Comparative genomics of vertebrate Fox cluster loci.

Published onlineComparative StudyJournal ArticleResearch Support, Non-U.S. Gov'tThis is the final version of the article. Available from BioMed Central via the DOI in this record.BACKGROUND: Vertebrate genomes contain numerous duplicate genes, many of which are organised into paralagous regions indicating duplication of linked groups of genes. Comparison of genomic organisation in different lineages can often allow the evolutionary history of such regions to be traced. A classic example of this is the Hox genes, where the presence of a single continuous Hox cluster in amphioxus and four vertebrate clusters has allowed the genomic evolution of this region to be established. Fox transcription factors of the C, F, L1 and Q1 classes are also organised in clusters in both amphioxus and humans. However in contrast to the Hox genes, only two clusters of paralogous Fox genes have so far been identified in the Human genome and the organisation in other vertebrates is unknown. RESULTS: To uncover the evolutionary history of the Fox clusters, we report on the comparative genomics of these loci. We demonstrate two further paralogous regions in the Human genome, and identify orthologous regions in mammalian, chicken, frog and teleost genomes, timing the duplications to before the separation of the actinopterygian and sarcopterygian lineages. An additional Fox class, FoxS, was also found to reside in this duplicated genomic region. CONCLUSION: Comparison of loci identifies the pattern of gene duplication, loss and cluster break up through multiple lineages, and suggests FoxS1 is a likely remnant of Fox cluster duplication.We thank Peter Holland and Dave Ferrier for their discussions on this project and John Mulley for supplying the Amia calva genomic DNA, and for comments on the manuscript. This work was supported by the BBSRC

Quantification of migrant hoverfly movements (Diptera: Syrphidae) on the West Coast of North America

This is the final version. Available on open access from the Royal Society via the DOI in this recordData accessibility: The datasets supporting this article have been uploaded as part of the electronic supplementary material. A full resolution version of electronic supplementary material, file S1 can be found here: https://doi.org/10.6084/m9.figshare.7584386.v1.The seasonal migration of huge numbers of hoverflies is frequently reported in Europe from mountain passes or spurs of land. The movement of such large numbers of beneficial insects is thought to provide significant ecosystem services in terms of pollination and pest control. Observations from the East Coast of the USA during the 1920s indicate the presence of migratory life histories among some hoverfly species there, but 90 years have now passed since the last reported observation of hoverfly migration in the USA. Here, we analyse video footage taken during a huge northward migration of hoverflies on 20 April 2017 on the West Coast of California. The quantification of migrant numbers from this footage allows us to estimate the passage of over 100 000 hoverflies in half an hour over a 200 m section of headland in Montaña de Oro State Park (San Luis Obispo County). Field collections and analysis of citizen science data indicate different species from the previously reported Eristalis tenax migrations on the East Coast of the USA and provide evidence for migration among North American hoverflies. We wish to raise awareness of this phenomenon and suggest approaches to advance the study of hoverfly migration in North America and elsewhere.Royal SocietyEuropean Union Horizon 202

tarsal-less is expressed as a gap gene but has no gap gene phenotype in the moth midge Clogmia albipunctata

This is the final version.Available from The Royal Society via the DOI in this record.Gap genes are involved in segment determination during early development of the vinegar fly Drosophila melanogaster and other dipteran insects (flies, midges and mosquitoes). They are expressed in overlapping domains along the antero-posterior (A–P) axis of the blastoderm embryo. While gap domains cover the entire length of the A–P axis in Drosophila, there is a region in the blastoderm of the moth midge Clogmia albipunctata, which lacks canonical gap gene expression. Is a non-canonical gap gene functioning in this area? Here, we characterize tarsal-less (tal) in C. albipunctata. The homologue of tal in the flour beetle Tribolium castaneum (called milles-pattes, mlpt) is a bona fide gap gene. We find that Ca-tal is expressed in the region previously reported as lacking gap gene expression. Using RNA interference, we study the interaction of Ca-tal with gap genes. We show that Ca-tal is regulated by gap genes, but only has a very subtle effect on tailless (Ca-tll), while not affecting other gap genes at all. Moreover, cuticle phenotypes of Ca-tal depleted embryos do not show any gap phenotype. We conclude that Ca-tal is expressed and regulated like a gap gene, but does not function as a gap gene in C. albipunctata.This work was funded by the MEC-EMBL agreement for the EMBL/CRG Research Unit in Systems Biology, SGR Grant 406 from the Catalan funding agency AGAUR, and by grant BFU2009-10184 and BFU2012-33775 from the Spanish Ministerio de Economia y Competitividad (MINECO). The Centre for Genomic Regulation (CRG) acknowledges support from MINECO, ‘Centro de Excelencia Severo Ochoa 2013–2017', SEV-2012-0208

