NMR secondary structure and interactions of recombinant human MOZART1 protein,a component of the gamma‐tubulin complex

Mitotic‐spindle organizing protein associated with a ring of γ‐tubulin 1 (MOZART1) is an 8.5 kDa protein linked to regulation of γ‐tubulin ring complexes (γTuRCs), which are involved in nucleation of microtubules. Despite its small size, MOZART1 represents a challenging target for detailed characterization in vitro. We described herein a protocol for efficient production of recombinant human MOZART1 in Escherichia coli and assessed the properties of the purified protein using a combination of size exclusion chromatography coupled with multiangle light scattering (SEC‐MALS), dynamic light scattering (DLS), and nuclear magnetic resonance (NMR) experiments. MOZART1 forms heterogeneous oligomers in solution. We identified optimal detergent and buffer conditions for recording well resolved NMR experiments allowing nearly full protein assignment and identification of three distinct alpha‐helical structured regions. Finally, using NMR, we showed that MOZART1 interacts with the N‐terminus (residues 1–250) of GCP3 (γ‐tubulin complex protein 3). Our data illustrate the capacity of MOZART1 to form oligomers, promoting multiple contacts with a subset of protein partners in the context of microtubule nucleation.     

When structure leads to sex: Untangling signals in population genetic data sets

A robust signal of population structure often provides the first glimpse into the evolutionary history of a species and its populations. In this issue of Molecular Ecology, new work from Louis Bernatchez's group (Benestan et al., 2017 ) starts with an investigation of apparent structure in two marine species and concludes with an identification of sex‐linked genes, and in the process provides a model for robust analysis. Structure is the genetic signal left by natural selection as well as by neutral processes like migration and gene flow. Neutral areas of the genome can reveal the geographical relationships and related gene flow between populations over time and space, while selection can resist the natural genomic turnover created by recombination and generate adaptive structure between populations that can be detected. However, artefacts in a data set can easily hide the true signal of structure; mutation, whether it is a true appearance of a recent, minor allele, or more commonly, an error in SNP calling or molecular library construction, can easily conceal patterns of population structure (e.g., geographical structure in mackerel, Rodriguez‐Ezpeleta et al. ( 2016 )). A demographic structure that results from the most “forceful” evolutionary processes can overwhelm another signal generated by other, unrelated phenotypes. For example, the structure among diverged freshwater and marine threespine stickleback populations results from such strong selection and linkage disequilibrium across the genome that it impairs the ability to disentangle the genetic basis of particular evolved morphological traits (e.g., opercle development, Alligood ( 2017 )). Finally, there might be conflicting inferences for what underlies structure patterns. Structure may be created by differential patterns of meiotic recombination, and genetic maps are a reliable means for identifying genomic regions that resist recombination. But, without additional information (Anderson et al., 2012 ), it can be difficult to distinguish the recombination‐suppressing effect of a segregating genomic inversion (Small et al., 2016 ) from that of sex‐linked selection.     

Male and female contributions to behavioral isolation in darters as a function of genetic distance and color distance

Determining which reproductive isolating barriers arise first between geographically isolated lineages is critical to understanding allopatric speciation. We examined behavioral isolation among four recently diverged allopatric species in the orangethroat darter clade (Etheostoma: Ceasia). We also examined behavioral isolation between each Ceasia species and the sympatric rainbow darter Etheostoma caeruleum. We asked (1) is behavioral isolation present between allopatric Ceasia species, and how does this compare to behavioral isolation with E. caeruleum, (2) does male color distance and/or genetic distance predict behavioral isolation between species, and (3) what are the relative contributions of female choice, male choice, and male competition to behavioral isolation? We found that behavioral isolation, genetic differentiation, and male color pattern differentiation were present between allopatric Ceasia species. Males, but not females, discerned between conspecific and heterospecific mates. Males also directed more aggression toward conspecific rival males. The high levels of behavioral isolation among Ceasia species showed no obvious pattern with genetic distance or male color distance. However, when the E. caeruleum was included in the analysis, an association between male aggression and male color distance was apparent. We discuss the possibility that reinforcement between Ceasia and E. caeruleum is driving behavioral isolation among allopatric Ceasia species.     

Bulk hydrogen stable isotope composition of seaweeds: Clear separation between Ulvophyceae and other classes

Little is known about the bulk hydrogen stable isotope composition (δ²H) of seaweeds. This study investigated the bulk δ²H in several different seaweed species collected from three different beaches in Brazil, Australia, and Argentina. Here, we show that Ulvophyceae (a group of green algae) had lower δ²H values (between ?94‰ and ?130‰) than red algae (Florideophyceae), brown algae (Phaeophyceae), and species from the class Bryopsidophyceae (another group of green algae). Overall the latter three groups of seaweeds had δ²H values between ?50‰ and ?90‰. These findings were similar at the three different geographic locations. Observed differences in δ²H values were probably related to differences in hydrogen (H) metabolism among algal groups, also observed in the δ²H values of their lipids. The marked difference between the δ²H values of Ulvophyecae and those of the other groups could be useful to trace the food source of food webs in coastal rocky shores, to assess the impacts of green tides on coastal ecosystems, and to help clarify aspects of their phylogeny. However, reference materials for seaweed δ²H are required before the full potential of using the δ²H of seaweeds for ecological studies can be exploited.     

