Computational prediction of Pho regulons in cyanobacteria

<p>Abstract</p> <p>Background</p> <p>Phosphorus is an essential element for all life forms. However, it is limiting in most ecological environments where cyanobacteria inhabit. Elucidation of the phosphorus assimilation pathways in cyanobacteria will further our understanding of the physiology and ecology of this important group of microorganisms. However, a systematic study of the Pho regulon, the core of the phosphorus assimilation pathway in a cyanobacterium, is hitherto lacking.</p> <p>Results</p> <p>We have predicted and analyzed the Pho regulons in 19 sequenced cyanobacterial genomes using a highly effective scanning algorithm that we have previously developed. Our results show that different cyanobacterial species/ecotypes may encode diverse sets of genes responsible for the utilization of various sources of phosphorus, ranging from inorganic phosphate, phosphodiester, to phosphonates. Unlike in <it>E. coli</it>, some cyanobacterial genes that are directly involved in phosphorus assimilation seem to not be under the regulation of the regulator SphR (orthologue of PhoB in <it>E coli</it>.) in some species/ecotypes. On the other hand, SphR binding sites are found for genes known to play important roles in other biological processes. These genes might serve as bridging points to coordinate the phosphorus assimilation and other biological processes. More interestingly, in three cyanobacterial genomes where no <it>sphR </it>gene is encoded, our results show that there is virtually no functional SphR binding site, suggesting that transcription regulators probably play an important role in retaining their binding sites.</p> <p>Conclusion</p> <p>The Pho regulons in cyanobacteria are highly diversified to accommodate to their respective living environments. The phosphorus assimilation pathways in cyanobacteria are probably tightly coupled to a number of other important biological processes. The loss of a regulator may lead to the rapid loss of its binding sites in a genome.</p

Blueprint for a minimal photoautotrophic cell: conserved and variable genes in Synechococcus elongatus PCC 7942

Background: Simpler biological systems should be easier to understand and to engineer towards pre-defined goals. One way to achieve biological simplicity is through genome minimization. Here we looked for genomic islands in the fresh water cyanobacteria Synechococcus elongatus PCC 7942 (genome size 2.7 Mb) that could be used as targets for deletion. We also looked for conserved genes that might be essential for cell survival.Results: By using a combination of methods we identified 170 xenologs, 136 ORFans and 1401 core genes in the genome of S. elongatus PCC 7942. These represent 6.5%, 5.2% and 53.6% of the annotated genes respectively. We considered that genes in genomic islands could be found if they showed a combination of: a) unusual G+C content; b) unusual phylogenetic similarity; and/or c) a small number of the highly iterated palindrome 1 (HIP1) motif plus an unusual codon usage. The origin of the largest genomic island by horizontal gene transfer (HGT) could be corroborated by lack of coverage among metagenomic sequences from a fresh water microbialite. Evidence is also presented that xenologous genes tend to cluster in operons. Interestingly, most genes coding for proteins with a diguanylate cyclase domain are predicted to be xenologs, suggesting a role for horizontal gene transfer in the evolution of Synechococcus sensory systems.Conclusions: Our estimates of genomic islands in PCC 7942 are larger than those predicted by other published methods like SIGI-HMM. Our results set a guide to non-essential genes in S. elongatus PCC 7942 indicating a path towards the engineering of a model photoautotrophic bacterial cell.Financial support was provided by grants BFU2009-12895-C02-01/BMC (Ministerio de Ciencia e Innovación, Spain), the European Community’s Seventh Framework Programme (FP7/2007-2013) under grant agreement number 212894 and Prometeo/2009/092 (Conselleria d’Educació, Generalitat Valenciana, Spain) to A. Moya. Work in the FdlC laboratory was supported by grants BFU2008-00995/BMC (Spanish Ministry of Education), RD06/0008/1012 (RETICS research network, Instituto de Salud Carlos III, Spanish Ministry of Health) and LSHM-CT- 2005_019023 (European VI Framework Program). Dr. González-Domenech was supported by grant from the University of Granada. LD, thanks to financial support from Facultad de Ciencias, Universidad Nacional Autónoma de México

Blueprint for a minimal photoautotrophic cell: conserved and variable genes in Synechococcus elongatus PCC 7942

