Genome phylogeny based on gene content.

Species phylogenies derived from comparisons of single genes are rarely consistent with each other, due to horizontal gene transfer, unrecognized paralogy and highly variable rates of evolution. The advent of completely sequenced genomes allows the construction of a phylogeny that is less sensitive to such inconsistencies and more representative of whole-genomes than are single-gene trees. Here, we present a distance-based phylogeny constructed on the basis of gene content, rather than on sequence identity, of 13 completely sequenced genomes of unicellular species. The similarity between two species is defined as the number of genes that they have in common divided by their total number of genes. In this type of phylogenetic analysis, evolutionary distance can be interpreted in terms of evolutionary events such as the acquisition and loss of genes, whereas the underlying properties (the gene content) can be interpreted in terms of function. As such, it takes a position intermediate to phylogenies based on single genes and phylogenies based on phenotypic characteristics. Although our comprehensive genome phylogeny is independent of phylogenies based on the level of sequence identity of individual genes, it correlates with the standard reference of prokarytic phylogeny based on sequence similarity of 16s rRNA. Thus, shared gene content between genomes is quantitatively determined by phylogeny, rather than by phenotype, and horizontal gene transfer has only a limited role in determining the gene content of genomes.

Gene context conservation of a higher order than operons.

Operons, co-transcribed and co-regulated contiguous sets of genes, are poorly conserved over short periods of evolutionary time. The gene order, gene content and regulatory mechanisms of operons can be very different, even in closely related species. Here, we present several lines of evidence which suggest that, although an operon and its individual genes and regulatory structures are rearranged when comparing the genomes of different species, this rearrangement is a conservative process. Genomic rearrangements invariably maintain individual genes in very specific functional and regulatory contexts. We call this conserved context an uber-operon.

Conservation of gene order: a fingerprint of proteins that physically interact.

A systematic comparison of nine bacterial and archaeal genomes reveals a low level of gene-order (and operon architecture) conservation. Nevertheless, a number of gene pairs are conserved. The proteins encoded by conserved gene pairs appear to interact physically. This observation can therefore be used to predict functions of, and interactions between, prokaryotic gene products.

Assessment of phylogenomic and orthology approaches for phylogenetic inference.

MOTIVATION: Phylogenomics integrates the vast amount of phylogenetic information contained in complete genome sequences, and is rapidly becoming the standard for reliably inferring species phylogenies. There are, however, fundamental differences between the ways in which phylogenomic approaches like gene content, superalignment, superdistance and supertree integrate the phylogenetic information from separate orthologous groups. Furthermore, they all depend on the method by which the orthologous groups are initially determined. Here, we systematically compare these four phylogenomic approaches, in parallel with three approaches for large-scale orthology determination: pairwise orthology, cluster orthology and tree-based orthology. RESULTS: Including various phylogenetic methods, we apply a total of 54 fully automated phylogenomic procedures to the fungi, the eukaryotic clade with the largest number of sequenced genomes, for which we retrieved a golden standard phylogeny from the literature. Phylogenomic trees based on gene content show, relative to the other methods, a bias in the tree topology that parallels convergence in lifestyle among the species compared, indicating convergence in gene content. CONCLUSIONS: Complete genomes are no guarantee for good or even consistent phylogenies. However, the large amounts of data in genomes enable us to carefully select the data most suitable for phylogenomic inference. In terms of performance, the superalignment approach, combined with restrictive orthology, is the most successful in recovering a fungal phylogeny that agrees with current taxonomic views, and allows us to obtain a high-resolution phylogeny. We provide solid support for what has grown to be a common practice in phylogenomics during its advance in recent years. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

[Examples of preimplantation genetic diagnosis versus prenatal diagnosis in carriers of genetic abnormalities: advantages and disadvantages]. / Voorbeelden van pre-implantatiegenetische diagnostiek versus prenatale diagnostiek bij dragerschap van genetische afwijkingen: voor- en nadelen.

Preimplantation genetic diagnosis (PGD) is a method for identifying genetic abnormalities in embryos obtained via in-vitro fertilisation before their implantation. There are many indications for PGD, which offers an alternative to prenatal diagnosis, although each modality has its advantages and drawbacks. In three female patients there were clear indications for PGD: carriage of the gene for myotubular myopathy, a balanced complex chromosomal translocation, and a Robertsonian translocation in the male partner, respectively. The first patient eventually abandoned PGD and chose prenatal diagnosis. She had a total of three abortions. A twin pregnancy with a so-called foetal reduction resulted in the birth of a genetically normal child. The second patient had one spontaneous abortion and subsequently underwent two PGD cycles, resulting in an uncomplicated pregnancy and the delivery of a genetically normal child. The third patient had one genetically normal child and six spontaneous abortions. She then underwent the PGD procedure successfully and is expecting twins; the pregnancy has been uncomplicated. During preconception counselling of couples at high genetic risk, physicians should be aware of PGD as an alternative to prenatal diagnosis.

