Population Genetics Research

MOLECULAR BASIS AND EVOLUTION OF REPRODUCTIVE ISOLATION

Mechanisms underlying speciation are of interest in life science since Darwin introduced his concept of the rise of species into modern biology. One major step in speciation is the evolution of reproductive isolation. Complete reproductive isolation is achieved if potential mating partners do not recognize each other (prezygotic isolation). On the other hand, mating partners might still try to reproduce but zygotes fail to develop at all or develop to sterile adults. In contrast, reproductive isolation can be incomplete were mates have either a reduced affinity to each other (incomplete prezygotic isolation) or zygotes develop to non sterile adults with reduced fitness. The case were two incompletely isolated species form a stable hybrid zone is a highly interesting model to investigate mechanisms underlying reproductive isolation and hence speciation.

We use the Mytilus edulis-species complex as a model to investigate the molecular basis of reproductive isolation. This species complex consists of three species: Mytilus edulisM. galloprovincialis, and M. trossulus.

The Mytilus edulis – species complex is well suited as model because of several reasons. It is a marine sessile invertebrate and males and females release their gametes into the free water. Thus, processes such as species recognition and mate choice mainly occur at the level of sperm-egg-interaction while processes such as behaviour are almost not relevant. Consequently, the identification of gamete proteins and the understanding of evolutionary forces acting on their corresponding genes will largely contribute to an understanding of reproductive isolation. In this respect, M. edulis is comparable to marine sessile invertebrates such as abalone and sea urchin that have been intensively studied in the context of reproductive traits, e.g., by investigating gamete recognition proteins such as the bindin protein of sea urchin or lysin of abalone. However, a distinctive feature of M. edulis is its ability to hybridize in zones of secondary contact with M. galloprovincialis and M. trossulus. This allows comparatively investigating the molecular features of a given candidate molecule in genetically pure and hybrid populations.

One basic precondition for investigating the molecular basis of reproductive isolation is the availability of marker proteins and their corresponding genes. Such genes should play a key role in several aspects of egg- and sperm function. Recently, research on Mytilus-reproduction focuses on a single protein, M7 lysin. This protein is located in the sperm acrosome and plays a role in gamete recognition and first polar body formation. However, to comprehensively understand the evolution of reproductive isolation it would be desirable to investigate a whole set of candidate genes being functionally important in gamete recognition as well as in gamete performance. Such a broad view is indeed justified. For instance, reproductive success in Mytilus-species does not only depend on gamete interaction but also on processes such as gamete competition. Gamete competition can be attributed to characteristics like sperm motility, efficiency to induce the acrosome reaction, or response to chemoattractants.

Thus, one focus of our research is the identification of new candidate proteins and their corresponding genes with relevance to gamete function. In order to do so, we recently employed selective antibody production.

This procedure used fragmented M. edulis sperm as antigen to immunize mice. Hybridoma cell clones were generated and each cell clone can theoretically produce antibodies against different factors. These antibodies can target virtually any factor present in sperm. Thus, these different monoclonal antibodies are subsequently screened (e.g., by Western blot or immunohistochemistry) and those showing a sperm-biased binding pattern are selected. Although the exact identity of each antibody target is unknown at first hand, the target molecules of the selected antibodies must be preferentially present in sperm; hence these target molecules most likely represent a molecule with functional relevance to sperm function and reproduction. Thus, monoclonal antibodies can be used as selective tools to isolate proteins directly from M. edulis sperm and to identify them, e.g., by performing de novo-sequencing of peptides using tandem mass spectrometry.

Alternatively, we use computational approaches to screen public and non-public databases to identify candidate genes with relevance to gamete function. In order to do so, candidate genes are listed that were experimentally characterized as factors important for gamete function in model organisms such as mouse. The corresponding nucleotide and/or amino acid sequences are used to identify Mytilus and non-Mytilus orthologs from genomic and transcriptomic databases.

Newly identified candidate genes (e.g., using either selective antibody production or computational approaches) are investigated in naturally occurring allopatric and sympatric populations. Molecular tools are employed to estimate diversity and divergence parameters, e.g., to study patterns of natural selection. In addition, we investigate gene flow across hybrid zones. If a gene plays an important role in the speciation process, one expects i) patterns of positive selection and/or ii) restricted interspecies gene flow.