Medium-throughput processing of whole mount in situ hybridisation experiments into gene expression domains

This is the final version of the article. Available from the publisher via the DOI in this record.Understanding the function and evolution of developmental regulatory networks requires the characterisation and quantification of spatio-temporal gene expression patterns across a range of systems and species. However, most high-throughput methods to measure the dynamics of gene expression do not preserve the detailed spatial information needed in this context. For this reason, quantification methods based on image bioinformatics have become increasingly important over the past few years. Most available approaches in this field either focus on the detailed and accurate quantification of a small set of gene expression patterns, or attempt high-throughput analysis of spatial expression through binary pattern extraction and large-scale analysis of the resulting datasets. Here we present a robust, "medium-throughput" pipeline to process in situ hybridisation patterns from embryos of different species of flies. It bridges the gap between high-resolution, and high-throughput image processing methods, enabling us to quantify graded expression patterns along the antero-posterior axis of the embryo in an efficient and straightforward manner. Our method is based on a robust enzymatic (colorimetric) in situ hybridisation protocol and rapid data acquisition through wide-field microscopy. Data processing consists of image segmentation, profile extraction, and determination of expression domain boundary positions using a spline approximation. It results in sets of measured boundaries sorted by gene and developmental time point, which are analysed in terms of expression variability or spatio-temporal dynamics. Our method yields integrated time series of spatial gene expression, which can be used to reverse-engineer developmental gene regulatory networks across species. It is easily adaptable to other processes and species, enabling the in silico reconstitution of gene regulatory networks in a wide range of developmental contexts.The laboratory of Johannes Jaeger and this study in particular was funded by the MEC-EMBL agreement for the EMBL/CRG Research Unit in Systems Biology, by grant 153 (MOPDEV) of the ERANet: ComplexityNET program, by SGR grant 406 from the Catalan funding agency AGAUR, by grant BFU2009-10184 from the Spanish Ministry of Science, and by European Commission grant FP7-KBBE-2011-5/289434 (BioPreDyn)

A staging scheme for the development of the moth midge Clogmia albipunctata.

Published onlineJournal ArticleResearch Support, Non-U.S. Gov'tThis is the final version of the article. Available from Public Library of Science via the DOI in this record.Model organisms, such as Drosophila melanogaster, allow us to address a wide range of biological questions with experimental rigour. However, studies in model species need to be complemented by comparative studies if we are to fully understand the functional properties and evolutionary history of developmental processes. The establishment of new model organisms is crucial for this purpose. One of the first essential steps to establish a species as an experimental model is to carefully describe its life cycle and development. The resulting staging scheme serves as a framework for molecular studies, and allows us to homologise developmental processes between species. In this paper, we have characterised the life cycle and development of an emerging non-drosophilid dipteran model system: the moth midge Clogmia albipunctata. In particular, we focus on early embryogenesis (cleavage and blastoderm cycles before gastrulation), on formation and retraction of extraembryonic tissues, and on formation of the germ line. Considering the large evolutionary distance between the two species (approximately 250 million years), we find that the development of C. albipunctata is remarkably conserved compared to D. melanogaster. On the other hand, we detect significant differences in morphology and timing affecting the development of extraembryonic tissues and the germ line. Moreover, C. albipunctata shows several heterochronic shifts, and lacks head involution and associated processes during late stages of development.The laboratory of Johannes Jaeger and this study in particular was funded by the MEC-EMBL agreement for the EMBL/CRG Research Unit in Systems Biology, by SGR grant 406 from the Catalan funding agency AGAUR, by grants BFU2009-10184 & BFU2012-33775 from the Spanish Ministry of Science (MICINN, now called MINECO), and by ERANet: ERASysBio+ grant EUI2009-04045 (MODHEART). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Non-canonical dorsoventral patterning in the moth midge Clogmia albipunctata

