An Optimized Proteomics Approach Reveals Novel Alternative Proteins in Mouse Liver Development

Alternative ORFs (AltORFs) are unannotated sequences in genome that encode novel peptides or proteins named alternative proteins (AltProts). Although ribosome profiling and bioinformatics predict a large number of AltProts, mass spectrometry as the only direct way of identification is hampered by the short lengths and relative low abundance of AltProts. There is an urgent need for improvement of mass spectrometry methodologies for AltProt identification. Here, we report an approach based on size-exclusion chromatography for simultaneous enrichment and fractionation of AltProts from complex proteome. This method greatly simplifies the variance of AltProts discovery by enriching small proteins smaller than 40 kDa. In a systematic comparison between 10 methods, the approach we reported enabled the discovery of more AltProts with overall higher intensities, with less cost of time and effort compared to other workflows. We applied this approach to identify 89 novel AltProts from mouse liver, 39 of which were differentially expressed between embryonic and adult mice. During embryonic development, the upregulated AltProts were mainly involved in biological pathways on RNA splicing and processing, whereas the AltProts involved in metabolisms were more active in adult livers. Our study not only provides an effective approach for identifying AltProts but also novel AltProts that are potentially important in developmental biology.     

Effect of Different Carbon and Nitrogen Sources on Clostridium argentinense Toxigenicity in Coculture withPseudomonas mendocina

The effect of different carbon and nitrogen sources on the production of toxin by Clostridium argentinense was examined. The toxin production by C. argentinense in coculture with Pseudomonas mendocina increased in all the cases in relation to that produced by monocultures independent of the nature of the source. Using dextrin as carbon source C. argentinense produced the highest levels of toxin both in monocultures (300 LD₅₀/mL) and in cocultures with P. mendocina (5000 LD₅₀/mL). Experiments run in a microfermenter showed that the slow growth of cocultures associated with the assimilation of dextrin and the pH and Eh profiles favoured the production of toxin. Of the nitrogen sources assayed, corn steep liquor sustained the highest levels of toxin in both monocultures and cocultures with 3 and 2.8 fold increases with respect to that obtained using proteose peptone. The toxin production by C. argentinense cultures and C. argentinense–P. mendocina cocultures was highly dependent on the nature of the carbon and nitrogen sources used in the culture media. Growth of C. argentinense on substrates slowly assimilated stimulated the production of toxin.     

Distinguishing Drift and Selection Empirically: “The Great Snail Debate” of the 1950s

Biologists and philosophers have been extremely pessimistic about the possibility of demonstrating random drift in nature, particularly when it comes to distinguishing random drift from natural selection. However, examination of a historical case – Maxime Lamotte’s study of natural populations of the land snail, Cepaea nemoralis in the 1950s – shows that while some pessimism is warranted, it has been overstated. Indeed, by describing a unique signature for drift and showing that this signature obtained in the populations under study, Lamotte was able to make a good case for a significant role for␣drift. It may be difficult to disentangle the causes of drift and selection acting in a population, but it is not (always) impossible.     

Climate oscillations and species interactions: large‐scale congruence but regional differences in the phylogeographic structures of an alpine plant and its monophagous insect

Aim To predict the fate of alpine interactions involving specialized species, using a monophagous beetle and its host plant as a case study. Location The Alps. Methods We investigated genetic structuring of the herbivorous beetle Oreina gloriosa and its specific host‐plant Peucedanum ostruthium. We used genome fingerprinting (in the insect and the plant) and sequence data (in the insect) to compare the distribution of the main gene pools in the two associated species and to estimate divergence time in the insect, a proxy for the temporal origin of the interaction. We quantified the similarity in spatial genetic structures by performing a Procrustes analysis, a tool from shape theory. Finally, we simulated recolonization of an empty space analogous to the deglaciated Alps just after ice retreat by two lineages from two species showing unbalanced dependence, to examine how timing of the recolonization process, as well as dispersal capacities of associated species, could explain the observed pattern. Results Contrasting with expectations based on their asymmetrical dependence, patterns in the beetle and plant were congruent at a large scale. Exceptions occurred at a regional scale in areas of admixture, matching known suture zones in Alpine plants. Simulations using a lattice‐based model suggested these empirical patterns arose during or soon after recolonization, long after the estimated origin of the interaction c. 0.5 million years ago. Main conclusions Species‐specific interactions are scarce in alpine habitats because glacial cycles have limited the opportunities for co‐evolution. Their fate, however, remains uncertain under climate change. Here we show that whereas most dispersal routes are paralleled at a large scale, regional incongruence implies that the destinies of the species might differ under changing climate. This may be a consequence of the host dependence of the beetle, which locally limits the establishment of dispersing insects.     

