Vertical migration of aggregated aerobic and anaerobic ammonium oxidizers enhances oxygen uptake in a stagnant water layer

Ammonium can be removed as dinitrogen gas by cooperating aerobic and anaerobic ammonium-oxidizing bacteria (AerAOB and AnAOB). The goal of this study was to verify putative mutual benefits for aggregated AerAOB and AnAOB in a stagnant freshwater environment. In an ammonium fed water column, the biological oxygen consumption rate was, on average, 76 kg O₂ ha⁻¹ day⁻¹. As the oxygen transfer rate of an abiotic control column was only 17 kg O₂ ha⁻¹ day⁻¹, biomass activity enhanced the oxygen transfer. Increasing the AnAOB gas production increased the oxygen consumption rate with more than 50% as a result of enhanced vertical movement of the biomass. The coupled decrease in dissolved oxygen concentration increased the diffusional oxygen transfer from the atmosphere in the water. Physically preventing the biomass from rising to the upper water layer instantaneously decreased oxygen and ammonium consumption and even led to the occurrence of some sulfate reduction. Floating of the biomass was further confirmed to be beneficial, as this allowed for the development of a higher AerAOB and AnAOB activity, compared to settled biomass. Overall, the results support mutual benefits for aggregated AerAOB and AnAOB, derived from the biomass uplifting effect of AnAOB gas production.     

Endonuclease G: a mitochondrial protein released in apoptosis and involved in caspase-independent DNA degradation.

A hallmark of apoptosis is the fragmentation of nuclear DNA. Although this activity involves the caspase-3-dependent DNAse CAD (caspase-activated DNAse), evidence exists that DNA fragmentation can occur independently of caspase activity. Here we report on the ability of truncated Bid (tBid) to induce the release of a DNAse activity from mitochondria. This DNAse activity was identified by mass spectrometry as endonuclease G, an abundant 30 kDa protein released from mitochondria under apoptotic conditions. No tBid-induced endonuclease G release could be observed in mitochondria from Bcl-2-transgenic mice. The in vivo occurrence of endonuclease G release from mitochondria during apoptosis was confirmed in the liver from mice injected with agonistic anti-Fas antibody and is completely prevented in Bcl-2 transgenic mice. These data indicate that endonuclease G may be involved in CAD-independent DNA fragmentation during cell death pathways in which truncated Bid is generated.     

PINK1 Kinase Catalytic Activity Is Regulated by Phosphorylation on Serines 228 and 402

Mutations in the PINK1 gene cause early-onset recessive Parkinson disease. PINK1 is a mitochondrially targeted kinase that regulates multiple aspects of mitochondrial biology, from oxidative phosphorylation to mitochondrial clearance. PINK1 itself is also phosphorylated, and this might be linked to the regulation of its multiple activities. Here we systematically analyze four previously identified phosphorylation sites in PINK1 for their role in autophosphorylation, substrate phosphorylation, and mitophagy. Our data indicate that two of these sites, Ser-228 and Ser-402, are autophosphorylated on truncated PINK1 but not on full-length PINK1, suggesting that the N terminus has an inhibitory effect on phosphorylation. We furthermore establish that phosphorylation of these PINK1 residues regulates the phosphorylation of the substrates Parkin and Ubiquitin. Especially Ser-402 phosphorylation appears to be important for PINK1 function because it is involved in Parkin recruitment and the induction of mitophagy. Finally, we identify Thr-313 as a residue that is critical for PINK1 catalytic activity, but, in contrast to previous reports, we find no evidence that this activity is regulated by phosphorylation. These data clarify the regulation of PINK1 through multisite phosphorylation.     

Reflectance spectra and mating patterns support intraspecific mimicry in the colour polymorphic damselfly <Emphasis Type="Italic">Ischnura elegans</Emphasis>

Coexistence of female colour morphs in animal populations is often considered the result of sexual conflict, where polymorphic females benefit from reduced male sexual harassment. Mate-searching males easily detect suitable partners when only one type of female is present, but become challenged when multiple female morphs coexist, which may result in frequency-dependent mate preferences. Intriguingly, in damselflies, one female morph often closely resembles the conspecific male in body coloration, which has lead to hypotheses regarding intra-specific male-mimicry. However, few studies have quantitatively evaluated the correspondence between colour reflectance spectra from males and male-like females, relying instead on qualitative visual assessments of coloration. Using colour analyses of reflectance spectra, we compared characteristics of the body coloration of ontogenetic male and female colour morphs of the damselfly Ischnura elegans. In addition, we evaluated whether males appear to (1) discriminate between immature and mature female colour morphs, and (2) whether male-like females experience reduced male mating attention and low mating frequencies as predicted from male-mimicry. Spectral reflectance data show that immature female morphs differ substantially in coloration from mature individuals. Mating frequencies were much lower for immature than mature female morphs. For the male-like female morph, measures of colour were statistically indistinguishable from that of both immature and mature conspecific males. Mating frequencies of male-like females were lower than those of other mature female morphs under field and experimental conditions. Together, our results indicate that males may use the observed spectral differences in mate choice decisions. Furthermore, male-like females may be regarded as functional mimics that have reduced attractiveness and lowered rates of sexual harassment by mate-searching males.     

