Notch ligand endocytosis generates mechanical pulling force dependent on dynamin, epsins, and actin

Notch signaling induced by cell surface ligands is critical to development and maintenance of many eukaryotic organisms. Notch and its ligands are integral membrane proteins that facilitate direct cell-cell interactions to activate Notch proteolysis and release the intracellular domain that directs Notch-specific cellular responses. Genetic studies suggest that Notch ligands require endocytosis, ubiquitylation, and epsin endocytic adaptors to activate signaling, but the exact role of ligand endocytosis remains unresolved. Here we characterize a molecularly distinct mode of clathrin-mediated endocytosis requiring ligand ubiquitylation, epsins, and actin for ligand cells to activate signaling in Notch cells. Using a cell-bead optical tweezers system, we obtained evidence for cell-mediated mechanical force dependent on this distinct mode of ligand endocytosis. We propose that the mechanical pulling force produced by endocytosis of Notch-bound ligand drives conformational changes in Notch that permit activating proteolysis.     

White matter differences between healthy young ApoE4 carriers and non-carriers identified with tractography and support vector machines

The apolipoprotein E4 (ApoE4) is an established risk factor for Alzheimer's disease (AD). Previous work has shown that this allele is associated with functional (fMRI) changes as well structural grey matter (GM) changes in healthy young, middle-aged and older subjects. Here, we assess the diffusion characteristics and the white matter (WM) tracts of healthy young (20-38 years) ApoE4 carriers and non-carriers. No significant differences in diffusion indices were found between young carriers (ApoE4+) and non-carriers (ApoE4-). There were also no significant differences between the groups in terms of normalised GM or WM volume. A feature selection algorithm (ReliefF) was used to select the most salient voxels from the diffusion data for subsequent classification with support vector machines (SVMs). SVMs were capable of classifying ApoE4 carrier and non-carrier groups with an extremely high level of accuracy. The top 500 voxels selected by ReliefF were then used as seeds for tractography which identified a WM network that included regions of the parietal lobe, the cingulum bundle and the dorsolateral frontal lobe. There was a non-significant decrease in volume of this WM network in the ApoE4 carrier group. Our results indicate that there are subtle WM differences between healthy young ApoE4 carriers and non-carriers and that the WM network identified may be particularly vulnerable to further degeneration in ApoE4 carriers as they enter middle and old age.     

Transmissible spongiform encephalopathy strain-associated diversity of N-terminal proteinase K cleavage sites of PrP(Sc) from scrapie-infected and bovine spongiform encephalopathy-infected mice.

Assessment of the different conformational states of the abnormal prion protein (PrP(Sc)) in the CNS provides an established basis for distinguishing transmissible spongiform encephalopathy (TSE) strains. PrP(Sc) conformers are variably resistant to N-terminal proteinase K (PK) digestion, and analysis of the consensus products (PrP(res)) by immunoassay enables effective, but relatively low-resolution differentiation. Determination of the precise N-terminal amino acid profile (N-TAAP) of PrP(res) presents a potential high-resolution means of TSE-strain typing, and thus of differential disease diagnosis. This approach was evaluated using individual mice affected by model scrapie (22A, ME7, 87V and 79A) and bovine spongiform encephalopathy (BSE) (301V) strains. Nano liquid chromatography-mass spectrometry (LC-MS) was used to determine PrP(res) N-terminal tryptic digestion products. Four major N-terminal tryptic peptides were generated from all mouse TSE strains investigated, corresponding with predominant N-termination of PrP(res) at G(81), G(85), G(89) and G(91). Both the mass spectrometric abundance of the individual peptides and the ratios of pairs of these peptides were evaluated as markers of conformation in relation to their potential for strain discrimination. The yield of peptides was significantly greater for BSE than scrapie strains and the relative quantities of particular peptide pairs differed between strains. Thus, whereas peptide G(91)-K(105) was a dominant peptide from 301V, this was not the case for other strains and, significantly, the ratio of peptides G(91)-K(105):G(89)-K(105) was substantially higher for BSE-infected compared with scrapie-infected mice. These data support the potential of the N-TAAP approach for high-resolution TSE strain typing and differential diagnosis.     