This is the final version of the article. Available from BioMed Central via the DOI in this record.Background Bone morphogenetic proteins (BMPs) are of central importance for dorsal–ventral (DV) axis specification. They are core components of a signalling cascade that includes the BMP ligand decapentaplegic (DPP) and its antagonist short gastrulation (SOG) in Drosophila melanogaster. These components are very ancient, with orthologs involved in DV patterning in both protostomes and deuterostomes. Despite such strong conservation, recent comparative work in insects has revealed interesting differences in the way the patterning function of the DV system is achieved in different species. Results In this paper, we characterise the expression patterns of the principal components of the BMP DV patterning system, as well as its signalling outputs and downstream targets, in the non-cyclorrhaphan moth midge Clogmia albipunctata (Diptera: Psychodidae). We previously reported ventral expression patterns of dpp in the pole regions of C. albipunctata blastoderm embryos. Strikingly, we also find ventral sog and posteriorly restricted tkv expression, as well as expanded polar activity of pMad. We use our results from gene knock-down by embryonic RNA interference to propose a mechanism of polar morphogen shuttling in C. albipunctata. We compare these results to available data from other species and discuss scenarios for the evolution of DV signalling in the holometabolan insects. Conclusions A comparison of gene expression patterns across hemipteran and holometabolan insects reveals that expression of upstream signalling factors in the DV system is very variable, while signalling output is highly conserved. This has two major implications: first, as long as ligand shuttling and other upstream regulatory mechanisms lead to an appropriately localised activation of BMP signalling at the dorsal midline, it is of less importance exactly where the upstream components of the DV system are expressed. This, in turn, explains why the early-acting components of the DV patterning system in insects exhibit extensive amounts of developmental systems drift constrained by highly conserved downstream signalling output.This work was funded by the MEC-EMBL agreement for the EMBL/CRG Research Unit in Systems Biology, SGR Grant 406, from the Catalan funding agency AGAUR and by grants BFU2009-10184 and BFU2012-33775 from the Spanish Ministerio de Economia y Competitividad (MINECO). The Centre for Genomic Regulation (CRG) acknowledges support from MINECO, “Centro de Excelencia Severo Ochoa 2013-2017”, SEV-2012-0208

Evolution and expression of BMP genes in flies.

PublishedJournal ArticleResearch Support, Non-U.S. Gov'tThis is the author accepted manuscript. The final version is available from Springer Verlag via the DOI in this record.Bone morphogenetic proteins (BMPs) play key roles in development. In Drosophila melanogaster, there are three BMP-encoding genes: decapentaplegic (dpp), glass bottom boat (gbb) and screw (scw). dpp and gbb are found in all groups of insects. In contrast, the origin of scw via duplication of an ancestral gbb homologue is more recent, with new evidence placing it within the Diptera. Recent studies show that scw appeared basal to the Schizophora, since scw orthologues exist in aschizan cyclorrhaphan flies. In order to further localise the origin of scw, we have utilised new genomic resources for the nematoceran moth midge Clogmia albipunctata (Psychodidae). We identified the BMP subclass members dpp and gbb from an early embryonic transcriptome and show that their expression patterns in the blastoderm differ considerably from those seen in cyclorrhaphan flies. Further searches of the genome of C. albipunctata were unable to identify a scw-like gbb duplicate, but confirm the presence of dpp and gbb. Our phylogenetic analysis shows these to be clear orthologues of dpp and gbb from other non-cyclorrhaphan insects, with C. albipunctata gbb branching ancestrally to the cyclorrhaphan gbb/scw split. Furthermore, our analysis suggests that scw is absent from all Nematocera, including the Bibionomorpha. We conclude that the gbb/scw duplication occurred between the separation of the lineage leading to Brachycera and the origin of cyclorrhaphan flies 200-150 Ma ago.This research was funded by the MEC/EMBL agreement for the EMBL/CRG Research Unit in Systems Biology, by AGAUR SGR grant 406 and by Grants BFU2009-10184 and BFU2009-09168 from the Spanish Ministry of Science and Innovation (MICINN). EJG is supported by ERASysBio+ Grant P#161 (MODHEART). AAC acknowledges the contribution of an internship by the Caixa Catalunya savings bank, which first brought her into contact with the Jaeger lab. Genome and transcriptome sequences used in this study were acquired, assembled and annotated in collaboration with the Genomics and Bioinformatics Core Facilities at the CRG. We thank Brenda Gavilán for the help with maintaining fly cultures

