Surveying DNA Elements within Functional Genes of Heterocyst-Forming Cyanobacteria

Abstract

Some cyanobacteria are capable of differentiating a variety of cell types in response to environmental factors. For instance, in low nitrogen conditions, some cyanobacteria form heterocysts, which are specialized for N2 fixation. Many heterocyst-forming cyanobacteria have DNA elements interrupting key N2 fixation genes, elements that are excised during heterocyst differentiation. While the mechanism for the excision of the element has been well-studied, many questions remain regarding the introduction of the elements into the cyanobacterial lineage and whether they have been retained ever since or have been lost and reintroduced. To examine the evolutionary relationships and possible function of DNA sequences that interrupt genes of heterocyst-forming cyanobacteria, we identified and compared 101 interruption element sequences within genes from 38 heterocyst-forming cyanobacterial genomes. The interruption element lengths ranged from about 1 kb (the minimum able to encode the recombinase responsible for element excision), up to nearly 1 Mb. The recombinase gene sequences served as genetic markers that were common across the interruption elements and were used to track element evolution. Elements were found that interrupted 22 different orthologs, only five of which had been previously observed to be interrupted by an element. Most of the newly identified interrupted orthologs encode proteins that have been shown to have heterocyst-specific activity. However, the presence of interruption elements within genes with no known role in N2 fixation, as well as in three non-heterocyst-forming cyanobacteria, indicates that the processes that trigger the excision of elements may not be limited to heterocyst development or that the elements move randomly within genomes. This comprehensive analysis provides the framework to study the history and behavior of these unique sequences, and offers new insight regarding the frequency and persistence of interruption elements in heterocyst-forming cyanobacteria.

Fig 1. The coxA3 element found in the Calothrix sp. PCC 7103 genome.

An example of an element interrupting a gene (split into coxA3₁ and coxA3₂) in the vegetative genome. The complete, functional coxA3 gene is present in the heterocyst genome due to the element excision by the protein encoded by the xis gene.

Fig 2. Heterocyst-forming cyanobacteria 16S phylogeny.

Maximum likelihood phylogenetic tree of 16S rRNA from 38 heterocyst-forming cyanobacterial genomes, rooted with Trichodesmium erythraeum IMS 101. The size of the genome and the number of interruption elements within the genome are shown within parentheses. Cyanobacteria of the order Nostocales are in shaded boxes, and all other cyanobacteria belong to the order Stigonematales, with the exception of T. erythraeum IMS101. Open circles at branch connections indicate a bootstrap value of at least 75%.

Fig 3. Serine recombinase phylogeny.

Maximum likelihood phylogenetic tree of serine recombinase gene (xis) nucleotide sequences and data characterizing the element each xis gene is found on. Shaded boxes group element variants together. Open circles at branch connections indicate a bootstrap value of at least 75.

Fig 4. Tyrosine recombinase phylogeny.

Maximum likelihood phylogenetic tree of tyrosine recombinase gene (xis) nucleotide sequences and data characterizing the element each xis gene is found on. Shaded boxes group element variants together. Open circles at branch connections indicate a bootstrap value of at least 75.