Complex archaea that bridge the gap between prokaryotes and eukaryotes.

The origin of the eukaryotic cell remains one of the most contentious puzzles in modern biology. Recent studies have provided support for the emergence of the eukaryotic host cell from within the archaeal domain of life, but the identity and nature of the putative archaeal ancestor remain a subject of debate. Here we describe the discovery of 'Lokiarchaeota', a novel candidate archaeal phylum, which forms a monophyletic group with eukaryotes in phylogenomic analyses, and whose genomes encode an expanded repertoire of eukaryotic signature proteins that are suggestive of sophisticated membrane remodelling capabilities. Our results provide strong support for hypotheses in which the eukaryotic host evolved from a bona fide archaeon, and demonstrate that many components that underpin eukaryote-specific features were already present in that ancestor. This provided the host with a rich genomic 'starter-kit' to support the increase in the cellular and genomic complexity that is characteristic of eukaryotes.

Testing sensory drive speciation in cichlid fish: Linking light conditions to opsin expression, opsin genotype and female mate preference.

Ecological speciation is facilitated when divergent adaptation has direct effects on selective mating. Divergent sensory adaptation could generate such direct effects, by mediating both ecological performance and mate selection. In aquatic environments, light attenuation creates distinct photic environments, generating divergent selection on visual systems. Consequently, divergent sensory drive has been implicated in the diversification of several fish species. Here, we experimentally test whether divergent visual adaptation explains the divergence of mate preferences in Haplochromine cichlids. Blue and red Pundamilia co-occur across south-eastern Lake Victoria. They inhabit different photic conditions and have distinct visual system properties. Previously, we documented that rearing fish under different light conditions influences female preference for blue versus red males. Here, we examine to what extent variation in female mate preference can be explained by variation in visual system properties, testing the causal link between visual perception and preference. We find that our experimental light manipulations influence opsin expression, suggesting a potential role for phenotypic plasticity in optimizing visual performance. However, variation in opsin expression does not explain species differences in female preference. Instead, female preference covaries with allelic variation in the long-wavelength-sensitive opsin gene (LWS), when assessed under broad-spectrum light. Taken together, our study presents evidence for environmental plasticity in opsin expression and confirms the important role of colour perception in shaping female mate preferences in Pundamilia. However, it does not constitute unequivocal evidence for the direct effects of visual adaptation on assortative mating.

DNA nuclear targeting sequences for non-viral gene delivery.

PURPOSE: To evaluate if introduction of DNA nuclear Targeting Sequences (DTS; i.e. recognition sequences for endogenous DNA-binding proteins) in plasmid DNA (pDNA) leads to increased transfection efficiency of non-viral gene delivery by virtue of enhanced nuclear import of the pDNA. METHODS: A set of DTS was identified and cloned into EGFP-reporter plasmids controlled by the CMV-promoter. These pDNA constructs were delivered into A431 and HeLa cells using standard electroporation, pEI-based polyfection or lipofection methods. The amount of pDNA delivered into the nucleus was determined by qPCR; transfection efficiency was determined by flow cytometry. RESULTS: Neither of these DTS increased transgene expression. We varied several parameters (mitotic activity, applied dose and delivery strategy), but without effect. Although upregulated transgene expression was observed after stimulation with TNF-α, this effect could be ascribed to non-specific upregulation of transcription rather than enhanced nuclear import. Nuclear copy numbers of plasmids containing or lacking a DTS did not differ significantly after lipofectamine-based transfection in dividing and non-dividing cells. CONCLUSION: No beneficial effects of DTS on gene expression or nuclear uptake were observed in this study.

Synthesis and characterization of hydroxyl-functionalized caprolactone copolymers and their effect on adhesion, proliferation, and differentiation of human mesenchymal stem cells.

The aim of this study was to develop new hydrophilic polyesters for tissue engineering applications. In our approach, poly(benzyloxymethyl glycolide-co-epsilon-caprolactone)s (pBHMG-CLs) were synthesized through melt copolymerization of epsilon-caprolactone (CL) and benzyl-protected hydroxymethyl glycolide (BHMG). Deprotection of the polymers yielded copolymers with pendant hydroxyl groups, poly(hydroxymethylglycolide-co-epsilon-caprolactone) (pHMG-CL). The synthesized polymers were characterized by GPC, NMR, and DSC techniques. The resulting copolymers consisting of up to 10% of HMG monomer were semicrystalline with a melting temperature above body temperature. Water contact angle measurements of polymeric films showed that increasing HMG content resulted in higher surface hydrophilicity, as evidenced from a decrease in receding contact angle from 68 degrees for PCL to 40 degrees for 10% HMG-CL. Human mesenchymal stem cells showed good adherence onto pHMG-CL films as compared to the more hydrophobic PCL surfaces. The cells survived and were able to differentiate toward osteogenic lineage on pHMG-CL surfaces. This study shows that the aforementioned hydrophilic polymers are attractive candidates for the design of scaffolds for tissue engineering applications.

Geographic variation in opsin expression does not align with opsin genotype in Lake Victoria cichlid populations.

