Unwinding snail chirality by a massive subtractive linkage analysis (MSLA)

Lead Research Organisation: University of Edinburgh
Department Name: Sch of Biological Sciences

Abstract

Although many animals are symmetric on the outside, very many of them are inwardly asymmetric, or chiral: vertebrates (including ourselves) have one or several organs displaced to one side, nematode worms have an asymmetric nervous system, and even fruitflies, long-supposed to have perfect 'mirror image' symmetry, have an asymmetric intestine. For an animal to become asymmetric, symmetry must somehow be broken during development. This raises the interesting problem of how one side is consistently distinguished from the other, given that the side that we call 'right' is essentially arbitrary. The solution is that in the early stages of development a hypothetical, asymmetric 'F-molecule' lines up with the front/back and top/bottom planes, so creating a left-right asymmetry. Later organ asymmetry comes about because the F-molecule sends some substance(s) toward one side or the other (i.e. to the right, or, to the left). To attempt to understand how asymmetry is established, scientists have focussed their research on model organisms such as the mouse, chick and zebrafish. In these animals, it has been found that rotational beating of small hairs ('cilia') in the early embryo create a fluid movement that is asymmetric, leading to a suggestion that this is the critical symmetry-breaking step: the asymmetry of motor proteins around the cilia leads to directional fluid movement, ultimately determining the molecular and organismal asymmetry. While these findings are elegant, some recent and also much older research indicates that the symmetry-breaking event is much earlier, putting the ultimate relevance of the above research into doubt. For instance, in the nematode worm, left-right asymmetry is established by the six-cell stage, and probably earlier. In the pond snail chirality is determined by a substance that the mother deposits in the unfertilised egg. In the frog, molecular asymmetry is established by the four-cell stage, and even in zebrafish and chick, there are indications of differences prior to the cilial stage. Together, the results suggest that in many animals, including our close relatives, molecular asymmetry is established early in the development of embryos, with morphological asymmetry only becoming apparent later. We believe that snails may be a crucial model organism in coming to understand the symmetry-breaking step, because their asymmetry is established very early, yet they have been almost completely neglected in recent years. The objective of this project, therefore, is to utilise the power of new DNA sequencing technologies to directly identify the gene sequence that determines chirality in snail eggs. As one idea is that this gene is also the F-molecule, then this work will lead in the future to an understanding of the symmetry-breaking event in snails. The results will then invigorate analyses of the same or related molecules in other organisms, including vertebrates. Finally, as the methodology that we will use is an entirely new application of ultrahigh-throughput DNA sequencing, then success in this project would be a springboard towards using the same method to identify other genes with the same methodology.

Technical Summary

Although multiple lines of enquiry remain, a deep-seated theoretical problem has stoked a burning interest in understanding the symmetry-breaking event during development / how is one side of an organism consistently distinguished from the other, given that the side that is called 'right' is essentially arbitrary? In the hypothetical view of Brown and Wolpert, the solution is provided by a pre-existing asymmetric molecular reference: an asymmetric distribution is created if an 'F-molecule' aligns with anterior-posterior and dorsal-ventral axes, so transporting an effector molecule towards the left or right. We believe that snails may be a crucial model organism in coming to understand the symmetry-breaking step because their asymmetry is established very early. The objective of the project, therefore, is to use the almost exhaustive power of massively parallel DNA sequencing to directly clone the maternal determinant of chirality in snails, by a method that we term 'massive subtractive linkage analysis' (MSLA). The basic principle is that if the dextral gene product is only present in snails that are genetically dextral, and the sinistral gene product is only present in snails that are genetically sinistral, then a comparative bioinformatic analysis of DNA sequence reads can be used to rapidly discover candidate genes, finally identifying the gene with a functional assay. As one hypothesis is that the maternal determinant is the F-molecule in snails, or else a molecule that interacts with it, then this work will not only lead in the future to a precise understanding of the symmetry-breaking event, but will also likely stimulate comparative / investigative analyses of the same or related molecules in other organisms, including vertebrates. As the methodology is a new application of ultrahigh-throughput DNA sequencing, then success would be a springboard towards identifying other genes via the same methodology.
 
Title Transmissions exhibition 
Description The Blaxter lab collaborated closely wth artists-in-residence (see http://www.ascus.org.uk/ciie-micro-residency-artists-announced/) in the Centre for immunity Infection and Evolution to inspire and be part of the final exhibition "Transmissions". Mark Blaxter appears in the film work produced by Anne Milne, and the work of the lab inspired Jo hodges and Robbie Coleman to produce a piece dedicated to the lab. 
Type Of Art Artwork 
Year Produced 2014 
Impact 'Transmissions' was showcased to the general public within a group exhibition 'Parallel Perspectives' in Summerhall as part of the Edinburgh International Science Festival 2015 art programme, How The Light Gets In . This exhibition of work susequently travelled LifeSpace, Dundee, returning to Edinburgh to showcase at the Tent Gallery, Edinburgh College of Art. 
URL http://www.ascus.org.uk/ciie-micro-residency-2/
 
Description We have identified a gene that controls left-right organisation of the bodies of snails. This gene is also involved in left-right asymmetry in frogs, and thus probably humans. It is a fundamental finding that may have implications for biomedicine
Exploitation Route Understanding of left-right asymmetry may aid in development of treatments for conditions where asymmetry is malformed and pathogenic.
Sectors Healthcare,Pharmaceuticals and Medical Biotechnology