This is an Open Access article distributed under the terms of the Creative Commons Attribution License.-- et al.Abstract Background Simpler biological systems should be easier to understand and to engineer towards pre-defined goals. One way to achieve biological simplicity is through genome minimization. Here we looked for genomic islands in the fresh water cyanobacteria Synechococcus elongatus PCC 7942 (genome size 2.7 Mb) that could be used as targets for deletion. We also looked for conserved genes that might be essential for cell survival. Results By using a combination of methods we identified 170 xenologs, 136 ORFans and 1401 core genes in the genome of S. elongatus PCC 7942. These represent 6.5%, 5.2% and 53.6% of the annotated genes respectively. We considered that genes in genomic islands could be found if they showed a combination of: a) unusual G+C content; b) unusual phylogenetic similarity; and/or c) a small number of the highly iterated palindrome 1 (HIP1) motif plus an unusual codon usage. The origin of the largest genomic island by horizontal gene transfer (HGT) could be corroborated by lack of coverage among metagenomic sequences from a fresh water microbialite. Evidence is also presented that xenologous genes tend to cluster in operons. Interestingly, most genes coding for proteins with a diguanylate cyclase domain are predicted to be xenologs, suggesting a role for horizontal gene transfer in the evolution of Synechococcus sensory systems. Conclusions Our estimates of genomic islands in PCC 7942 are larger than those predicted by other published methods like SIGI-HMM. Our results set a guide to non-essential genes in S. elongatus PCC 7942 indicating a path towards the engineering of a model photoautotrophic bacterial cell.Financial support was provided by grants BFU2009-12895-C02-01/BMC (Ministerio de Ciencia e Innovación, Spain), the European Community’s Seventh Framework Programme (FP7/2007-2013) under grant agreement number 212894 and Prometeo/2009/092 (Conselleria d’Educació, Generalitat Valenciana, Spain) to A. Moya. Work in the FdlC laboratory was supported by grants BFU2008-00995/BMC (Spanish Ministry of Education), RD06/0008/1012 (RETICS research network, Instituto de Salud Carlos III, Spanish Ministry of Health) and LSHM-CT-2005_019023 (European VI Framework Program). Dr. González-Domenech was supported by grant from the University of Granada. LD, thanks to financial support from Facultad de Ciencias, Universidad Nacional Autónoma de México.Peer Reviewe

Biocomputational prediction of non-coding RNAs in model cyanobacteria

<p>Abstract</p> <p>Background</p> <p>In bacteria, non-coding RNAs (ncRNA) are crucial regulators of gene expression, controlling various stress responses, virulence, and motility. Previous work revealed a relatively high number of ncRNAs in some marine cyanobacteria. However, for efficient genetic and biochemical analysis it would be desirable to identify a set of ncRNA candidate genes in model cyanobacteria that are easy to manipulate and for which extended mutant, transcriptomic and proteomic data sets are available.</p> <p>Results</p> <p>Here we have used comparative genome analysis for the biocomputational prediction of ncRNA genes and other sequence/structure-conserved elements in intergenic regions of the three unicellular model cyanobacteria <it>Synechocystis </it>PCC6803, <it>Synechococcus elongatus </it>PCC6301 and <it>Thermosynechococcus elongatus </it>BP1 plus the toxic <it>Microcystis aeruginosa </it>NIES843. The unfiltered numbers of predicted elements in these strains is 383, 168, 168, and 809, respectively, combined into 443 sequence clusters, whereas the numbers of individual elements with high support are 94, 56, 64, and 406, respectively. Removing also transposon-associated repeats, finally 78, 53, 42 and 168 sequences, respectively, are left belonging to 109 different clusters in the data set. Experimental analysis of selected ncRNA candidates in <it>Synechocystis </it>PCC6803 validated new ncRNAs originating from the <it>fabF-hoxH </it>and <it>apcC-prmA </it>intergenic spacers and three highly expressed ncRNAs belonging to the Yfr2 family of ncRNAs. Yfr2a promoter-<it>luxAB </it>fusions confirmed a very strong activity of this promoter and indicated a stimulation of expression if the cultures were exposed to elevated light intensities.</p> <p>Conclusion</p> <p>Comparison to entries in Rfam and experimental testing of selected ncRNA candidates in <it>Synechocystis </it>PCC6803 indicate a high reliability of the current prediction, despite some contamination by the high number of repetitive sequences in some of these species. In particular, we identified in the four species altogether 8 new ncRNA homologs belonging to the Yfr2 family of ncRNAs. Modelling of RNA secondary structures indicated two conserved single-stranded sequence motifs that might be involved in RNA-protein interactions or in the recognition of target RNAs. Since our analysis has been restricted to find ncRNA candidates with a reasonable high degree of conservation among these four cyanobacteria, there might be many more, requiring direct experimental approaches for their identification.</p

