Purification and analysis of a 5kDa component of enamel matrix derivative

High performance liquid chromatography (HPLC) methods were used to analyse a 5kDa component purified from enamel matrix derivative (EMD), the active ingredient in Emdogain, a commercial product for periodontal tissue regeneration. After initial purification by size-exclusion chromatography (SEC) on a 100 cm x 5 cm column (Bio-Gel P-30 Fine, 280 nm), collected fractions were analysed by size-exclusion HPLC (SE HPLC; TSK-Gel Super SW2000, 220 nm). The fractions containing only the 5kDa component were analysed by reversed-phase high-pressure chromatography (RP HPLC; YMC-Pack ODS-A, 200 nm), revealing four peaks of the 5kDa component. From 1200 mg of EMD (of which 9% is the 5kDa component), approximately 65 mg of lyophilised 5kDa component were obtained, corresponding to a recovery of 60%. The SE HPLC method was mainly suitable for qualitative analysis, whereas the RP HPLC method was appropriate for both qualitative and quantitative analysis.     

Evolution after gene duplication: models, mechanisms, sequences, systems, and organisms

Gene duplication is postulated to have played a major role in the evolution of biological novelty. Here, gene duplication is examined across levels of biological organization in an attempt to create a unified picture of the mechanistic process by which gene duplication can have played a role in generating biodiversity. Neofunctionalization and subfunctionalization have been proposed as important processes driving the retention of duplicate genes. These models have foundations in population genetic theory, which is now being refined by explicit consideration of the structural constraints placed upon genes encoding proteins through physical chemistry. Further, such models can be examined in the context of comparative genomics, where an integration of gene-level evolution and species-level evolution allows an assessment of the frequency of duplication and the fate of duplicate genes. This process, of course, is dependent upon the biochemical role that duplicated genes play in biological systems, which is in turn dependent upon the mechanism of duplication: whole genome duplication involving a co-duplication of interacting partners vs. single gene duplication. Lastly, the role that these processes may have played in driving speciation is examined.     

An optimized MALDI MSI protocol for spatial detection of tryptic peptides in fresh frozen prostate tissue

MALDI MS imaging (MSI) is a powerful analytical tool for spatial peptide detection in heterogeneous tissues. Proper sample preparation is crucial to achieve high quality, reproducible measurements. Here we developed an optimized protocol for spatially resolved proteolytic peptide detection with MALDI time-of-flight MSI of fresh frozen prostate tissue sections. The parameters tested included four different tissue washes, four methods of protein denaturation, four methods of trypsin digestion (different trypsin densities, sprayers, and incubation times), and five matrix deposition methods (different sprayers, settings, and matrix concentrations). Evaluation criteria were the number of detected and excluded peaks, percentage of high mass peaks, signal-to-noise ratio, spatial localization, and average intensities of identified peptides, all of which were integrated into a weighted quality evaluation scoring system. Based on these scores, the optimized protocol included an ice-cold EtOH+H₂O wash, a 5 min heating step at 95°C, tryptic digestion incubated for 17h at 37°C and CHCA matrix deposited at a final amount of 1.8 μg/mm². Including a heat-induced protein denaturation step after tissue wash is a new methodological approach that could be useful also for other tissue types. This optimized protocol for spatial peptide detection using MALDI MSI facilitates future biomarker discovery in prostate cancer and may be useful in studies of other tissue types.     

Modulation of the central carbon metabolism of Corynebacterium glutamicum improves malonyl-CoA availability and increases plant polyphenol synthesis