One particular focus of our research is on analysing the Baltic contact zone between M. edulis and M. trossulus. A comparable contact zone between both species is found in North America. However, remarkable differences between both contact zones exist: while interspecies gene flow in North America is limited, massive introgression is observed in the Baltic and these populations are considered as hybrid swarm. Therefore, the question arose whether the Baltic hybrid swarm is the result of non-existing reproductive barriers and/or the result of adaptation to the extreme environmental conditions (e.g., salinity gradient). The ongoing research projects aim at using the newly identified markers to analyse whether or not Baltic M. edulis and M. trossulus are at least partly reproductively isolated. In addition, we are currently establishing research activities to analyse molecular mechanisms of environmental adaptation to find out whether gene flow may be the basis for a better adaptation to environmental conditions such as low salinity.

SELECTED PROJECTS
  • Molecular analyses of Mytilus-sperm using monoclonal antibodies. (Funded by Paul-Ungerer-Foundation, February 2010 – November 2010)
  • Molecular basis of reproductive isolation in blue mussels (Mytilus, Bivalvia): Identification and characterization of gamete-specific factors. (Funded by Deutsche Forschungsgemeinschaft, DFG, STU 519/2-1. January 2011-March 2012)
  • Molecular evolution of proteins with functional relevance to sperm function. (January 2012-present)

 

ADAPTIVE VARIATION AND HYBRID ZONES

The genetic structure of a population is substantially shaped by genetic drift, i.e., allele frequencies at a given coding or non-coding locus are controlled by random effects rather than selection. However, genetic loci that are functionally related to adaptive traits might be under positive selection, e.g., genes related to tolerance towards temperature or salinity in marine habitats. The identification of genetic loci that are functionally related to adaptation is an important issue to understand mechanisms underlying the distribution of species or the formation of interspecies hybrids. Thus, the genetic basis of adaptive traits in pure species and interspecies hybrids is the subject of several research projects.

One focus is on understanding of adaptive traits in Baltic Mytilus-populations. As mentioned in the context of reproductive isolation, gene flow between Baltic Mytilus-species is massive (hybrid swarm) but the morphological and genetic integrity between Baltic M. edulis and M. trossulus is maintained. The overall goal of our research is to unravel the relative contribution of reproductive isolation and adaptive traits to maintain the integrity of Baltic Mytilus-species. Recent research activities aim at investigating how mussels can adapt to the low salinity conditions in the Baltic and whether hybridization plays a key role in that process. An additional focus is on understanding the molecular basis of temperature adaptation of butterflies (Hyles spec., Lepidoptera, Insecta). By understanding salinity adaptation of Mytilus-species and temperature tolerance of butterflies we are also contributing to applied issues, i.e., the prediction of consequences related to climate change.

SELECTED PROJECTS
  • Analyses of interspecific gene flow in Baltic Mytilus-populations (M. edulisM. edulis x M. trossulus hybrids) using RNAseq data: Functional importnace of introgressive hybridization in extreme environments. (March 2013-present, collaboration with Prof. Dr. Frank Melzner and Corinna Breusing, GEOMAR, Helmholtz Centre for Ocean Reserach Kiel)
  • Molecular basis of temperature adaptation in Hyles species (Insecta, Lepidoptera). (Collaboration with Dr. Anna Hundsdörfer, haed of DNA-Laboratory, Senckenberg Dresden)

GENETIC STRUCTURE OF POPULATIONS

The methodological profile of our research group allows analysing the genetic composition of populations and enables to infer mechanisms that shaped a particular genetic structure of populations. These methods are the basis for answering many biological questions in fields such as evolutionary biology, phylogeography or conservation genetics. This leaves room to perform not only projects related to understanding the genetic structure of Mytilus-populations but also to establish external collaborations or research links to other sections and subdivisions at Senckenberg Dresden.

SELECTED PROJECTS
  • Speciation processes in  two co-distributed freshwater turtle complexes (Emys specMauremys spec.).(2010-present; collaboration with Prof. Dr. Uwe Fritz  and Melita Vamberger, phylogeography subdivision, Senckenberg Dresden)
  • Population genetic analysis of marine organisms. (Jan 2013-present; collaboration with Prof. Dr. Pedro Martinez Arbizu, Dr. Michael Raupach, Senckenberg am Meer, German Centre for Marine Biodiversity Research, Wilhelmshaven)