Corals shed bacteria as a potential mechanism of resilience to organic matter enrichment

Understanding the mechanisms of resilience of coral reefs to anthropogenic stressors is a critical step toward mitigating their current global decline. Coral–bacteria associations are fundamental to reef health and disease, but direct observations of these interactions remain largely unexplored. Here, we use novel technology, high-speed laser scanning confocal microscopy on live coral (Pocillopora damicornis), to test the hypothesis that corals exert control over the abundance of their associated bacterial communities by releasing (‘shedding'') bacteria from their surface, and that this mechanism can counteract bacterial growth stimulated by organic inputs. We also test the hypothesis that the coral pathogen Vibrio coralliilyticus can evade such a defense mechanism. This first report of direct observation with high-speed confocal microscopy of living coral and its associated bacterial community revealed a layer (3.3–146.8 μm thick) on the coral surface where bacteria were concentrated. The results of two independent experiments showed that the bacterial abundance in this layer was not sensitive to enrichment (5 mg l⁻¹ peptone), and that coral fragments exposed to enrichment released significantly more bacteria from their surfaces than control corals (P<0.01; 35.9±1.4 × 10⁵ cells cm⁻² coral versus 1.3±0.5 × 10⁵ cells cm⁻² coral). Our results provide direct support to the hypothesis that shedding bacteria may be an important mechanism by which coral-associated bacterial abundances are regulated under organic matter stress. Additionally, the novel ability to watch this ecological behavior in real-time at the microscale opens an unexplored avenue for mechanistic studies of coral–microbe interactions.     

Amazonian biodiversity: assessing conservation priorities with taxonomic data

Data from 3991 records of museum collections representing 421 species of plants, arthropods, amphibians, fish, and primates were analyzed with GIS to identify areas of high species diversity and endemism in Amazonia. Of the 472 1 × 1° grid cells in Amazonia, only nine cells are included in the highest species diversity category (43–67 total species) and nine in the highest endemic species diversity category (4–13 endemic species). Over one quarter of the grid cells have no museum records of any of the organisms in our study. Little correspondence exists between the centers of species diversity identified by our collections-based data and those areas recommended for conservation in an earlier qualitative study of Amazonian biodiversity. Museum collections can play a vital role in identifying species-rich areas for potential conservation in Amazonia, but a concerted and structured effort to increase the number and distribution of collections is needed to take maximum advantage of the information they contain.     

Modelling minirhizotron observations to test experimental procedures

In order to help design experiments with minirhizotrons or interpret data from such experiments, a modelling approach is a valuable tool to complement empirical approaches. The general principle of this modelling approach is to calculate and to study the part of a theoretical root system that is intersected by passes through a virtual minirhizotron tube (modelled here as a cylinder). Various outputs can be calculated from this part of the root system, and related to the surrounding root system which is perfectly known, since it has been simulated and stored in a data structure. Therefore, the method involves two levels of modelling that are presented and discussed: the root system architecture of a crop, and the observations that can be achieved with minirhizotron tubes. Illustrations of the method are presented to study the effect of several factors on the rooting depth curves, and to show how images may be calculated to mimic what can actually be viewed from inside the tube. These first results show that the maximum rooting depth curves, as virtually observed in the minirhizotron tube, present large variations and strongly underestimate the maximum rooting depth of the modelled root system (up to 60 cm in average). The underestimation is still more critical when the radius of the tube is lower than 3 cm, and when the tube is close to the vertical (angle lower than 0.2 rad). The use of the 0.9 quantile instead of the average value, for each of the observation dates, leads to a better estimation of the maximum rooting depth.     

Disrupted phylogeographical microsatellite and chloroplast DNA patterns indicate a vicariance rather than long‐distance dispersal origin for the disjunct distribution of the Chilean endemic Dioscorea biloba (Dioscoreaceae) around the Atacama Desert

Aim The Chilean endemic Dioscorea biloba (Dioscoreaceae) is a dioecious geophyte that shows a remarkable 600 km north–south disjunction in the peripheral arid area of the Atacama Desert. Its restricted present‐day distribution and probable Neogene origin indicate that its populations have a history linked to that of the Atacama Desert, making this an ideal model species with which to investigate the biogeography of the region. Location Chile, Atacama Desert and peripheral arid area. Methods Two hundred and seventy‐five individuals from nine populations were genotyped for seven nuclear microsatellite loci, and plastid trnL–F and trnT–L sequences were obtained for a representative subset of these. Analyses included the estimation of genetic diversity and population structure through clustering, Bayesian and analysis of molecular variance analyses, and statistical parsimony networks of chloroplast haplotypes. Isolation by distance was tested against alternative dispersal hypotheses. Results Microsatellite markers revealed moderate to high levels of genetic diversity within populations, with those from the southern Limarí Valley showing the highest values and northern populations showing less exclusive alleles. Bayesian analysis of microsatellite data identified three genetic groups that corresponded to geographical ranges. Chloroplast phylogeography revealed no haplotypes shared between northern and southern ranges, and little haplotype sharing between the two neighbouring southern valleys. Dispersal models suggested the presence of extinct hypothetical populations between the southern and northern ranges. Main conclusions Our results are consistent with prolonged isolation of the northern and southern groups, mediated by the life‐history traits of the species. Significant isolation was revealed at both large and moderate distances as gene flow was not evident even between neighbouring valleys. Bayesian analyses of microsatellite and chloroplast haplotype diversity identified the southern area of Limarí as the probable area of origin of the species. Our data do not support recent dispersal of D. biloba from the southern range into Antofagasta, but indicate the fragmentation of an earlier wider range, concomitant with the Pliocene–Pleistocene climatic oscillations, with subsequent extinctions of the Atacama Desert populations and the divergence of the peripheral ones as a consequence of genetic drift.