EVOLUTION AND DYNAMICS OF TRANSCRIPTIONAL REGULATION IN BACTERIA

Although transcription is one of the most important biological functions of cells, our understanding of its regulation is still limited. In this dissertation, we have studied the transcriptional regulation in prokaryotes in three aspects. First, we investigated the extent to which cis-regulatory elements are conserved during the course of evolution using the LexA regulons in cyanobacteria as an example. We found that in most cyanobacterial genomes analyzed, LexA appears to function as the transcriptional regulator of the key SOS response genes. The loss of lexA in some genomes might lead to the degradation of its binding sites. Second, directional RNA-seq techniques have recently become the workhorse for transcriptome profiling in prokaryotes, however, it is a challenging task to accurately assemble highly labile prokaryotic transcriptomes for further analyses. To fill this gap, we have developed a hidden Markov model based transcriptome assembler which outperforms the state-of-the-art assemblers. Using our tool, we characterized alternative operon structures in E. coli K12 under various growth conditions and growth phases, and found that they are more complex and dynamic than previously anticipated. Lastly, we determined anti-sense and non-coding transcription patterns in E. coli K12 under various growth conditions and time points. We found that a large portion of genes have antisense transcription in a condition-dependent manner. Most antisense transcripts are initiated and restricted to the 5?-end of the gene on the sense strand, and their expression levels are correlated with those of the genes on the sense strand, suggesting that these antisense transcripts might play an important role in transcriptional regulation

Computational analysis of LexA regulons in Cyanobacteria

<p>Abstract</p> <p>Background</p> <p>The transcription factor LexA plays an important role in the SOS response in <it>Escherichia coli </it>and many other bacterial species studied. Although the <it>lexA </it>gene is encoded in almost every bacterial group with a wide range of evolutionary distances, its precise functions in each group/species are largely unknown. More recently, it has been shown that <it>lexA </it>genes in two cyanobacterial genomes <it>Nostoc sp</it>. PCC 7120 and <it>Synechocystis sp</it>. PCC 6803 might have distinct functions other than the regulation of the SOS response. To gain a general understanding of the functions of LexA and its evolution in cyanobacteria, we conducted the current study.</p> <p>Results</p> <p>Our analysis indicates that six of 33 sequenced cyanobacterial genomes do not harbor a <it>lexA </it>gene although they all encode the key SOS response genes, suggesting that LexA is not an indispensable transcription factor in these cyanobacteria, and that their SOS responses might be regulated by different mechanisms. Our phylogenetic analysis suggests that <it>lexA </it>was lost during the course of evolution in these six cyanobacterial genomes. For the 26 cyanobacterial genomes that encode a <it>lexA </it>gene, we have predicted their LexA-binding sites and regulons using an efficient binding site/regulon prediction algorithm that we developed previously. Our results show that LexA in most of these 26 genomes might still function as the transcriptional regulator of the SOS response genes as seen in <it>E. coli </it>and other organisms. Interestingly, putative LexA-binding sites were also found in some genomes for some key genes involved in a variety of other biological processes including photosynthesis, drug resistance, etc., suggesting that there is crosstalk between the SOS response and these biological processes. In particular, LexA in both <it>Synechocystis sp. </it>PCC6803 and <it>Gloeobacter violaceus </it>PCC7421 has largely diverged from those in other cyanobacteria in the sequence level. It is likely that LexA is no longer a regulator of the SOS response in <it>Synechocystis sp</it>. PCC6803.</p> <p>Conclusions</p> <p>In most cyanobacterial genomes that we analyzed, LexA appears to function as the transcriptional regulator of the key SOS response genes. There are possible couplings between the SOS response and other biological processes. In some cyanobacteria, LexA has adapted distinct functions, and might no longer be a regulator of the SOS response system. In some other cyanobacteria, <it>lexA </it>appears to have been lost during the course of evolution. The loss of <it>lexA </it>in these genomes might lead to the degradation of its binding sites.</p

Computational prediction of the osmoregulation network in Synechococcus sp. WH8102