Responses of efferent octopaminergic thoracic unpaired median neurons in the locust to visual and mechanosensory signals

Insect thoracic ganglia contain efferent octopaminergic unpaired median neurons (UM neurons) located in the midline, projecting bilaterally and modulating neuromuscular transmission, muscle contraction kinetics, sensory sensitivity and muscle metabolism. In locusts, these neurons are located dorsally or ventrally (DUM- or VUM-neurons) and divided into functionally different sub-populations activated during different motor tasks. This study addresses the responsiveness of locust thoracic DUM neurons to various sensory stimuli. Two classes of sense organs, cuticular exteroreceptor mechanosensilla (tactile hairs and campaniform sensilla), and photoreceptors (compound eyes and ocelli) elicited excitatory reflex responses. Chordotonal organ joint receptors caused no responses. The tympanal organ (Müller's organ) elicited weak excitatory responses most likely via generally increased network activity due to increased arousal. Vibratory stimuli to the hind leg subgenual organ never elicited responses. Whereas DUM neurons innervating wing muscles are not very responsive to sensory stimulation, those innervating leg and other muscles are very responsive to stimulation of exteroreceptors and hardly responsive to stimulation of proprioceptors. After cutting both cervical connectives all mechanosensory excitation is lost, even for sensory inputs from the abdomen. This suggests that, in contrast to motor neurons, the sensory inputs to octopaminergic efferent neuromodulatory cells are pre-processed in the suboesophageal ganglion.     

Atomic Layer Deposition of Functional Layers for on Chip 3D Li‐Ion All Solid State Microbattery

Nowadays, millimeter scale power sources are key devices for providing autonomy to smart, connected, and miniaturized sensors. However, until now, planar solid state microbatteries do not yet exhibit a sufficient surface energy density. In that context, architectured 3D microbatteries appear therefore to be a good solution to improve the material mass loading while keeping small the footprint area. Beside the design itself of the 3D microbaterry, one important technological barrier to address is the conformal deposition of thin films (lithiated or not) on 3D structures. For that purpose, atomic layer deposition (ALD) technology is a powerful technique that enables conformal coatings of thin film on complex substrate. An original, robust, and highly efficient 3D scaffold is proposed to significantly improve the geometrical surface of miniaturized 3D microbattery. Four functional layers composing the 3D lithium ion microbattery stacking has been successfully deposited on simple and double microtubes 3D templates. In depth synchrotron X‐ray nanotomography and high angle annular dark field transmission electron microscope analyses are used to study the interface between each layer. For the first time, using ALD, anatase TiO₂ negative electrode is coated on 3D tubes with Li₃PO₄ lithium phosphate as electrolyte, opening the way to all solid‐state 3D microbatteries. The surface capacity is significantly increased by the proposed topology (high area enlargement factor – “thick” 3D layer), from 3.5 μA h cm^?2 for a planar layer up to 0.37 mA h cm^?2 for a 3D thin film (105 times higher).     

Homozygosity mapping of a Weill-Marchesani syndrome locus to chromosome 19p13.3-p13.2

Weill-Marchesani syndrome (WMS) is a rare disease characterized by short stature, brachydactyly, joint stiffness, and characteristic eye abnormalities, including microspherophakia, ectopia lentis, and glaucoma. Both autosomal recessive and autosomal dominant modes of inheritance have been described in association with WMS. We have performed a genome-wide search in two large consanguineous families of Lebanese and Saudian origin consistent with an autosomal recessive mode of inheritance. Here, we report the linkage of the disease gene to chromosome 19p13.3-p13.2 (Zmax=5.99 at theta=0 at locus D19S906). A recombination event between loci D19S905 and D19S901 defines the distal boundary, and a second recombination event between loci D19S221 and D19S840 defines the proximal boundary of the genetic interval encompassing the WMS gene (12.4 cM). We hope that our ongoing studies will lead to the identification of the disease-causing gene.     

Metabolic Tumour Burden Measured by 18F-FDG PET/CT Predicts Malignant Transformation in Patients with Neurofibromatosis Type-1

Background

To investigate the diagnostic and prognostic performances of ¹⁸F-FDG PET/CT measures of metabolic tumour burden in patients with neurofibromatosis type-1 (NF1), suspect of malignant transformation.