Sensory adaptation to the local environment can contribute to speciation. Aquatic environments are well suited for studying this process: The natural attenuation of light through water results in heterogeneous light environments, to which vision-dependent species must adapt for communication and survival. Here, we study visual adaptation in sympatric Pundamilia cichlids from southeastern Lake Victoria. Species with blue or red male nuptial coloration co-occur at many rocky islands but tend to be depth-differentiated, entailing different visual habitats, more strongly at some islands than others. Divergent visual adaptation to these environments has been implicated as a major factor in the divergence of P. pundamilia and P. nyererei, as they show consistent differentiation in the long-wavelength-sensitive visual pigment gene sequence (LWS opsin). In addition to sequence variation, variation in the opsin gene expression levels may contribute to visual adaptation. We characterized opsin gene expression and LWS genotype across Pundamilia populations inhabiting turbid and clear waters, to examine how different mechanisms of visual tuning contribute to visual adaptation. As predicted, the short-wavelength-sensitive opsin (SWS2b) was expressed exclusively in a population from clear water. Contrary to prediction however, expression levels of the other opsins were species- and island-dependent and did not align with species differences in LWS genotype. Specifically, in two locations with turbid water, the shallow-water dwelling blue species expressed more LWS and less RH2A than the deeper-dwelling red species, while the opposite pattern occurred in the two locations with clear water. Visual modeling suggests that the observed distribution of opsin expression profiles and LWS genotypes does not maximize visual performance, implying the involvement of additional visual tuning mechanisms and/or incomplete adaptation. OPEN RESEARCH BADGE: This article has earned an Open Data Badge for making publicly available the digitally-shareable data necessary to reproduce the reported results. The data is available at https://hdl.handle.net/10411/I1IUUQ.

Last male sperm precedence is modulated by female remating rate in Drosophila melanogaster.

Following multiple matings, sperm from different males compete for fertilization within the female reproductive tract. In many species, this competition results in an unequal sharing of paternity that favors the most recent mate, termed last male sperm precedence (LMSP). Much of our understanding of LMSP comes from studies in Drosophila melanogaster that focus on twice-mated females with standardized latencies between successive matings. Despite accumulating evidence indicating that females often mate with more than two males and exhibit variation in the latency between matings, the consequences of mating rate on LMSP are poorly understood. Here, we developed a paradigm utilizing D. melanogaster in which females remated at various time intervals with either two or three transgenic males that produce fluorescent sperm (green, red, or blue). This genetic manipulation enables paternity assessment of offspring and male-specific sperm fate examination in female reproductive tracts. We found that remating latency had no relationship with LMSP in females that mated with two males. However, LMSP was significantly reduced in thrice-mated females with short remating intervals; coinciding with reduced last-male sperm storage. Thus, female remating rate influences the relative share of paternity, the overall clutch paternity diversity, and ultimately the acquisition of indirect genetic benefits to potentially maximize female reproductive success.

Serotonin Transporter Genotype Modulates the Gut Microbiota Composition in Young Rats, an Effect Augmented by Early Life Stress.

The neurotransmitter serotonin (5-HT) plays a vital regulatory role in both the brain and gut. 5-HT is crucial for regulating mood in the brain as well as gastrointestinal motility and secretion peripherally. Alterations in 5-HT transmission have been linked to pathological symptoms in both intestinal and psychiatric disorders and selective 5-HT transporter (5-HTT) inhibitors, affecting the 5-HT system by blocking the 5-HT transporter (5-HTT) have been successfully used to treat CNS- and intestinal disorders. Humans that carry the short allele of the 5-HTT-linked polymorphic region (5-HTTLPR) are more vulnerable to adverse environmental stressors, in particular early life stress. Although, early life stress has been shown to alter the composition of the gut microbiota, it is not known whether a lower 5-HTT expression is also associated with an altered microbiome composition. To investigate this, male and female wild type (5-HTT+/+), heterozygous (5-HTT+/-), and knockout (5-HTT-/-) 5-HT transporter rats were maternally separated for 6 h a day from postnatal day 2 till 15. On postnatal day 21, fecal samples were collected and the impact of 5-HTT genotype and maternal separation (MS) on the microbiome was analyzed using high-throughput sequencing of the bacterial 16S rRNA gene. MS showed a shift in the ratio between the two main bacterial phyla characterized by a decrease in Bacteroidetes and an increase in Firmicutes. Interestingly, the 5-HTT genotype caused a greater microbal dysbiosis (microbial imbalance) compared with MS. A significant difference in microbiota composition was found segregating 5-HTT-/- apart from 5-HTT+/- and 5-HTT+/+ rats. Moreover, exposure of rats with 5-HTT diminished expression to MS swayed the balance of their microbiota away from homeostasis to 'inflammatory' type microbiota characterized by higher abundance of members of the gut microbiome including Desulfovibrio, Mucispirillum, and Fusobacterium, all of which are previously reported to be associated with a state of intestinal inflammation, including inflammation associated with MS and brain disorders like multiple depressive disorders. Overall, our data show for the first time that altered expression of 5-HTT induces disruptions in male and female rat gut microbes and these 5-HTT genotype-related disruptions are augmented when combined with early life stress.

How to screen non-viral gene delivery systems in vitro?

Screening of new gene delivery candidates regarding transfection efficiency and toxicity is usually performed by reading out transgene expression levels relative to a reference formulation after in vitro transfection. However, over the years and among different laboratories, this screening has been performed in a variety of cell lines, using a variety of conditions and read-out systems, and by comparison to a variety of reference formulations. This makes a direct comparison of results difficult, if not impossible. Reaching a consensus would enable placing new results into context of previous findings and estimate the overall contribution to the improvement of non-viral gene delivery. In this paper we illustrate the sensitivity of transfection outcomes on testing conditions chosen, and propose a screening protocol with the aim of standardization within the field.