A tale of time and depth: intralacustrine radiation in endemic Gammarus species flock from the ancient Lake Ohrid

Tentatively dated, the Plio‐/Pleistocene origin of the ancient Lake Ohrid on the Balkan Peninsula makes it the oldest ancient lake in Europe. Given the surface area of the lake and the adjusted endemicity rate, it may be also defined as the most diverse of all the ancient lakes in the world. From all the animal groups endemic to this lake, gammarids are amongst the most scarcely known in terms of their diversity and phylogenetic relationships. Partial DNA sequences of two mitochondrial genes, cytochrome oxidase subunit I (cox1) and 16S ribosomal RNA (16S rRNA) of eight known endemic Gammarus species from the Lake Ohrid valley were analysed. Phylogenetic analyses showed that endemic Gammarus species comprise an ancient species flock, with Gammarus sketi from the feeder springs being their sister taxon outside the lake. Amongst the species inhabiting the lake, Gammarus solidus and Gammarus salemaai are morphologically and molecularly well defined. By contrast, Gammarus ochridensis, Gammarus parechiniformis, Gammarus lychnidensis, and Gammarus stankokaramani revealed high discrepancy between morphological and genetic data. None of these morphospecies form a monophyletic clade and a significant degree of apparent gene flow occurs between them. This could be caused by incomplete lineage sorting and/or hybridization events. Two novel mtDNA lineages were found within the lake, possibly constituting two new species (Gammarus sp. 1 and Gammarus sp. 2). Molecular clock analysis showed that the split between G. sketi and the Gammarus species flock from the lake occurred approximately 5–7 Mya, whereas within the flock there were at least two intralacustrine radiations: one estimated at 2–3 Mya and the second at less than 1 Mya. The first one could be associated with the origin of the lake and the second with the lake water‐level fluctuations during Pleistocene. © 2013 The Linnean Society of London     

Subfamilies of CR1 non-LTR retrotransposons have different 5'UTR sequences but are otherwise conserved

CR1 elements and CR1-related (CR1-like) elements are a novel family of non-LTR retrotransposons that are found in all vertebrates (reptilia, amphibia, fish, and mammals), whereas more distantly related elements are found in several invertebrate species. CR1 elements have several features that distinguish them from other non-LTR retrotransposons. Most notably, their 3' termini lack a polyadenylic acid (poly A) tail and instead contain 2-4 copies of a unique 8 bp repeat. CR1 elements are present at approximately 100,000 copies in the chicken genome. The vast majority of these elements are severely 5' truncated and mutated; however, six subfamilies (CR1-A through CR1-F) are resolved by sequence comparisons. One of these subfamilies (i.e. CR1-B) previously was analyzed in detail. In the present study, we identified several full-length elements from the CR1-F subfamily. Although regions within the open reading frames and 3' untranslated regions of CR1-F and CR1-B elements are well conserved, their respective 5' untranslated regions are unrelated. Thus, our results suggest that new CR1 subfamilies form when elements with intact open reading frames acquire new 5' UTRs, which could, in principle, function as promoters.     

Using MT(-/-) mice to study metallothionein and oxidative stress

Mice with null mutations for metallothionein genes MT-1 and MT-2 were used to study the role that metallothionein plays in protecting cellular targets in vivo from oxidative stress. Wild-type (MT(+/+)) and MT-null (MT(-/-)) mice were treated with either saline or zinc and exposed to two types of oxidative stress: gamma-irradiation or 2-nitropropane. There was no alteration in the antioxidant defense system (superoxide dismutase, catalase, or glutathione peroxidase and glutathione levels) to compensate for the lack of the metallothionein in the MT(-/-) mice. The amount of oxidative damage to liver DNA, lipids, and proteins were similar for the MT(-/-) and MT(+/+) mice even though the levels of metallothionein in the livers of the saline- or zinc-pretreated MT(+/+) mice were 5- to 100-fold greater than found in the MT(-/-) mice. To determine if metallothionein can protect mice from the lethal effects of ionizing radiation, the mean survivals of MT(-/-) and MT(+/+) mice exposed to whole body gamma-irradiation were measured and found to be similar. However, the mean survival increased significantly after zinc pretreatment for both the MT(-/-) and MT(+/+) mice. These results demonstrate that tissue levels of metallothionein do not protect mice in vivo against oxidative stress.     

Fluorescence probes in biochemistry: an examination of the non-fluorescent behavior of dansylamide by photoacoustic calorimetry.

