Independent duplications of the acetylcholinesterase gene conferring insecticide resistance in the mosquito Culex pipiens

Gene duplication is thought to be the main potential source of material for the evolution of new gene functions. Several models have been proposed for the evolution of new functions through duplication, most based on ancient events (Myr). We provide molecular evidence for the occurrence of several (at least 3) independent duplications of the ace-1 locus in the mosquito Culex pipiens, selected in response to insecticide pressure that probably occurred very recently (<40 years ago). This locus encodes the main target of several insecticides, the acetylcholinesterase. The duplications described consist of 2 alleles of ace-1, 1 susceptible and 1 resistant to insecticide, located on the same chromosome. These events were detected in different parts of the world and probably resulted from distinct mechanisms. We propose that duplications were selected because they reduce the fitness cost associated with the resistant ace-1 allele through the generation of persistent, advantageous heterozygosis. The rate of duplication of ace-1 in C. pipiens is probably underestimated, but seems to be rather high.     

Gene expression specificity of the mussel antifungal mytimycin (MytM)

We previously reported the nucleotide sequences and diversity of mytimycin (MytM) from the Mediterranean mussel, Mytilus galloprovincialis. Using real-time PCR (q-PCR), we observed that the MytM gene was mainly expressed in circulating hemocytes and to a less extent in the mantle. In vivo challenge with bacteria or with the yeast, Candida albicans, did not increase the expression as measured by q-PCR in hemocytes. By contrast, injection of the filamentous fungus, Fusarium oxysporum, induced a sudden and strong increase of expression at 9h p.i. (stimulation index of 25.7 ± 2.1). Optimum stimulating dose was 10⁴ spores of F. oxysporum per mussel. In the same samples, AMP mytilin and myticin showed no stimulation. Consequently, we hypothesized the existence of 2 different signal transduction pathways, one activated by bacteria and yeast, the other triggered by filamentous fungi. A second challenge performed with F. oxysporum 24 h after the first challenge induced an increase of MytM gene expression (stimulation index of 3.5 ± 1.7). However, this second increase was significantly lower than the first, suggesting less efficient response rather than significant protection.     

Wolbachia Divergence and the Evolution of Cytoplasmic Incompatibility in Culex pipiens

Many insect species harbor Wolbachia bacteria that induce cytoplasmic incompatibility (CI), i.e. embryonic lethality in crosses between infected males and uninfected females, or between males and females carrying incompatible Wolbachia strains. The molecular mechanism of CI remains unknown, but the available data are best interpreted under a modification–rescue model, where a mod function disables the reproductive success of infected males’ sperm, unless the eggs are infected and express a compatible resc function. Here we examine the evolution of CI in the mosquito Culex pipiens, harbouring a large number of closely related Wolbachia strains structured in five distinct phylogenetic groups. Specifically, we used a worldwide sample of mosquito lines to assess the hypothesis that genetic divergence should correlate with the divergence of CI properties on a low evolutionary scale. We observed a significant association of Wolbachia genetic divergence with CI patterns. Most Wolbachia strains from the same group were compatible whereas those from different groups were often incompatible. Consistently, we found a strong association between Wolbachia groups and their mod-resc properties. Finally, lines from the same geographical area were rarely incompatible, confirming the conjecture that the spatial distribution of Wolbachia compatibility types should be constrained by selection. This study indicates a clear correlation between Wolbachia genotypes and CI properties, paving the way toward the identification of the molecular basis of CI through comparative genomics.     

The Relationship between Muscle Fiber Type-Specific PGC-1α Content and Mitochondrial Content Varies between Rodent Models and Humans