A staging scheme for the development of the scuttle fly Megaselia abdita

This is the final version of the article. Available from the publisher via the DOI in this record.Model organisms, such as Drosophila melanogaster, provide powerful experimental tools for the study of development. However, approaches using model systems need to be complemented by comparative studies for us to gain a deeper understanding of the functional properties and evolution of developmental processes. New model organisms need to be established to enable such comparative work. The establishment of new model system requires a detailed description of its life cycle and development. The resulting staging scheme is essential for providing morphological context for molecular studies, and allows us to homologise developmental processes between species. In this paper, we provide a staging scheme and morphological characterisation of the life cycle for an emerging non-drosophilid dipteran model system: the scuttle fly Megaselia abdita. We pay particular attention to early embryogenesis (cleavage and blastoderm stages up to gastrulation), the formation and retraction of extraembryonic tissues, and the determination and formation of germ (pole) cells. Despite the large evolutionary distance between the two species (approximately 150 million years), we find that M. abdita development is remarkably similar to D. melanogaster in terms of developmental landmarks and their relative timing.Funding: The laboratory of Johannes Jaeger and this study in particular was funded by the MEC-EMBL agreement for the EMBL/CRG Research Unit in Systems Biology, by SGR grant 406 from the Catalan funding agency AGAUR, by grants BFU2009-10184 & BFU2012-33775 from the Spanish Ministry of Science (MICINN, now called MINECO), and by ERANet: ERASysBio+ grant EUI2009-04045 (MODHEART). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Efficient reverse-engineering of a developmental gene regulatory network

This is the final version of the article. Available from the publisher via the DOI in this record.Understanding the complex regulatory networks underlying development and evolution of multi-cellular organisms is a major problem in biology. Computational models can be used as tools to extract the regulatory structure and dynamics of such networks from gene expression data. This approach is called reverse engineering. It has been successfully applied to many gene networks in various biological systems. However, to reconstitute the structure and non-linear dynamics of a developmental gene network in its spatial context remains a considerable challenge. Here, we address this challenge using a case study: the gap gene network involved in segment determination during early development of Drosophila melanogaster. A major problem for reverse-engineering pattern-forming networks is the significant amount of time and effort required to acquire and quantify spatial gene expression data. We have developed a simplified data processing pipeline that considerably increases the throughput of the method, but results in data of reduced accuracy compared to those previously used for gap gene network inference. We demonstrate that we can infer the correct network structure using our reduced data set, and investigate minimal data requirements for successful reverse engineering. Our results show that timing and position of expression domain boundaries are the crucial features for determining regulatory network structure from data, while it is less important to precisely measure expression levels. Based on this, we define minimal data requirements for gap gene network inference. Our results demonstrate the feasibility of reverse-engineering with much reduced experimental effort. This enables more widespread use of the method in different developmental contexts and organisms. Such systematic application of data-driven models to real-world networks has enormous potential. Only the quantitative investigation of a large number of developmental gene regulatory networks will allow us to discover whether there are rules or regularities governing development and evolution of complex multi-cellular organisms.Funding: The laboratory of Johannes Jaeger and this study in particular was funded by the MEC-EMBL agreement for the EMBL/CRG Research Unit in Systems Biology, by Grant 153 (MOPDEV) of the ERANet: ComplexityNET program, by SGR Grant 406 from the Catalan funding agency AGAUR, by grant BFU2009-10184 from the Spanish Ministry of Science, and by European Commission grant FP7-KBBE-2011-5/289434 (BioPreDyn). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Quantitative system drift compensates for altered maternal inputs to the gap gene network of the scuttle fly Megaselia abdita.

Published onlineJournal ArticleThis is the final version of the article. Available from eLife Sciences Publications via the DOI in this record.The segmentation gene network in insects can produce equivalent phenotypic outputs despite differences in upstream regulatory inputs between species. We investigate the mechanistic basis of this phenomenon through a systems-level analysis of the gap gene network in the scuttle fly Megaselia abdita (Phoridae). It combines quantification of gene expression at high spatio-temporal resolution with systematic knock-downs by RNA interference (RNAi). Initiation and dynamics of gap gene expression differ markedly between M. abdita and Drosophila melanogaster, while the output of the system converges to equivalent patterns at the end of the blastoderm stage. Although the qualitative structure of the gap gene network is conserved, there are differences in the strength of regulatory interactions between species. We term such network rewiring 'quantitative system drift'. It provides a mechanistic explanation for the developmental hourglass model in the dipteran lineage. Quantitative system drift is likely to be a widespread mechanism for developmental evolution.Ministerio de Economía y Competitividad MEC/EMBL Agreement/ BFU2009-10184/ BFU2012-33775/ SEV-2012-0208 Agència de Gestió d'Ajuts Universitaris I de Recerca SGR Grant 406 European Commission FP7-KBBE-2011-5/289434 National Science Foundation IOS-0719445/IOS-112121