 
Description The snail chirality work was featured in BBC Scotland radio news (MB interview); BBC national news; BBC radio 4 news; The Atlantic science blog; other newspapers and blogs, and extensively on twitter. The Current Biology paper has an altmetric score of 467.
First Year Of Impact 2016
Sector Pharmaceuticals and Medical Biotechnology
Impact Types Societal

 
Title MolluscDB 
Description MolluscDB is a PartiGene database covering the transcriptomes of a number of mollusc species. 
Type Of Material Database/Collection of data 
Provided To Others? No  
Impact Mollusc DB has been used in several published works, including our own on Lymnaea stagnalis pond snails 
URL http://www.nematodes.org/NeglectedGenomes/MOLLUSCA/index.html
 
Title ngenomes 
Description ngenomes is a database for the display of genome assemblies from the Blaxter lab. It used the genomeHubs - LepBase ENSEMBL code 
Type Of Material Database/Collection of data 
Year Produced 2016 
Provided To Others? Yes  
Impact Colleagues globally have used the database for identification of target loci, exploration of relationships and download of data. 
URL http://ensembl.ngenomes.org
 
Description Marine Alliance for Science & Technology for Scotland 
Organisation National Oceanography Centre
Country United Kingdom 
Sector Academic/University 
PI Contribution Experimental support: assistance with understanding technology and experimental design; Delivery of sequencing service; Horizon scanning consultation to SAMS collaborators
Collaborator Contribution The MASTS collaboration is coordinating marine science in Scotland, and has recommended GenePool as its sequencing provider
Impact Several projects running in GenePool; participation in SAMS genomics workshop.
Start Year 2010
 
Description Blaxter group presentations and outreach 2015 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Mark Blaxter and research team communication and outreach 2015

Globodera genomics and blobtools software
25/02/2015 JHI Postgraduate Student Competition 2015 James Hutton Institute, Aberdeen, UK A tale of Two Peaks: Analysing Genomic Data from Potato Cyst Nematodes Talk
26/03/2015 JHI Cell and Molecular Sciences (CMS) seminar James Hutton Institute, Invergowrie, Dundee, UK Frustration and happiness : (De)-constructing parasite genomes Talk
16/06/2015 JHI Dundee effector consortium (DEC) meeting 2015 Birnam Arts and Conference Centre, Birnam, UK Variation within the Globodera pallida species complex: preliminary results Talk
03/09/2015 Molecular and Cellular Biology of Helminth Parasites IX Bratsera Hotel, Hydra, Greece Inter- and intra-specific analyses of the effector complement in potato cyst nematodes Poster
18/09/2015 UoE Postgraduate Poster Day University of Edinburgh, Edinburgh, UK Inter- and intra-specific analyses of the effector complement in potato cyst nematodes Poster
26/09/2015 Edinburgh University Doors Open Day University of Edinburgh, Edinburgh, UK Potato Cyst Nematodes (PCN) - Nematode parasites of potatoes Poster
30/11/2015 NextGenBug University of Edinburgh, Edinburgh, UK Blobtools: Blobology 2.0 Talk
01/12/2015 UK pollinator genomics meeting Roslin Institute, Edinburgh, UK Bees and Blobs Talk

LepBase
06/03/2015 EMARES Cambridge, UK The Bicyclus Genome Project Talk
06/03/2015 EMARES Cambridge, UK An introduction to Lepbase Talk
17/06/2015 Arthropod Genomics Manhattan, Kansas, USA Lepbase - A multi genome database for the Lepidoptera Poster
24/07/2015 10th Heliconius Meeting Gamboa, Panama Lepbase - A multi genome database for the Lepidoptera (API demonstration) Workshop
24/07/2015 10th Heliconius Meeting Panama Lepbase - A multi genome database for the Lepidoptera Poster
26/07/2015 10th Heliconius Meeting Panama Lepbase Workshop Talk
04/09/2015 Edinburgh Bioinformatics Edinburgh, UK Lepbase - A multi genome database for the Lepidoptera Talk
26/09/2015 Open Doors Day "Make a butterfly" interactive exhibition
26/09/2015 Edinburgh University Doors Open Day Edinburgh, UK Lepbase Multiple Sequence Alignments game Poster+Game
28/10/2015 NextgenBUG Dundee, UK Lepbase - an Ensembl (and more) for the Lepidoptera Talk

Nematode genomics
24.06.2015 20th International C. elegans Meeting Los Angeles USA A new evolutionary framework for the phylum Nematoda: a case study of HOX cluster evolution Poster
24.06.2015 20th International C. elegans Meeting Los Angeles USA Caenorhabditis Genomes Project Workshop (organiser and chair) Talk
24.06.2015 20th International C. elegans Meeting Los Angeles USA Current status of the CGP in Edinburgh Talk

Meloidogyne genomics
10-14 August 2015 ESEB Lausanne-Switzerland Genomic consequences of hybridization and the loss of meiotic recombination in Root-knot nematodes poster
15-18 December 2015 PopGroup Edinburgh-UK Genomic consequences of hybridization and the loss of meiotic recombination in Root-knot nematodes talk
23 February 2016 NextGenBug Edinburgh-UK Genomics of Root-knot nematodes talk
Year(s) Of Engagement Activity 2015
 
Description Press releases and website 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Public/other audiences
Results and Impact We have engaged actively with the University press office to promote press coverage of our research outcomes, particularly major publications (which have had coverage in national and international newspapers) and in blogs and other online media. We have also promoted major new initiatives such as additional core funding of the Edinburgh genomics facility.

Increased visibility of Edinburgh Genomics within the community; requests for comment by funders and government on matters pertaining to genomics.
Year(s) Of Engagement Activity 2009,2010,2011,2012,2013,2014,2015,2016