts; this phenotype was exacerbated by high light intensity. We hypothesize that the putative operon all2875-all2874 senses and relays information about light conditions and this information is integrated into the decision to form heterocysts. Finally, we identified the all0187 gene, which is expressed at 9 h, a time when cells that have initiated differentiation commit to complete the process. In nitrogen-free medium, all0187 mutant filaments formed abnormally long heterocysts and were unable to grow diazotrophically. Septum formation between heterocysts and their flanking vegetative cells was incomplete, leaving one or both poles of the heterocysts more opened and potentially more permeable to oxygen. Despite having nitrogenase activity, the all0187 mutant was unable to grow diazotrophically. We hypothesize that the diazotrophic growth defect is caused by the inability of the heterocysts to transport fixed nitrogen to the neighboring vegetative cells.