PGC-1α regulates critical processes in muscle physiology, including mitochondrial biogenesis, lipid metabolism and angiogenesis. Furthermore, PGC-1α was suggested as an important regulator of fiber type determination. However, whether a muscle fiber type-specific PGC-1α content exists, whether PGC-1α content relates to basal levels of mitochondrial content, and whether such relationships are preserved between humans and classically used rodent models are all questions that have been either poorly addressed or never investigated. To address these issues, we investigated the fiber type-specific content of PGC-1α and its relationship to basal mitochondrial content in mouse, rat and human muscles using in situ immunolabeling and histochemical methods on muscle serial cross-sections. Whereas type IIa fibers exhibited the highest PGC-1α in all three species, other fiber types displayed a hierarchy of type IIx>I>IIb in mouse, type I = IIx> IIb in rat, and type IIx>I in human. In terms of mitochondrial content, we observed a hierarchy of IIa>IIx>I>IIb in mouse, IIa >I>IIx> IIb in rat, and I>IIa> IIx in human skeletal muscle. We also found in rat skeletal muscle that type I fibers displayed the highest capillarization followed by type IIa >IIx>IIb. Finally, we found in human skeletal muscle that type I fibers display the highest lipid content, followed by type IIa>IIx. Altogether, our results reveal that (i) the fiber type-specific PGC-1α and mitochondrial contents were only matched in mouse, (ii) the patterns of PGC-1α and mitochondrial contents observed in mice and rats do not correspond to that seen in humans in several respects, and (iii) the classical phenotypes thought to be regulated by PGC-1α do not vary exclusively as a function of PGC-1α content in rat and human muscles.     

Ocular application of the kinin B1 receptor antagonist LF22-0542 inhibits retinal inflammation and oxidative stress in streptozotocin-diabetic rats

Purpose

Kinin B₁ receptor (B₁R) is upregulated in retina of Streptozotocin (STZ)-diabetic rats and contributes to vasodilation of retinal microvessels and breakdown of the blood-retinal barrier. Systemic treatment with B₁R antagonists reversed the increased retinal plasma extravasation in STZ rats. The present study aims at determining whether ocular application of a water soluble B₁R antagonist could reverse diabetes-induced retinal inflammation and oxidative stress.

Methods

Wistar rats were made diabetic with STZ (65 mg/kg, i.p.) and 7 days later, they received one eye drop application of LF22-0542 (1% in saline) twice a day for a 7 day-period. The impact was determined on retinal vascular permeability (Evans blue exudation), leukostasis (leukocyte infiltration using Fluorescein-isothiocyanate (FITC)-coupled Concanavalin A lectin), retinal mRNA levels (by qRT-PCR) of inflammatory (B₁R, iNOS, COX-2, ICAM-1, VEGF-A, VEGF receptor type 2, IL-1β and HIF-1α) and anti-inflammatory (B₂R, eNOS) markers and retinal level of superoxide anion (dihydroethidium staining).

Results

Retinal plasma extravasation, leukostasis and mRNA levels of B₁R, iNOS, COX-2, VEGF receptor type 2, IL-1β and HIF-1α were significantly increased in diabetic retinae compared to control rats. All these abnormalities were reversed to control values in diabetic rats treated with LF22-0542. B₁R antagonist also significantly inhibited the increased production of superoxide anion in diabetic retinae.

Conclusion

B₁R displays a pathological role in the early stage of diabetes by increasing oxidative stress and pro-inflammatory mediators involved in retinal vascular alterations. Hence, topical application of kinin B₁R antagonist appears a highly promising novel approach for the treatment of diabetic retinopathy.     

Myostatin inactivation increases myotube size through regulation of translational initiation machinery

Myostatin deficiency leads in skeletal muscle overgrowth but the precise molecular mechanisms underlying this hypertrophy are not well understood. In this study, to gain insight into the role of endogenous myostatin in the translational regulation, we used an in vitro model of cultured satellite cells derived from myostatin knock‐out mice. Our results show that myostatin knock‐out myotubes are larger than control myotubes and that this phenotype is associated with an increased activation of the Akt/mTOR signaling pathway, a known regulator of muscle hypertrophy. These results demonstrate that hypertrophy due to myostatin deficiency is preserved in vitro and suggest that myostatin deletion results in an increased protein synthesis. Accordingly, the rates of global RNA content, polysome formation and protein synthesis are all increased in myostatin‐deficient myotubes while they are counteracted by the addition of recombinant myostatin. We furthermore demonstrated that genetic deletion of myostatin stimulates cap‐dependent translation by positively regulating assembly of the translation preinitiation complex. Together the data indicate that myostatin controls muscle hypertrophy in part by regulating protein synthesis initiation rates, that is, translational efficiency. J. Cell. Biochem. 112: 3531–3542, 2011. © 2011 Wiley Periodicals, Inc.     