Giant tortoises spread to western Indian Ocean islands by sea drift in pre‐Holocene times,not by later human agency – response to Wilmé et al. (2016a)

Evidence from DNA phylogeny, Plio‐Pleistocene ocean currents, giant tortoise dispersal, evolution of plant defences, radiocarbon dates and archaeology indicates that the endemic giant tortoises on the Mascarenes and Seychelles colonized naturally and were not translocated there by humans.     

Leaf‐IT: An Android application for measuring leaf area

The use of plant functional traits has become increasingly popular in ecological studies because plant functional traits help to understand key ecological processes in plant species and communities. This also includes changes in diversity, inter‐ and intraspecific interactions, and relationships of species at different spatiotemporal scales. Leaf traits are among the most important traits as they describe key dimensions of a plant's life history strategy. Further, leaf area is a key parameter with relevance for other traits such as specific leaf area, which in turn correlates with leaf chemical composition, photosynthetic rate, leaf longevity, and carbon investment. Measuring leaf area usually involves the use of scanners and commercial software and can be difficult under field conditions. We present Leaf‐IT, a new smartphone application for measuring leaf area and other trait‐related areas. Leaf‐IT is free, designed for scientific purposes, and runs on Android 4 or higher. We tested the precision and accuracy using objects with standardized area and compared the area measurements of real leaves with the well‐established, commercial software WinFOLIA using the Altman–Bland method. Area measurements of standardized objects show that Leaf‐IT measures area with high accuracy and precision. Area measurements with Leaf‐IT of real leaves are comparable to those of WinFOLIA. Leaf‐IT is an easy‐to‐use application running on a wide range of smartphones. That increases the portability and use of Leaf‐IT and makes it possible to measure leaf area under field conditions typical for remote locations. Its high accuracy and precision are similar to WinFOLIA. Currently, its main limitation is margin detection of damaged leaves or complex leaf morphologies.     

Evaluation of several protein a resins for application to multicolumn chromatography for the rapid purification of fed‐batch bioreactors

Most of the existing production capacity is based on fed‐batch bioreactors. Thanks to the development of more efficient cell lines and the development of high‐performance culture media, cell productivity dramatically increased. In a manufacturing perspective, it is necessary to clear as quickly as possible the protein A capture step to respect the manufacturing agenda. This article describes the methodology applied for the design of a multicolumn chromatography process with the objective of purifying as quickly as possible 1,000 and 15,000 L fed‐batch bioreactors. Several recent and reference protein A resins are compared based on characteristic values obtained from breakthrough curves. The importance and relevance of resin parameters are explained, and purposely simple indicators are proposed to quickly evaluate the potential of each candidate. Based on simulation data, the optimum BioSC systems associated with each resin are then compared. The quality of the elution delivered by each resin is also compared to complete the assessment. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:941–953, 2017     

A variant ECE1 allele contributes to reduced pathogenicity of Candida albicans during vulvovaginal candidiasis

Vulvovaginal candidiasis (VVC), caused primarily by the human fungal pathogen Candida albicans, results in significant quality-of-life issues for women worldwide. Candidalysin, a toxin derived from a polypeptide (Ece1p) encoded by the ECE1 gene, plays a crucial role in driving immunopathology at the vaginal mucosa. This study aimed to determine if expression and/or processing of Ece1p differs across C. albicans isolates and whether this partly underlies differential pathogenicity observed clinically. Using a targeted sequencing approach, we determined that isolate 529L harbors a similarly expressed, yet distinct Ece1p isoform variant that encodes for a predicted functional candidalysin; this isoform was conserved amongst a collection of clinical isolates. Expression of the ECE1 open reading frame (ORF) from 529L in an SC5314-derived ece1Δ/Δ strain resulted in significantly reduced vaginopathogenicity as compared to an isogenic control expressing a wild-type (WT) ECE1 allele. However, in vitro challenge of vaginal epithelial cells with synthetic candidalysin demonstrated similar toxigenic activity amongst SC5314 and 529L isoforms. Creation of an isogenic panel of chimeric strains harboring swapped Ece1p peptides or HiBiT tags revealed reduced secretion with the ORF from 529L that was associated with reduced virulence. A genetic survey of 78 clinical isolates demonstrated a conserved pattern between Ece1p P2 and P3 sequences, suggesting that substrate specificity around Kex2p-mediated KR cleavage sites involved in protein processing may contribute to differential pathogenicity amongst clinical isolates. Therefore, we present a new mechanism for attenuation of C. albicans virulence at the ECE1 locus.