In vivo relevance of citrullinated proteins and the challenges in their detection

Citrullination is a posttranslational modification of arginine. It plays both a physiological role, for instance during apoptosis and epigenetics, and a pathological role in cancer or diseases of the central nervous system. Most research on citrullination to date focuses on its role in auto-immune diseases such as multiple sclerosis and rheumatoid arthritis. In this context, the exact knowledge of citrullination sites in a protein can provide invaluable information about the etiological importance of these citrullinated proteins. However, few techniques exist that can accurately detect citrullination on the peptide level. This review aims to give an overview of the different methods available to date for the detection of citrullinated proteins and peptides. These include 2D-SDS-PAGE and immunodetection, as well as specific mass spectrometry (MS) approaches, both labeled and unlabeled. These MS approaches have been developed to pinpoint the exact location of citrullination on the peptide level. Improving the currently existing detection strategies while focusing on the role of citrullinated proteins will be invaluable to elucidate the importance of this posttranslational modification in vivo.     

Extracting histones for the specific purpose of label‐free MS

Extracting histones from cells is the first step in studies that aim to characterize histones and their post‐translational modifications (hPTMs) with MS. In the last decade, label‐free quantification is more frequently being used for MS‐based histone characterization. However, many histone extraction protocols were not specifically designed for label‐free MS. While label‐free quantification has its advantages, it is also very susceptible to technical variation. Here, we adjust an established histone extraction protocol according to general label‐free MS guidelines with a specific focus on minimizing sample handling. These protocols are first evaluated using SDS‐PAGE. Hereafter, a selection of extraction protocols was used in a complete histone workflow for label‐free MS. All protocols display nearly identical relative quantification of hPTMs. We thus show that, depending on the cell type under investigation and at the cost of some additional contaminating proteins, minimizing sample handling can be done during histone isolation. This allows analyzing bigger sample batches, leads to reduced technical variation and minimizes the chance of in vitro alterations to the hPTM snapshot. Overall, these results allow researchers to determine the best protocol depending on the resources and goal of their specific study. Data are available via ProteomeXchange with identifier PXD002885.     

Vegetation development and human occupation in the Damascus region of southwestern Syria from the Late Pleistocene to Holocene

Rockshelter Baaz in the Damascus region of Syria provided a variety of botanical remains from the Late Pleistocene and Early Holocene period. These remains provide new information about the vegetation evolution in this region. The earliest occupational levels correspond with a moisture peak during the Late Pleistocene, between ca. 34–32 kyr b.p., when pine expanded. The next occupations took place during extreme arid conditions, ca. 23–21 kyr b.p., and probably during the Last Glacial Maximum when a steppe vegetation was established. The occupation level of the Younger Dryas, represented by Natufian remains, suggests that the area had been covered by almond-pistachio steppe, similar to later periods of the Early Holocene, and was probably located just outside the range of dense wild cereal stands. There is no drastic impact of the Younger Dryas visible on the vegetation in the botanical remains. The lack of fruits and seeds at Baaz indicates that the site was more likely to have been a temporary hunting post rather than a plant processing site for much of its history. It is ideally suited to this purpose because of its location over the Jaba′deen Pass and the associated springs. However, archaeological remains from the Natufian period, suggest that the site was more permanently occupied during this time. A. Dodonov deceased.     

Highly efficient peptide separations in proteomics: Part 2: Bi- and multidimensional liquid-based separation techniques

Multidimensional liquid-based separation techniques are described for maximizing the resolution of the enormous number of peptides generated upon tryptic digestion of proteomes, and hence, reduce the spatial and temporal complexity of the sample to a level that allows successful mass spectrometric analysis. This review complements the previous contribution on unidimensional high performance liquid chromatography (HPLC). Both chromatography and electrophoresis will be discussed albeit with reversed-phase HPLC (RPLC) as the final separation dimension prior to MS analysis.     

Llama antibody fragments with cross-subtype human immunodeficiency virus type 1 (HIV-1)-neutralizing properties and high affinity for HIV-1 gp120

Members of the Camelidae family produce immunoglobulins devoid of light chains. We have characterized variable domains of these heavy chain antibodies, the VHH, from llamas immunized with human immunodeficiency virus type 1 (HIV-1) envelope protein gp120 in order to identify VHH that can inhibit HIV-1 infection. To increase the chances of isolating neutralizing VHH, we employed a functional selection approach, involving panning of phage libraries expressing the VHH repertoire on recombinant gp120, followed by a competitive elution with soluble CD4. By immunizing with gp120 derived from an HIV-1 subtype B′/C primary isolate, followed by panning on gp120 from HIV-1 isolates of subtypes A, B, and C, we could select for VHH with cross-subtype neutralizing activity. Three VHH able to neutralize HIV-1 primary isolates of subtypes B and C were characterized. These bound to recombinant gp120 with affinities close to the suggested affinity ceiling for in vivo-maturated antibodies and competed with soluble CD4 for this binding, indicating that their mechanism of neutralization involves interacting with the functional envelope spike prior to binding to CD4. The most potent VHH in terms of low 50% inhibitory concentration (IC₅₀) and IC₉₀ values and cross-subtype reactivity was A12. These results indicate that camelid VHH can be potent HIV-1 entry inhibitors. Since VHH are stable and can be produced at a relatively low cost, they may be considered for applications such as HIV-1 microbicide development. Antienvelope VHH might also prove useful in defining neutralizing and nonneutralizing epitopes on HIV-1 envelope proteins, with implications for HIV-1 vaccine design.