Mothers with higher twinning propensity had lower fertility in pre-industrial Europe.

Historically, mothers producing twins gave birth, on average, more often than non-twinners. This observation has been interpreted as twinners having higher intrinsic fertility - a tendency to conceive easily irrespective of age and other factors - which has shaped both hypotheses about why twinning persists and varies across populations, and the design of medical studies on female fertility. Here we show in >20k pre-industrial European mothers that this interpretation results from an ecological fallacy: twinners had more births not due to higher intrinsic fertility, but because mothers that gave birth more accumulated more opportunities to produce twins. Controlling for variation in the exposure to the risk of twinning reveals that mothers with higher twinning propensity - a physiological predisposition to producing twins - had fewer births, and when twin mortality was high, fewer offspring reaching adulthood. Twinning rates may thus be driven by variation in its mortality costs, rather than variation in intrinsic fertility.

A robust sequencing assay of a thousand amplicons for the high-throughput population monitoring of Alpine ibex immunogenetics.

Polymorphism for immune functions can explain significant variation in health and reproductive success within species. Drastic loss in genetic diversity at such loci constitutes an extinction risk and should be monitored in species of conservation concern. However, effective implementations of genome-wide immune polymorphism sets into high-throughput genotyping assays are scarce. Here, we report the design and validation of a microfluidics-based amplicon sequencing assay to comprehensively capture genetic variation in Alpine ibex (Capra ibex). This species represents one of the most successful large mammal restorations recovering from a severely depressed census size and a massive loss in diversity at the major histocompatibility complex (MHC). We analysed 65 whole-genome sequencing sets of the Alpine ibex and related species to select the most representative markers and to prevent primer binding failures. In total, we designed ~1,000 amplicons densely covering the MHC, further immunity-related genes as well as randomly selected genome-wide markers for the assessment of neutral population structure. Our analysis of 158 individuals shows that the genome-wide markers perform equally well at resolving population structure as RAD-sequencing or low-coverage genome sequencing data sets. Immunity-related loci show unexpectedly high degrees of genetic differentiation within the species. Such information can now be used to define highly targeted individual translocations. Our design strategy can be realistically implemented into genetic surveys of a large range of species. In conclusion, leveraging whole-genome sequencing data sets to design targeted amplicon assays allows the simultaneous monitoring of multiple genetic risk factors and can be translated into species conservation recommendations.

Heritable spouse effects increase evolutionary potential of human reproductive timing.

Sexual reproduction is inherently interactive, especially in animal species such as humans that exhibit extended pair bonding. Yet we have little knowledge of the role of male characteristics and their evolutionary impact on reproductive behavioural phenotypes, to the extent that biologists typically consider component traits (e.g. reproductive timing) as female-specific. Based on extensive genealogical data detailing the life histories of 6435 human mothers born across four centuries of modern history, we use an animal modelling approach to estimate the indirect genetic effect of men on the reproductive phenotype of their partners. These analyses show that a woman's reproductive timing (age at first birth) is influenced by her partner's genotype. This indirect genetic effect is positively correlated with the direct genetic effect expressed in women, such that total heritable variance in this trait is doubled when heritable partner effects are considered. Our study thus suggests that much of the heritable variation in women's reproductive timing is mediated via partner effects, and that the evolutionary potential of this trait is far greater than previously appreciated.

Simian virus 40 strains with novel properties generated by replacing the viral enhancer with synthetic oligonucleotides.

Typical enhancers of viral or cellular genes are approximately 100 to 400 bp long and contain several transcription factor binding sites. Previously, we have shown that simian virus 40 (SV40) genomic DNA that lacks its own enhancer can be used as an "enhancer trap" since it reacquires infectivity upon incorporation of heterologous enhancers. Here, we show that SV40 infectivity can be restored with synthetic enhancers that are assembled by the host cell. We found that several oligonucleotides, cotransfected with enhancerless SV40 DNA into host cells, were incorporated into the viral genome via cellular DNA end joining. The oligonucleotides tested included metal response elements (MREs), the binding sites for the transcription factor MTF-1, which induces gene activity in response to heavy metals. These recombinant SV40 strains showed preferential growth on cells overloaded with zinc or cadmium. We also cotransfected enhancerless SV40 DNA with oligonucleotides corresponding to enhancer motifs of human and mouse cytomegalovirus (HCMV and MCMV, respectively). In contrast to SV40 wild type, the viruses with cytomegalovirus-derived patchwork enhancers strongly expressed T-antigen in human HEK293 cells, accompanied by viral DNA replication. Occasionally, we also observed the assembly of functional viral genomes by incorporation of fragments of bovine DNA, an ingredient of the fetal calf serum in the medium. These fragments contained, among other sites, binding sites for AP-1 and CREB transcription factors. Taken together, our studies show that viruses with novel properties can be generated by intracellular incorporation of synthetic enhancer DNA motifs.

Dissection of Drosophila MTF-1 reveals a domain for differential target gene activation upon copper overload vs. copper starvation.

Metal-responsive transcription factor-1 (MTF-1) is a zinc finger protein conserved from mammals to insects. It mediates protection against heavy metal load by activating the expression of metallothionein and other genes. In Drosophila, MTF-1 serves a dual function in that it not only helps to protect against heavy metal load but also induces the expression of Ctr1B, the gene for an intestinal copper importer, upon copper starvation. By dissecting Drosophila MTF-1 function, we have identified determinants for nuclear import and export, and characterized a phosphorylation site mutant (T127A) that differentially affects MTF-1 target genes. Further, by generating a series of fusion proteins with the heterologous DNA binding domain of Gal4 we identified a strong, constitutive activation domain in the central region of MTF-1 (aa 352-540). By contrast, an extended fusion protein that includes MTF-1's C-terminus (aa 352-791) is not active in standard conditions but induced by copper load. The paramount regulatory importance of the C-terminal part, that harbors a cysteine-rich "metallothionein-like" domain, was corroborated by different experiments. Transgenic flies expressing C-terminally truncated MTF-1 variants displayed high constitutive transcription of both, the genes for metallothioneins and the copper importer Ctr1B. The indiscriminate activation of these genes that are normally induced under opposite conditions of copper load and copper starvation manifested itself in a shortened lifespan, crippled wings, and female sterility.