<p>Abstract</p> <p>Background</p> <p>Osmotic stress is caused by sudden changes in the impermeable solute concentration around a cell, which induces instantaneous water flow in or out of the cell to balance the concentration. Very little is known about the detailed response mechanism to osmotic stress in marine <it>Synechococcus</it>, one of the major oxygenic phototrophic cyanobacterial genera that contribute greatly to the global CO₂fixation.</p> <p>Results</p> <p>We present here a computational study of the osmoregulation network in response to hyperosmotic stress of <it>Synechococcus sp </it>strain <it>WH8102 </it>using comparative genome analyses and computational prediction. In this study, we identified the key transporters, synthetases, signal sensor proteins and transcriptional regulator proteins, and found experimentally that of these proteins, 15 genes showed significantly changed expression levels under a mild hyperosmotic stress.</p> <p>Conclusions</p> <p>From the predicted network model, we have made a number of interesting observations about <it>WH8102</it>. Specifically, we found that (i) the organism likely uses glycine betaine as the major osmolyte, and others such as glucosylglycerol, glucosylglycerate, trehalose, sucrose and arginine as the minor osmolytes, making it efficient and adaptable to its changing environment; and (ii) σ³⁸, one of the seven types of σ factors, probably serves as a global regulator coordinating the osmoregulation network and the other relevant networks.</p

Identification of cyanobacterial non-coding RNAs by comparative genome analysis

BACKGROUND: Whole genome sequencing of marine cyanobacteria has revealed an unprecedented degree of genomic variation and streamlining. With a size of 1.66 megabase-pairs, Prochlorococcus sp. MED4 has the most compact of these genomes and it is enigmatic how the few identified regulatory proteins efficiently sustain the lifestyle of an ecologically successful marine microorganism. Small non-coding RNAs (ncRNAs) control a plethora of processes in eukaryotes as well as in bacteria; however, systematic searches for ncRNAs are still lacking for most eubacterial phyla outside the enterobacteria. RESULTS: Based on a computational prediction we show the presence of several ncRNAs (cyanobacterial functional RNA or Yfr) in several different cyanobacteria of the Prochlorococcus-Synechococcus lineage. Some ncRNA genes are present only in two or three of the four strains investigated, whereas the RNAs Yfr2 through Yfr5 are structurally highly related and are encoded by a rapidly evolving gene family as their genes exist in different copy numbers and at different sites in the four investigated genomes. One ncRNA, Yfr7, is present in at least seven other cyanobacteria. In addition, control elements for several ribosomal operons were predicted as well as riboswitches for thiamine pyrophosphate and cobalamin. CONCLUSION: This is the first genome-wide and systematic screen for ncRNAs in cyanobacteria. Several ncRNAs were both computationally predicted and their presence was biochemically verified. These RNAs may have regulatory functions and each shows a distinct phylogenetic distribution. Our approach can be applied to any group of microorganisms for which more than one total genome sequence is available for comparative analysis

The Effects of Chromium on Cellular Viability and Chromium Tolerant Gene Expression within Synechococcus sp. IU 625

Heavy metal contamination in the environment is always a big concern. Many microorganisms have developed metal tolerant /resistant mechanisms to survive in such environment. Cyanobacteria Synechococcus sp. IU 625 (S IU 625) formerly known as Anacystis nidulans have been used an indicator for studying many EPA targeted heavy metals such as Zn+2, Cu+2, Hg+2. This microorganism has been reported to have resistant mechanisms to mercury and zinc. In this study, in silico analysis was used to determine the pANL plasmid-mediated chromium resistant genes of Synechococcus sp. IU 625. DOOR and STRING were used to determine the operons and identity potential transcription factors. PCR primers were designed to amplify each gene within the proposed operon. Sequencing revealed high homology to S. elongatus PCC 7942 conservation among the Synechococcus genus. To monitor gene expression, S. IU 625 cultures were supplemented with increasing concentrations of chromium and isolated on hours 0, 8, 24, and 48. Samples were subject to RT-PCR and q-PCR to quantify the proposed chromium resistance operon and transcription regulators in the presences and absence of chromium. Results showed that all genes encoding the srpCDE operon are co-expressed in high concentrations of chromium. These also, a chromosomal transcription regulator showed increased co-expression with the operon. Furthermore, the pANL plasmid, in which the operon is located, increased in number as an adaptive response to chromium stress