Deformation of dynamin helices damped by membrane friction

Dynamin and other proteins of the dynamin superfamily are widely used by cells to sever lipid bilayers. During this process, a short helical dynamin polymer (one to three helical turns) assembles around a membrane tubule and reduces its radius and pitch upon guanosine triphosphate hydrolysis. This deformation is thought to be crucial for dynamin's severing action and results in an observable twisting of the helix. Here, we quantitatively characterize the dynamics of this deformation by studying long dynamin helices (many helical turns). We perform in vitro experiments where we attach small beads to the dynamin helix and track their rotation in real time, thus collecting information about the space and time dependence of the deformation. We develop a theoretical formalism to predict the dynamics of a mechanically continuous helix deforming on long timescales. Longer helices deform more slowly, as predicted by theory. This could account for the previously reported observation that they are less fission-competent. Comparison between experiments and our model indicates that the deformation dynamics is dominated by the draining of the membrane out of the helix, allowing quantification of helix-membrane interactions.     

Exploring the difficulties of studying futures in ecology: what do ecological scientists think?

In a mainly experimental science based traditionally on hypothesis testing such as ecology, studying futures may be difficult. However, in the last few decades, predicting the consequences of global changes on the dynamics and function of ecological systems has become a major challenge in ecological research. To study how ecological scientists deal with potential difficulties in studying futures, we adopted a reflexive viewpoint on how scientists address the study of ecological futures. To do so we questioned a panel of ecological scientists on their practical involvement and point of view. Quantitative and qualitative analyses of their responses showed that predictions or predictive models were the dominant theme. Many quantitative models, based on statistical correlations, empirical rules or processes have been developed and their methodological limitations explored by the researchers we interviewed. In a small proportion of studies, qualitative scenarios have been elaborated to explore the range of possible futures. Interviewees emphasized the problem of dealing with ecological complexity and multiple future possibilities. Specificities of futures compared to past or present events were not fully identified. In fact, researchers studying futures mainly adopted a reductionist approach, trying to simplify complex ecological systems. But methods and tools promoted by such an approach to science may not always be appropriate to deal with future ecological complexity. Indeed, an emphasis on prediction prevents ecologists from acknowledging the multiplicity and undetermined nature of futures.     

Structural and antimicrobial properties of human pre-elafin/trappin-2 and derived peptides against <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis>

Background  

Pre-elafin/trappin-2 is a human innate defense molecule initially described as a potent inhibitor of neutrophil elastase. The full-length protein as well as the N-terminal "cementoin" and C-terminal "elafin" domains were also shown to possess broad antimicrobial activity, namely against the opportunistic pathogen P. aeruginosa. The mode of action of these peptides has, however, yet to be fully elucidated. Both domains of pre-elafin/trappin-2 are polycationic, but only the structure of the elafin domain is currently known. The aim of the present study was to determine the secondary structures of the cementoin domain and to characterize the antibacterial properties of these peptides against P. aeruginosa.     

Systemic vasculopathy with altered vasoreactivity in a transgenic mouse model of scleroderma

Introduction  

Vasculopathy, including altered vasoreactivity and abnormal large vessel biomechanics, is a hallmark of systemic sclerosis (SSc). However, the pathogenic link with other aspects of the disease is less clear. To assess the potential role of transforming growth factor beta (TGF-β) overactivity in driving these cardiovascular abnormalities, we studied a novel transgenic mouse model characterized by ligand-dependent activation of TGF-β signaling in fibroblasts.     

Differential expression level of cytokeratin 8 in cells of the bovine nucleus pulposus complicates the search for specific intervertebral disc cell markers

Introduction  

Development of cell therapies for repairing the intervertebral disc is limited by the lack of a source of healthy human disc cells. Stem cells, particularly mesenchymal stem cells, are seen as a potential source but differentiation strategies are limited by the lack of specific markers that can distinguish disc cells from articular chondrocytes.     

