Dataset Information

Functional annotation of orthologs in metagenomes: a case study of genes for the transformation of oceanic dimethylsulfoniopropionate.

ABSTRACT: Dimethylsulfoniopropionate (DMSP) is produced mainly by phytoplankton and bacteria. It is relatively abundant and ubiquitous in the marine environment, where bacterioplankton make use of it readily as both carbon and sulfur sources. In one transformation pathway, part of the molecule becomes dimethylsulfide (DMS), which escapes into the atmosphere and plays an important role in the sulfur exchange between oceans and atmosphere. Through its other dominant catabolic pathway, bacteria are able to use it as sulfur source. During the past few years, a number of genes involved in its transformation have been characterized. Identifying genes in taxonomic groups not amenable to conventional methods of cultivation is challenging. Indeed, functional annotation of genes in environmental studies is not straightforward, considering that particular taxa are not well represented in the available sequence databases. Furthermore, many genes belong to families of paralogs with similar sequences but perhaps different functions. In this study, we develop in silico approaches to infer protein function of an environmentally important gene (dmdA) that carries out the first step in the sulfur assimilation from DMSP. The method combines a set of tools to annotate a targeted gene in genome databases and metagenome assemblies. The method will be useful to identify genes that carry out key biochemical processes in the environment.

SUBMITTER: Gonzalez JM

PROVIDER: S-EPMC6474240 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

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Functional annotation of orthologs in metagenomes: a case study of genes for the transformation of oceanic dimethylsulfoniopropionate.

González José M JM Hernández Laura L Manzano Iris I Pedrós-Alió Carlos C

The ISME journal 20190114 5

Dimethylsulfoniopropionate (DMSP) is produced mainly by phytoplankton and bacteria. It is relatively abundant and ubiquitous in the marine environment, where bacterioplankton make use of it readily as both carbon and sulfur sources. In one transformation pathway, part of the molecule becomes dimethylsulfide (DMS), which escapes into the atmosphere and plays an important role in the sulfur exchange between oceans and atmosphere. Through its other dominant catabolic pathway, bacteria are able to u ...[more]

PMID: 30643200

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Project description:BackgroundShotgun metagenomes are often assembled prior to annotation of genes which biases the functional capacity of a community towards its most abundant members. For an unbiased assessment of community function, short reads need to be mapped directly to a gene or protein database. The ability to detect genes in short read sequences is dependent on pre- and post-sequencing decisions. The objective of the current study was to determine how library size selection, read length and format, protein database, e-value threshold, and sequencing depth impact gene-centric analysis of human fecal microbiomes when using DIAMOND, an alignment tool that is up to 20,000 times faster than BLASTX.ResultsUsing metagenomes simulated from a database of experimentally verified protein sequences, we find that read length, e-value threshold, and the choice of protein database dramatically impact detection of a known target, with best performance achieved with longer reads, stricter e-value thresholds, and a custom database. Using publicly available metagenomes, we evaluated library size selection, paired end read strategy, and sequencing depth. Longer read lengths were acheivable by merging paired ends when the sequencing library was size-selected to enable overlaps. When paired ends could not be merged, a congruent strategy in which both ends are independently mapped was acceptable. Sequencing depths of 5 million merged reads minimized the error of abundance estimates of specific target genes, including an antimicrobial resistance gene.ConclusionsShotgun metagenomes of DNA extracted from human fecal samples sequenced using the Illumina platform should be size-selected to enable merging of paired end reads and should be sequenced in the PE150 format with a minimum sequencing depth of 5 million merge-able reads to enable detection of specific target genes. Expecting the merged reads to be 180-250?bp in length, the appropriate e-value threshold for DIAMOND would then need to be more strict than the default. Accurate and interpretable results for specific hypotheses will be best obtained using small databases customized for the research question.

Project description:Despite over a billion years of evolutionary divergence, several thousand human genes possess clearly identifiable orthologs in yeast, and many have undergone lineage-specific duplications in one or both lineages. These duplicated genes may have been free to diverge in function since their expansion, and it is unclear how or at what rate ancestral functions are retained or partitioned among co-orthologs between species and within gene families. Thus, in order to investigate how ancestral functions are retained or lost post-duplication, we systematically replaced hundreds of essential yeast genes with their human orthologs from gene families that have undergone lineage-specific duplications, including those with single duplications (1 yeast gene to 2 human genes, 1:2) or higher-order expansions (1:>2) in the human lineage. We observe a variable pattern of replaceability across different ortholog classes, with an obvious trend toward differential replaceability inside gene families, and rarely observe replaceability by all members of a family. We quantify the ability of various properties of the orthologs to predict replaceability, showing that in the case of 1:2 orthologs, replaceability is predicted largely by the divergence and tissue-specific expression of the human co-orthologs, i.e., the human proteins that are less diverged from their yeast counterpart and more ubiquitously expressed across human tissues more often replace their single yeast ortholog. These trends were consistent with in silico simulations demonstrating that when only one ortholog can replace its corresponding yeast equivalent, it tends to be the least diverged of the pair. Replaceability of yeast genes having more than 2 human co-orthologs was marked by retention of orthologous interactions in functional or protein networks as well as by more ancestral subcellular localization. Overall, we performed >400 human gene replaceability assays, revealing 50 new human-yeast complementation pairs, thus opening up avenues to further functionally characterize these human genes in a simplified organismal context.

Dataset Information

Functional annotation of orthologs in metagenomes: a case study of genes for the transformation of oceanic dimethylsulfoniopropionate.

Publications

Functional annotation of orthologs in metagenomes: a case study of genes for the transformation of oceanic dimethylsulfoniopropionate.

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