Methods

This retrospective study included 49 patients (15–60 years old, 30 women) with a diagnosis of NF1, followed in our Reference Centre for Rare Neuromuscular Diseases, who presented clinical signs of tumour progression (pain, neurological deficit, tumour growth). Quantitative metabolic parameters were measured on 149 tumoral targets, using semi-automatic software and the best cut off values to predict transformation was assessed by Receiver Operating Characteristics (ROC) analysis. Prognostic value of PET/CT metabolic parameters was assessed by Kaplan-Meier estimates of overall survival.

Results

Lesions were histologically documented in 40 patients: a sarcomatous transformation was found in 16, a dysplastic neurofibroma (NF) in 7, and a benign NF in 17; in the remaining 9 patients, a minimal follow-up of 12 mo (median 59 mo) confirmed the absence of transformation. The optimal cut off values for detection of malignant transformation were, in decreasing order of area under the ROC curves, a tumour-to-liver (T/L) ratio >2.5, SUV_max > 4.5, total lesion glycolysis (TLG) > 377, total metabolic tumour volume (TMTV) > 88 cm³, and heterogeneity index (HI_suv) > 1.69. The best prognostic marker was the TLG: the 4-y estimates of survival were 97% [95% CI, 90% - 100%] in patients with TLG ≤ 377 vs. 27% [95% CI, 5% - 49%] in patients with TLG > 377 (P < 0.0001; χ² 27.85; hazard ratio 13.27 [95% CI, 3.72–47.35]). T/L ratio, SUV_max and TMTV demonstrated slightly lower performance to predict survival, with χ² ranging 14.41–19.12. The HI_suv index was not predictive of survival.

Conclusion

Our study demonstrates that TLG and TMTV, as PET/CT measures of metabolic tumour burden, may be used clinically to identify sarcomatous transformation in patients with NF1 and predict overall survival, with a higher specificity for the TLG. Conventional measures such as the SUV_max, and T/L ratio also demonstrate high prognostic value.     

Haplotype sharing refines the location of an imprinted quantitative trait locus with major effect on muscle mass to a 250-kb chromosome segment containing the porcine IGF2 gene

We herein describe the fine mapping of an imprinted QTL with major effect on muscle mass that was previously assigned to distal SSC2p in the pig. The proposed approach exploits linkage disequilibrium in combination with QTL genotyping by marker-assisted segregation analysis. By identifying a haplotype shared by all "Q" chromosomes, we map the QTL to an approximately 250-kb chromosome segment containing INS and IGF2 as the only known paternally expressed genes. This considerably reinforces the candidacy of these genes, justifying their detailed analysis.     

Modeling and E-M estimation of haplotype-specific relative risks from genotype data for a case-control study of unrelated individuals

The US National Cancer Institute has recently sponsored the formation of a Cohort Consortium (http://2002.cancer.gov/scpgenes.htm) to facilitate the pooling of data on very large numbers of people, concerning the effects of genes and environment on cancer incidence. One likely goal of these efforts will be generate a large population-based case-control series for which a number of candidate genes will be investigated using SNP haplotype as well as genotype analysis. The goal of this paper is to outline the issues involved in choosing a method of estimating haplotype-specific risk estimates for such data that is technically appropriate and yet attractive to epidemiologists who are already comfortable with odds ratios and logistic regression. Our interest is to develop and evaluate extensions of methods, based on haplotype imputation, that have been recently described (Schaid et al., Am J Hum Genet, 2002, and Zaykin et al., Hum Hered, 2002) as providing score tests of the null hypothesis of no effect of SNP haplotypes upon risk, which may be used for more complex tasks, such as providing confidence intervals, and tests of equivalence of haplotype-specific risks in two or more separate populations. In order to do so we (1) develop a cohort approach towards odds ratio analysis by expanding the E-M algorithm to provide maximum likelihood estimates of haplotype-specific odds ratios as well as genotype frequencies; (2) show how to correct the cohort approach, to give essentially unbiased estimates for population-based or nested case-control studies by incorporating the probability of selection as a case or control into the likelihood, based on a simplified model of case and control selection, and (3) finally, in an example data set (CYP17 and breast cancer, from the Multiethnic Cohort Study) we compare likelihood-based confidence interval estimates from the two methods with each other, and with the use of the single-imputation approach of Zaykin et al. applied under both null and alternative hypotheses. We conclude that so long as haplotypes are well predicted by SNP genotypes (we use the Rh2 criteria of Stram et al. [1]) the differences between the three methods are very small and in particular that the single imputation method may be expected to work extremely well.