Inversions and the dynamics of eukaryotic gene order.

Comparisons of the gene order in closely related genomes reveal a major role for inversions in the genome shuffling process. In contrast to prokaryotes, where the inversions are predominantly large, half of the inversions between Saccharomyces cerevisiae and Candida albicans appear to be small, often encompassing only a single gene. Overall the genome rearrangement rate appears higher in eukaryotes than in prokaryotes, and the current genome data do not indicate that functional constraints on the co-expression of neighboring genes have a large role in conserving eukaryotic gene order. Nevertheless, qualitatively interesting examples of conservation of gene order in eukaryotes can be observed.

Exploitation of gene context.

Recently, a number of techniques have been proposed that use completely sequenced genomes for the function prediction of individual proteins encoded therein. They use the fusion of genes, their conserved location in operons or merely their co-occurrence in genomes to predict the existence of functional interactions between the proteins they encode. This type of information complements functional features that are predicted by classical homology-based search techniques.

Predicting disease genes using protein-protein interactions.

BACKGROUND: The responsible genes have not yet been identified for many genetically mapped disease loci. Physically interacting proteins tend to be involved in the same cellular process, and mutations in their genes may lead to similar disease phenotypes. OBJECTIVE: To investigate whether protein-protein interactions can predict genes for genetically heterogeneous diseases. METHODS: 72,940 protein-protein interactions between 10,894 human proteins were used to search 432 loci for candidate disease genes representing 383 genetically heterogeneous hereditary diseases. For each disease, the protein interaction partners of its known causative genes were compared with the disease associated loci lacking identified causative genes. Interaction partners located within such loci were considered candidate disease gene predictions. Prediction accuracy was tested using a benchmark set of known disease genes. RESULTS: Almost 300 candidate disease gene predictions were made. Some of these have since been confirmed. On average, 10% or more are expected to be genuine disease genes, representing a 10-fold enrichment compared with positional information only. Examples of interesting candidates are AKAP6 for arrythmogenic right ventricular dysplasia 3 and SYN3 for familial partial epilepsy with variable foci. CONCLUSIONS: Exploiting protein-protein interactions can greatly increase the likelihood of finding positional candidate disease genes. When applied on a large scale they can lead to novel candidate gene predictions.

STRING: a web-server to retrieve and display the repeatedly occurring neighbourhood of a gene.

The repeated occurrence of genes in each other's neighbourhood on genomes has been shown to indicate a functional association between the proteins they encode. Here we introduce STRING (search tool for recurring instances of neighbouring genes), a tool to retrieve and display the genes a query gene repeatedly occurs with in clusters on the genome. The tool performs iterative searches and visualises the results in their genomic context. By finding the genomically associated genes for a query, it delineates a set of potentially functionally associated genes. The usefulness of STRING is illustrated with an example that suggests a functional context for an RNA methylase with unknown specificity.

Re-annotating the Mycoplasma pneumoniae genome sequence: adding value, function and reading frames.

Four years after the original sequence submission, we have re-annotated the genome of Mycoplasma pneumoniae to incorporate novel data. The total number of ORFss has been increased from 677 to 688 (10 new proteins were predicted in intergenic regions, two further were newly identified by mass spectrometry and one protein ORF was dismissed) and the number of RNAs from 39 to 42 genes. For 19 of the now 35 tRNAs and for six other functional RNAs the exact genome positions were re-annotated and two new tRNA(Leu) and a small 200 nt RNA were identified. Sixteen protein reading frames were extended and eight shortened. For each ORF a consistent annotation vocabulary has been introduced. Annotation reasoning, annotation categories and comparisons to other published data on M.pneumoniae functional assignments are given. Experimental evidence includes 2-dimensional gel electrophoresis in combination with mass spectrometry as well as gene expression data from this study. Compared to the original annotation, we increased the number of proteins with predicted functional features from 349 to 458. The increase includes 36 new predictions and 73 protein assignments confirmed by the published literature. Furthermore, there are 23 reductions and 30 additions with respect to the previous annotation. mRNA expression data support transcription of 184 of the functionally unassigned reading frames.

Long-term morbidity of adjuvant whole abdominal radiotherapy (WART) or chemotherapy for early stage ovarian cancer.