Photoacoustic calorimetry is shown to be a simple, precise, and accurate method for the quantification of the photophysics of a fluorescence probe, e.g., dansylamide, in a variety of solvents. This technique, which is described in detail, provides a direct measurement of the energy that is released nonradiatively following photostimulation, and can therefore be used to indirectly determine the amount of energy released via luminescent pathways. Photoacoustic calorimetry combined with established absorption and fluorescence methodologies provides a complete arsenal for characterizing the photophysical properties of many systems. Comparison of the photoacoustic signal for dansylamide versus standard compounds (ferrocene, tetraphenylethylene, 8-anilinonaphthalene-1-sulfonate, and/or 5,5'-dithiobis(2-nitrobenzoic acid) in 12 different solvents gave fh values (fraction of each absorbed 337.1-nm photon returned as heat) from a low of 0.530 in 1,4-dioxane to a high of 0.973 in water. The trend noted with solvent polarity is different and more revealing than that determined by the more classical approach of examining either the wavelength of the emission maximum or the fluorescence quantum yield.     

Posttranslational processing of human lysosomal acid beta-glucosidase: a continuum of defects in Gaucher disease type 1 and type 2 fibroblasts.

The major processing steps in the maturation of the lysosomal hydrolase, acid beta-glucosidase, were examined in fibroblasts from normal individuals and from patients with types 1 and 2 Gaucher disease. In pulse-chase studies with normal fibroblasts, remodeling of N-linked oligosaccharides resulted in the temporal appearance of three molecular-weight forms of acid beta-glucosidase. An initial 64-kDa form, containing high mannose-type oligosaccharide side chains, was processed quantitatively, within 24 h, to a sialylated 69-kDa form. During the subsequent 96 h, some of the 69-kDa form is processed to 59 kDa. Glycosidase digestion studies revealed that the increase in the apparent molecular weight of the normal enzyme from 64 kDa to 69 kDa resulted primarily from the addition to sialic acid residues in the Golgi apparatus. The polypeptide backbone of both the 64-kDa and 69-kDa forms was 55.3 kDa. Processing of acid beta-glucosidase in fibroblasts from three of four type 1 (nonneuronopathic) Ashkenazi Jewish Gaucher disease patients was nearly normal. With fibroblasts from one Ashkenazi Jewish and three non-Jewish type 1 as well as from two type 2 (acute neuronopathic) Gaucher disease patients, only a 64-kDa form of acid beta-glucosidase was detected. Inefficient and incomplete processing to the 69-kDa form was found in one type 2 cell line (GM2627). These results indicate that no firm correlation exists between the type or degree of abnormal processing of acid beta-glucosidase in fibroblasts and the phenotype of Gaucher disease.     

HIV Shedding from Male Circumcision Wounds in HIV-Infected Men: A Prospective Cohort Study

BackgroundA randomized trial of voluntary medical male circumcision (MC) of HIV—infected men reported increased HIV transmission to female partners among men who resumed sexual intercourse prior to wound healing. We conducted a prospective observational study to assess penile HIV shedding after MC.ConclusionPenile HIV shedding is significantly reduced after healing of MC wounds. Lower plasma VL is associated with decreased frequency and quantity of HIV shedding from MC wounds. Starting ART prior to MC should be considered to reduce male-to-female HIV transmission risk. Research is needed to assess the time on ART required to decrease shedding, and the acceptability and feasibility of initiating ART at the time of MC.     

Activation of p38 Mitogen-Activated Protein Kinase in Gaucher’s Disease

Gaucher’s disease is caused by defects in acid β-glucosidase 1 (GBA1) and has been also proposed as an inflammatory disease. GBA1 cleaves glucosylceramide to form ceramide, an established bioactive lipid, and defects in GBA1 lead to aberrant accumulation in glucosylceramide and insufficient formation of ceramide. We investigated if the pro-inflammatory kinase p38 is activated in Gaucher’s disease, since ceramide has been proposed to suppress p38 activation. Three Gaucher’s disease mouse models were employed, and p38 was found to be activated in lung and liver tissues of all Gaucher’s disease mice. Most interestingly, neuronopathic Gaucher’s disease type mice, but not non-neuronopathic ones, displayed significant activation of p38 and up-regulation of p38-inducible proinflammatory cytokines in brain tissues. In addition, all type of Gaucher’s disease mice also showed increases in serum IL-6. As cellular signalling is believed to represent an in vivo inflammatory phenotype in Gaucher’s disease, activation of p38 and possibly its-associated formation of proinflammatory cytokines were assessed in fibroblasts established from neuronopathic Gaucher’s disease mice. In mouse Gaucher’s disease cells, p38 activation and IL-6 formation by TNF-α treatment were enhanced as compared to those of wild type. Furthermore, human fibroblasts from Gaucher’s disease patients also displayed increases in p38 activation and IL-6 formation as comparison to healthy counterpart. These results raise the potential that proinflammatory responses such as p38 activation and IL-6 formation are augmented in Gaucher’s disease.