The ultrathin structure of amoeboid flagellate <Emphasis Type="Italic">Thaumatomastix</Emphasis> sp. (Thaumatomonadida (Shirkina) Karpov, 1990)

The ultrathin structure of amoeboid flagellate Thaumatomastix sp. is considered. The cell is surrounded by two-layered triangular scales. They are formed on the surface of mitochondria. Pseudopodia grabbing bacteria run from ventricular furrow, which is armored with two longitudinal bands of microtubules. Heterodynamic flagella run from small flagellar pocket. Long back flagellum has thin mastigonemes. Proximal area of short flagellum is covered with flat oval scales. Transitional flagellant zone has no spiral or other additional elements. Transverse plate is localized above cell surface. Kinetosomes are parallel to each other. Vesicular nucleus and Golgi apparatus have typical structure. Oval mitochondria contain tubular cristae. Within cytoplasm, extrusive organelles (kinetocysts) containing amorphous material and capsule were found. The latter consists of muff and cylinder. Plasmodial and cystic phases of development have not been discovered. Contractile vacuole is absent. The resemblance between Thaumatomastix sp. and other thaumatomonads has been discussed.     

Diversity of Coding Sequences and Gene Structures of the Antifungal Peptide Mytimycin (MytM) from the Mediterranean Mussel, <Emphasis Type="Italic">Mytilus galloprovincialis</Emphasis>

Knowledge on antifungal biomolecules is limited compared to antibacterial peptides. A strictly antifungal peptide from the blue mussel, Mytilus edulis named mytimycin (MytM) was reported in 1996 as partial NH₂ 33 amino acid sequence. Using back-translations of the previous sequence, MytM-related nucleotide sequences were identified from a normalized Mytilus galloprovincialis expressed sequence tag library. Primers designed from a consensus sequence have been used to obtain a fragment of 560 nucleotides, including the complete coding sequence of 456 nucleotides. Precursor is constituted by a signal peptide of 23 amino acids, followed by MytM of 54 amino acids (6.2–6.3 kDa, 12 cysteines) and C-terminal extension of 75 amino acids. Only two major amino acid precursor sequences emerged, one shared by M. galloprovincialis from Venice and Vigo, the other belonging to M. galloprovincialis from Palavas, with nine amino acid differences between the two MytM. Predicted disulfide bonds suggested the presence of two constrained domains joined by amino acidic NIFG track. Intriguing was the presence of conserved canonical EF hand-motif located in the C-terminus extension of the precursor. The MytM gene was found interrupted by two introns. Intron 2 existed in two forms, a long (1,112 nucleotides) and a short (716 nucleotides) one resulting from the removal of the central part of the long one. Both the short (GenBank FJ804479) and the long (GenBank FJ804478) genes are simultaneously present in the mussel genome.     

Effect of sarcopenia on cardiovascular disease risk factors in obese postmenopausal women

Objective: To compare sarcopenic‐obese and obese postmenopausal women for risk factors predisposing to cardiovascular disease (CVD) and determine whether there may be a relationship between muscle mass and metabolic risk in obese postmenopausal women. Research Methods and Procedures: In this cross‐sectional study, 22 healthy obese postmenopausal women (mean age, 66 ± 5 years; mean BMI, 27 ± 3 kg/m²) were divided into two groups matched for age (±2 years) and fat mass (FM) (±2%). Sarcopenia was defined as a muscle mass index of <14.30 kg fat‐free mass (FFM)/m² (which corresponds to 1 standard deviation below the values of a young reference population), and obesity was defined as an FM of >35% (which corresponds to the World Health Organization guidelines). FM, FFM (measured by DXA), daily energy expenditure (accelerometry), dietary intake (3‐day dietary record), and blood biochemical analyses (lipid profile, insulin, glucose, and C‐reactive protein) were obtained. Visceral fat mass (VFM) was calculated by the equation of Bertin, which estimates VFM from DXA measurements. Results: Obese women had more FFM (p = 0.006), abdominal FM (p = 0.047), and VFM (p = 0.041) and a worse lipid profile [p = 0.040 for triglycerides; p = 0.004 for high‐density lipoprotein (HDL); p = 0.026 for total cholesterol/HDL] than sarcopenic‐obese postmenopausal women. Obese women also ingested significantly more animal (p = 0.001) and less vegetal proteins (p = 0.013), although both groups had a similar total protein intake (p = 0.967). Discussion: Sarcopenia seems to be associated with lower risk factors predisposing to CVD in obese postmenopausal women. With the increase in the number of aging people, the health implications of being sarcopenic‐obese merit more attention.