Gender affects liver desaturase expression in a rat model of n−3 fatty acid repletion

Dietary n?3 polyunsaturated fatty acids (PUFA) are major components of cell membranes and have beneficial effects on human health. Docosahexaenoic acid (DHA; 22:6n?3) is the most biologically important n?3 PUFA and can be synthesized from its dietary essential precursor, α-linolenic acid (ALA; 18:3n?3). Gender differences in the efficiency of DHA bioconversion have been reported, but underlying molecular mechanisms are unknown. We compared the capacity for DHA synthesis from ALA and the expression of related enzymes in the liver and cerebral cortex between male and female rats. Wistar rats, born with a low-DHA status, were supplied with a suboptimal amount of ALA from weaning to 8 weeks of age. Fatty acid composition was determined by gas chromatography, the mRNA expression of different genes involved in PUFA metabolism was determined by RT-PCR (low-density array) and the expression of proteins was determined by Western blot analysis. At 8 weeks, DHA content was higher (+20 to +40%) in each phospholipid class of female livers compared to male livers. The “Δ4,” Δ5 and Δ6 desaturation indexes were 1.2–3 times higher in females than in males. The mRNA expression of Δ5- and Δ6-desaturase genes was 3.8 and 2.5 times greater, respectively, and the Δ5-desaturase protein was higher in female livers (+50%). No gender difference was observed in the cerebral cortex. We conclude that female rats replete their DHA status more readily than males, probably due to a higher expression of liver desaturases. Our results support the hypothesis on hormonal regulation of PUFA metabolism, which should be taken into account for specific nutritional recommendations.     

Transrectal ultrasonography and plasma progestin profiles identifies feto-placental compromise in mares with experimentally induced placentitis

Transrectal ultrasonography of the caudal uterus and a progestin profile were evaluated for accuracy in identifying mares with feto-placental compromise in a model of placentitis. Twenty-two pregnant ponies were divided into four groups: (1) control mares (n=5); (2) instrumented controls (n=2); (3) instrumented inoculated mares (n=11); (4) inoculated mares (n=4). Mares in Groups 3 and 4 were inoculated with Streptococcus equi subsp. zooepidemicus. Maternal plasma progestins, vulvar discharge, mammary gland development, combined thickness of the uterus and placenta (CTUP) and placental separation were evaluated weekly before instrumentation, inoculation or Day 320 (Groups 1 and 2) and, thereafter, either daily (first three measurements) or several times weekly (last two measurements). Plasma progestin profiles were plotted to identify pattern characteristics. An abbreviated profile was created, consisting of four progestin samples collected at 48-h intervals, with Sample 1 collected the day before inoculation or on Day 285 in controls. Profiles were considered abnormal if Samples 2, 3, or 4 increased or decreased by more than 50% of Sample 1. A CTUP>1.0 cm or placental separation were considered abnormal. Placentitis was confirmed by histology of fetal membranes. Control mares had normal progestin profiles, transrectal ultrasonographic and clinical examinations. Control foals were born after Day 329; six were viable and one died after dystocia. All inoculated mares developed placentitis and foaled before Day 314. Thirteen of 15 foals were not viable. All inoculated mares had abnormal progestin profiles and 13 of the 15 were identified by the abbreviated progestin profile. Transrectal CTUP was affected by gestational age and increased after inoculation (P<0.05). Nine of 15 inoculated mares had a CTUP>1.0 cm by 5-day post-inoculation. By performing both tests, 20 of 22 mares were correctly identified with respect to pregnancy outcome. However, three inoculated mares exhibited minimal clinical signs and likely would not be examined in a clinical setting. These tests were diagnostic for identifying feto-placental compromise in the mare.     

Correlates of preserved CD4(+) T cell homeostasis during natural, nonpathogenic simian immunodeficiency virus infection of sooty mangabeys: implications for AIDS pathogenesis

In contrast to HIV-infected humans, naturally SIV-infected sooty mangabeys (SMs) very rarely progress to AIDS. Although the mechanisms underlying this disease resistance are unknown, a consistent feature of natural SIV infection is the absence of the generalized immune activation associated with HIV infection. To define the correlates of preserved CD4(+) T cell counts in SMs, we conducted a cross-sectional immunological study of 110 naturally SIV-infected SMs. The nonpathogenic nature of the infection was confirmed by an average CD4(+) T cell count of 1,076 +/- 589/mm(3) despite chronic infection with a highly replicating virus. No correlation was found between CD4(+) T cell counts and either age (used as a surrogate marker for length of infection) or viremia. The strongest correlates of preserved CD4(+) T cell counts were a low percentage of circulating effector T cells (CD28(-)CD95(+) and/or IL-7R/CD127(-)) and a high percentage of CD4(+)CD25(+) T cells. These findings support the hypothesis that the level of immune activation is a key determinant of CD4(+) T cell counts in SIV-infected SMs. Interestingly, we identified 14 animals with CD4(+) T cell counts of <500/mm(3), of which two show severe and persistent CD4(+) T cell depletion (<50/mm(3)). Thus, significant CD4(+) T cell depletion does occasionally follow SIV infection of SMs even in the context of generally low levels of immune activation, lending support to the hypothesis of multifactorial control of CD4(+) T cell homeostasis in this model of infection. The absence of AIDS in these "CD4(low)" naturally SIV-infected SMs defines a protective role of the reduced immune activation even in the context of a significant CD4(+) T cell depletion.     