UNLABELLED: The aim of the study was to evaluate long-term toxicity of adjuvant treatment in early stage ovarian cancer survivors. Data from all patients treated in one hospital for early stage ovarian cancer diagnosed between 1980 and 1990 were collected using a structured data form. In 93 FIGO stages I and II patients, cytoreductive and staging surgery was performed; 15 received no adjuvant treatment (controls), 39 whole abdominal radiotherapy (WART) and 39 platin-based chemotherapy. Median age at diagnosis was 54 years (range 21-83 years). During follow-up, 49/93 (53%) patients have died with a median overall survival of 18.4 years (95% CI 12.8-23.9). In both the radiotherapy and the chemotherapy group, 50% of patients reported long-term side-effects (all grades) versus 13% of controls. Two patients in the WART group died from bowel complications. Secondary malignancies were observed in 16 patients. Of all patients alive at the last follow-up, 12/17 (71%) patients treated with radiotherapy and 11/18 (61%) treated with chemotherapy experienced long-term morbidity versus 2/9 (22%) controls (P=0.03). IN CONCLUSION: Long-term follow-up of early stage ovarian cancer patients showed lasting GI morbidity in the survivors treated with adjuvant radiotherapy, which has therefore become obsolete. Cisplatin-based chemotherapy caused peripheral neuropathy versus virtual absence of problems in the survivors of just surgery, emphasising the need for strict criteria before instigating adjuvant treatment.

Predicting protein function by genomic context: quantitative evaluation and qualitative inferences.

Various new methods have been proposed to predict functional interactions between proteins based on the genomic context of their genes. The types of genomic context that they use are Type I: the fusion of genes; Type II: the conservation of gene-order or co-occurrence of genes in potential operons; and Type III: the co-occurrence of genes across genomes (phylogenetic profiles). Here we compare these types for their coverage, their correlations with various types of functional interaction, and their overlap with homology-based function assignment. We apply the methods to Mycoplasma genitalium, the standard benchmarking genome in computational and experimental genomics. Quantitatively, conservation of gene order is the technique with the highest coverage, applying to 37% of the genes. By combining gene order conservation with gene fusion (6%), the co-occurrence of genes in operons in absence of gene order conservation (8%), and the co-occurrence of genes across genomes (11%), significant context information can be obtained for 50% of the genes (the categories overlap). Qualitatively, we observe that the functional interactions between genes are stronger as the requirements for physical neighborhood on the genome are more stringent, while the fraction of potential false positives decreases. Moreover, only in cases in which gene order is conserved in a substantial fraction of the genomes, in this case six out of twenty-five, does a single type of functional interaction (physical interaction) clearly dominate (>80%). In other cases, complementary function information from homology searches, which is available for most of the genes with significant genomic context, is essential to predict the type of interaction. Using a combination of genomic context and homology searches, new functional features can be predicted for 10% of M. genitalium genes.

The phylogenetic distribution of frataxin indicates a role in iron-sulfur cluster protein assembly.

Much has been learned about the cellular pathology of Friedreich's ataxia, a recessive neurodegenerative disease resulting from insufficient expression of the mitochondrial protein frataxin. However, the biochemical function of frataxin has remained obscure, hampering attempts at therapeutic intervention. To predict functional interactions of frataxin with other proteins we investigated whether its gene specifically co-occurs with any other genes in sequenced genomes. In 56 available genomes we identified two genes with identical phylogenetic distributions to the frataxin/cyaY gene: hscA and hscB/JAC1. These genes have not only emerged in the same evolutionary lineage as the frataxin gene, they have also been lost at least twice with it, and they have been horizontally transferred with it in the evolution of the mitochondria. The proteins encoded by hscA and hscB, the chaperone HSP66 and the co-chaperone HSP20, have been shown to be required for the synthesis of 2Fe-2S clusters on ferredoxin in proteobacteria. JAC1, an ortholog of hscB, and SSQ1, a paralog of hscA, have been shown to be required for iron-sulfur cluster assembly in mitochondria of Saccharomyces cerevisiae. Combining data on the co-occurrence of genes in genomes with experimental and predicted cellular localization data of their proteins supports the hypothesis that frataxin is directly involved in iron-sulfur cluster protein assembly. They indicate that frataxin is specifically involved in the same sub-process as HSP20/Jac1p.

Pathway alignment: application to the comparative analysis of glycolytic enzymes.

Comparative analysis of metabolic pathways in different genomes yields important information on their evolution, on pharmacological targets and on biotechnological applications. In this study on glycolysis, three alternative ways of comparing biochemical pathways are combined: (1) analysis and comparison of biochemical data, (2) pathway analysis based on the concept of elementary modes, and (3) a comparative genome analysis of 17 completely sequenced genomes. The analysis reveals a surprising plasticity of the glycolytic pathway. Isoenzymes in different species are identified and compared; deviations from the textbook standard are detailed. Several potential pharmacological targets and by-passes (such as the Entner-Doudoroff pathway) to glycolysis are examined and compared in the different species. Archaean, bacterial and parasite specific adaptations are identified and described.