In recent years microorganisms have been engineered towards synthesizing interesting plant polyphenols such as flavonoids and stilbenes from glucose. Currently, the low endogenous supply of malonyl-CoA, indispensable for plant polyphenol synthesis, impedes high product titers. Usually, limited malonyl-CoA availability during plant polyphenol production is avoided by supplementing fatty acid synthesis-inhibiting antibiotics such as cerulenin, which are known to increase the intracellular malonyl-CoA pool as a side effect. Motivated by the goal of microbial polyphenol synthesis being independent of such expensive additives, we used rational metabolic engineering approaches to modulate regulation of fatty acid synthesis and flux into the tricarboxylic acid cycle (TCA cycle) in Corynebacterium glutamicum strains capable of flavonoid and stilbene synthesis. Initial experiments showed that sole overexpression of genes coding for the native malonyl-CoA-forming acetyl-CoA carboxylase is not sufficient for increasing polyphenol production in C. glutamicum. Hence, the intracellular acetyl-CoA availability was also increased by reducing the flux into the TCA cycle through reduction of citrate synthase activity. In defined cultivation medium, the constructed C. glutamicum strains accumulated 24 mg·L ⁻¹ (0.088 mM) naringenin or 112 mg·L ⁻¹ (0.49 mM) resveratrol from glucose without supplementation of phenylpropanoid precursor molecules or any inhibitors of fatty acid synthesis.     

p53 Represses the Mevalonate Pathway to Mediate Tumor Suppression

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Importance of infection of haemosporidia blood parasites during different life history stages for long‐term reproductive fitness of collared flycatchers

The interaction between birds and haemosporidia blood parasites is a well‐used system in the study of parasite biology. However, where, when and how parasites are transmitted is often unclear and defining parasite transmission dynamics is essential because of how they influence parasite‐mediated costs to the host. In this study, we used cross‐sectional and longitudinal data taken from a collared flycatcher Ficedula albicollis population to investigate the temporal dynamics of haemosporidia parasite infection and parasite‐mediated costs to host fitness. We investigated host–parasite interactions starting at the nestling stage of the bird's life‐cycle and then followed their progress over three breeding attempts to quantify their fitness – measured as the number of offspring they produced that recruited back into the breeding population. We found that the majority of haemosporidia blood parasite infections occurred within the first year of life and that the most common parasite lineages that infected the breeding population also infected juvenile birds in the natal environment. Moreover, our findings suggest that collared flycatcher nestlings in poorer condition could be at a higher risk of haemosporidia blood parasite infection. In this study, only female and not male bird fitness was adversely affected by parasite infection and the cost of infection on female fitness depended on the timing of transmission. In conclusion, our study indicates that in collared flycatchers, early‐life is potentially important for many of the interactions with haemosporidia parasite lineages, and evidence of parasite‐mediated costs to fitness suggest that these parasites may have influenced the host population dynamics.     

Are tropical coastal reefs sinks or sources of mesozooplankton? A case study in a Brazilian marine protected area

Coral Reefs - In spite of the paramount ecological and socioeconomic relevance of tropical reef ecosystems, the dynamics of their meroplankton abundance remain poorly characterized. The small-scale...     

Transient growth‐enhancing effects of elevated maternal thyroid hormones at no apparent oxidative cost during early postnatal period

Maternal thyroid hormones (THs) have been proven crucial for embryonic development in humans, but their influence within the natural variation on wild animals remains unknown. So far the only two studies that experimentally investigated the potential fitness consequences of maternal THs in birds found inconsistent results. More studies are thus required to assess the general effects of maternal THs and their influences on more behavioral and physiological parameters. In this study, we experimentally elevated yolk TH content in a wild migratory passerine species, the collared flycatcher Ficedula albicollis, to investigate the effects on hatching success, nestling growth and oxidative stress. We found that TH‐injected eggs had a higher hatching success, and the nestlings hatched from TH‐injected eggs were heavier and larger than control nestlings, but only during the early postnatal period. These differences vanished by fledging. Nestlings from TH‐injected eggs exhibited lower activity of the glutathione‐s‐transferase, a major antioxidant enzyme, than control nestlings at day 12, a few days before fledging, but they did not differ in oxidative damage and overall intracellular oxidative state. These results suggest that the early growth‐enhancing effects incurred no observable oxidative stress. We hypothesize that such a transient growth‐enhancing effect might be adaptive in advancing the development and maturation of the offspring so they are well‐prepared in time for the upcoming migration. Further studies investigating whether such advancing effects can influence long‐term fitness, will be more than valuable.