Microscopic analysis of chromatin localization and dynamics in C. elegans

During development, the genome undergoes drastic reorganization within the nuclear space. To determine tridimensional genome folding, genome-wide techniques (damID/Hi-C) can be applied using cell populations, but these have to be calibrated using microscopy and single-cell analysis of gene positioning. Moreover, the dynamic behavior of chromatin has to be assessed on living samples. Combining fast stereotypic development with easy genetics and microscopy, the nematode C. elegans has become a model of choice in recent years to study changes in nuclear organization during cell fate acquisition. Here we present two complementary techniques to evaluate nuclear positioning of genes either by fluorescence in situ hybridization in fixed samples or in living worm embryos using the GFP-lacI/lacO chromatin-tagging system.

Nuclear organization in the nematode C. elegans.

With its invariant cell lineage, easy genetics and small genome, the nematode Caenorhabditis elegans has emerged as one of the prime models in developmental biology over the last 50 years. Surprisingly however, until a decade ago very little was known about nuclear organization in worms, even though it is an ideal model system to explore the link between nuclear organization and cell fate determination. Here, we review the latest findings that exploit the repertoire of genetic tools developed in worms, leading to the identification of important sequences and signals governing the changes in chromatin tridimensional architecture. We also highlight parallels and differences to other model systems.

Promoter- and RNA polymerase II-dependent hsp-16 gene association with nuclear pores in Caenorhabditis elegans.

Some inducible yeast genes relocate to nuclear pores upon activation, but the general relevance of this phenomenon has remained largely unexplored. Here we show that the bidirectional hsp-16.2/41 promoter interacts with the nuclear pore complex upon activation by heat shock in the nematode Caenorhabditis elegans. Direct pore association was confirmed by both super-resolution microscopy and chromatin immunoprecipitation. The hsp-16.2 promoter was sufficient to mediate perinuclear positioning under basal level conditions of expression, both in integrated transgenes carrying from 1 to 74 copies of the promoter and in a single-copy genomic insertion. Perinuclear localization of the uninduced gene depended on promoter elements essential for induction and required the heat-shock transcription factor HSF-1, RNA polymerase II, and ENY-2, a factor that binds both SAGA and the THO/TREX mRNA export complex. After induction, colocalization with nuclear pores increased significantly at the promoter and along the coding sequence, dependent on the same promoter-associated factors, including active RNA polymerase II, and correlated with nascent transcripts.

Building silent compartments at the nuclear periphery: a recurrent theme.

In eukaryotes, the genetic material is stored in the nucleus, which is enclosed in a double lipid bilayer, the nuclear envelope (NE). It protects the genome from physical stress and separates it from the rest of the cell. On top of this physical function, growing evidence shows that the nuclear periphery contributes to the 3D organization of the genome. In turn, tridimensional organization of chromatin in the nuclear space influences genome expression. Here we review recent findings on the function of this physical barrier in gene repression and latest models on how silent subnuclear compartments at the NE are built in yeast as well as in the nematode C. elegans and mammalian cells; trying to draw parallels between the three systems.

Phylogeography of silver pheasant (Lophura nycthemera L.) across China: aggregate effects of refugia, introgression, and riverine barriers

The role of Pleistocene glacial cycles in forming the contemporary genetic structure of organisms has been well studied in China with a particular focus on the Tibetan Plateau. However, China has a complex topography and diversity of local climates, and how glacial cycles may have shaped the subtropical and tropical biota of the region remains mostly unaddressed. To investigate the factors that affected the phylogeography and population history of a widely distributed and nondeciduous forest species, we analysed morphological characters, mitochondrial DNA sequences and nuclear microsatellite loci in the Silver Pheasant (Lophura nycthemera). In a pattern generally consistent with phenotypic clusters, but not nominal subspecies, deeply divergent mitochondrial lineages restricted to different geographic regions were detected. Coalescent simulations indicated that the time of main divergence events corresponded to major glacial periods in the Pleistocene and gene flow was only partially lowered by drainage barriers between some populations. Intraspecific cytonuclear discordance was revealed in mitochondrial lineages from Hainan Island and the Sichuan Basin with evidence of nuclear gene flow from neighbouring populations into the latter. Unexpectedly, hybridization was revealed in Yingjiang between the Silver Pheasant and Kalij Pheasant (Lophura leucomelanos) with wide genetic introgression at both the mtDNA and nuclear levels. Our results highlight a novel phylogeographic pattern in a subtropical area generated from the combined effects of climate oscillation, partial drainage barriers and interspecific hybridization. Cytonuclear discordance combined with morphological differentiation implies that complex historical factors shaped the divergence process in this biodiversity hot spot area of southern China.

Temporal genetic structure and relatedness in the Tufted Duck Aythya fuligula suggests limited kin association in winter

Conspecific aggregation of waterfowl in winter is a common example of animal flocking behaviour, yet patterns of relatedness and temporal substructure in such social groups remain poorly understood even in common species. A previous study based on mark-recapture data showed that Tufted Ducks Aythya fuligula caught on the same day were re-caught together in subsequent winters more often than expected by chance, suggesting stable assortments of ‘socially familiar’ individuals between wintering periods. The genetic relationships within these social groups were not clear. Based on 191 individuals genotyped at 10 microsatellite markers, we investigated the temporal genetic structure and patterns of relatedness among wintering Tufted Ducks at Lake Sempach, Switzerland, in two consecutive winters. We found no evidence of genetic differentiation between temporal groups within or between winters. The average levels of relatedness in temporal groups were low and not higher than expected in random assortments of individuals. However, Mantel tests performed for each sex separately revealed significant negative correlations between the pairwise relatedness coefficients and the number of days between the capture dates of pairs of wintering Tufted Duck in males and females. This pattern suggests the presence of a small number of co-migrating same-sex sibling pairs in wintering flocks of Tufted Ducks. Our findings provide one of the first genetic analyses of a common duck species outside the breeding season and contribute to the understanding of social interactions in long-distance migratory birds.

Divergent evolutionary processes associated with colonization of offshore islands

Oceanic islands have been a test ground for evolutionary theory, but here, we focus on the possibilities for evolutionary study created by offshore islands. These can be colonized through various means and by a wide range of species, including those with low dispersal capabilities. We use morphology, modern and ancient sequences of cytochrome b (cytb) and microsatellite genotypes to examine colonization history and evolutionary change associated with occupation of the Orkney archipelago by the common vole (Microtus arvalis), a species found in continental Europe but not in Britain. Among possible colonization scenarios, our results are most consistent with human introduction at least 5100 bp (confirmed by radiocarbon dating). We used approximate Bayesian computation of population history to infer the coast of Belgium as the possible source and estimated the evolutionary timescale using a Bayesian coalescent approach. We showed substantial morphological divergence of the island populations, including a size increase presumably driven by selection and reduced microsatellite variation likely reflecting founder events and genetic drift. More surprisingly, our results suggest that a recent and widespread cytb replacement event in the continental source area purged cytb variation there, whereas the ancestral diversity is largely retained in the colonized islands as a genetic ‘ark’. The replacement event in the continental M. arvalis was probably triggered by anthropogenic causes (land-use change). Our studies illustrate that small offshore islands can act as field laboratories for studying various evolutionary processes over relatively short timescales, informing about the mainland source area as well as the island.