A subpopulation of macrophages infiltrates hypertrophic adipose tissue and is activated by free fatty acids via Toll-like receptors 2 and 4 and JNK-dependent pathways

Obesity and type 2 diabetes are characterized by decreased insulin sensitivity, elevated concentrations of free fatty acids (FFAs), and increased macrophage infiltration in adipose tissue (AT). Here, we show that FFAs can cause activation of RAW264.7 cells primarily via the JNK signaling cascade and that TLR2 and TLR4 are upstream of JNK and help transduce FFA proinflammatory signals. We also demonstrate that F4/80(+)CD11b(+)CD11c(+) bone marrow-derived dendritic cells (BMDCs) have heightened proinflammatory activity compared with F4/80(+)CD11b(+)CD11c(-) bone marrow-derived macrophages and that the proinflammatory activity and JNK phosphorylation of BMDCs, but not bone marrow-derived macrophages, was further increased by FFA treatment. F4/80(+)CD11b(+)CD11c(+) cells were found in AT, and the proportion and number of these cells in AT is increased in ob/ob mice and by feeding wild type mice a high fat diet for 1 and 12 weeks. AT F4/80(+)CD11b(+)CD11c(+) cells express increased inflammatory markers compared with F4/80(+)CD11b(+)CD11c(-) cells, and FFA treatment increased inflammatory responses in these cells. In addition, we found that CD11c expression is increased in skeletal muscle of high fat diet-fed mice and that conditioned medium from FFA-treated wild type BMDCs, but not TLR2/4 DKO BMDCs, can induce insulin resistance in L6 myotubes. Together our results show that FFAs can activate CD11c(+) myeloid proinflammatory cells via TLR2/4 and JNK signaling pathways, thereby promoting inflammation and subsequent cellular insulin resistance.     

Rat liver carnitine palmitoyltransferase 1 forms an oligomeric complex within the outer mitochondrial membrane

Carnitine palmitoyltransferase (CPT) 1A catalyzes the rate-limiting step in the transport of long chain acyl-CoAs from cytoplasm to the mitochondrial matrix by converting them to acylcarnitines. Located within the outer mitochondrial membrane, CPT1A activity is inhibited by malonyl-CoA, its allosteric inhibitor. In this study, we investigate for the first time the quaternary structure of rat CPT1A. Chemical cross-linking studies using intact mitochondria isolated from fed rat liver or from Saccharomyces cerevisiae expressing CPT1A show that CPT1A self-assembles into an oligomeric complex. Size exclusion chromatography experiments using solubilized mitochondrial extracts suggest that the fundamental unit of its quaternary structure is a trimer. When studied in blue native-PAGE, the CPT1A hexamer could be observed, however, suggesting that under these native conditions CPT1A trimers might be arranged as dimers. Moreover, the oligomeric state of CPT1A was found unchanged by starvation and by streptozotocin-induced diabetes, conditions characterized by changes in malonyl-CoA sensitivity of CPT1A. Finally, gel filtration analysis of several yeast-expressed chimeric CPTs demonstrates that the first 147 N-terminal residues of CPT1A, encompassing its two transmembrane segments, trigger trimerization independently of its catalytic C-terminal domain. Deletion of residues 1-82, including transmembrane 1, did not abrogate oligomerization, but the latter is limited to a trimer by the presence of the large catalytic C-terminal domain on the cytosolic face of mitochondria. Based on these findings, we proposed that the oligomeric structure of CPT1A would allow the newly formed acylcarnitines to gain direct access into the intermembrane space, hence facilitating substrate channeling.