270 research outputs found
Imaging Trans-Cellular Neurexin-Neuroligin Interactions by Enzymatic Probe Ligation
Neurexin and neuroligin are transmembrane adhesion proteins that play an important role in organizing the neuronal synaptic cleft. Our lab previously reported a method for imaging the trans-synaptic binding of neurexin and neuroligin called BLINC (Biotin Labeling of INtercellular Contacts). In BLINC, biotin ligase (BirA) is fused to one protein while its 15-amino acid acceptor peptide substrate (AP) is fused to the binding partner. When the two fusion proteins interact across cellular junctions, BirA catalyzes the site-specific biotinylation of AP, which can be read out by staining with streptavidin-fluorophore conjugates. Here, we report that BLINC in neurons cannot be reproduced using the reporter constructs and labeling protocol previously described. We uncover the technical reasons for the lack of reproducibilty and then re-design the BLINC reporters and labeling protocol to achieve neurexin-neuroligin BLINC imaging in neuron cultures. In addition, we introduce a new method, based on lipoic acid ligase instead of biotin ligase, to image trans-cellular neurexin-neuroligin interactions in human embryonic kidney cells and in neuron cultures. This method, called ID-PRIME for Interaction-Dependent PRobe Incorporation Mediated by Enzymes, is more robust than BLINC due to higher surface expression of lipoic acid ligase fusion constructs, gives stronger and more localized labeling, and is more versatile than BLINC in terms of signal readout. ID-PRIME expands the toolkit of methods available to study trans-cellular protein-protein interactions in living systems.National Institutes of Health (U.S.) (DP1 OD003961
Meta-analysis Reveals Genome-Wide Significance at 15q13 for Nonsyndromic Clefting of Both the Lip and the Palate, and Functional Analyses Implicate GREM1 As a Plausible Causative Gene
Nonsyndromic orofacial clefts are common birth defects with multifactorial etiology. The
most common type is cleft lip, which occurs with or without cleft palate (nsCLP and nsCLO,
respectively). Although genetic components play an important role in nsCLP, the genetic
factors that predispose to palate involvement are largely unknown. In this study, we carried
out a meta-analysis on genetic and clinical data from three large cohorts and identified
strong association between a region on chromosome 15q13 and nsCLP (P = 8.13×10−14 for
rs1258763; relative risk (RR): 1.46, 95% confidence interval (CI): 1.32–1.61)) but not
nsCLO (P = 0.27; RR: 1.09 (0.94–1.27)). The 5 kb region of strongest association maps
downstream of Gremlin-1 (GREM1), which encodes a secreted antagonist of the BMP4
pathway. We show during mouse embryogenesis, Grem1 is expressed in the developing lip
and soft palate but not in the hard palate. This is consistent with genotype-phenotype correlations
between rs1258763 and a specific nsCLP subphenotype, since a more than two-fold
increase in risk was observed in patients displaying clefts of both the lip and soft palate but
who had an intact hard palate (RR: 3.76, CI: 1.47–9.61, Pdiff<0.05). While we did not find lip
or palate defects in Grem1-deficient mice, wild type embryonic palatal shelves developed
divergent shapes when cultured in the presence of ectopic Grem1 protein (P = 0.0014). The
present study identified a non-coding region at 15q13 as the second, genome-wide significant
locus specific for nsCLP, after 13q31. Moreover, our data suggest that the closely
located GREM1 gene contributes to a rare clinical nsCLP entity. This entity specifically
involves abnormalities of the lip and soft palate, which develop at different time-points and
in separate anatomical regions.Clefts of the lip and palate are common birth defects, and require long-term multidisciplinary
management. Their etiology involves genetic factors and environmental influences
and/or a combination of both, however, these interactions are poorly defined. Moreover,
although clefts of the lip may or may not involve the palate, the determinants predisposing
to specific subphenotypes are largely unknown. Here we demonstrate that variations in
the non-coding region near the GREM1 gene show a highly significant association with a
particular phenotype in which cleft lip and cleft palate co-occ
Selective Induction of Cell Death in Melanoma Cell Lines through Targeting of Mcl-1 and A1
Melanoma is an often fatal form of skin cancer which is remarkably resistant against radio- and chemotherapy. Even new strategies that target RAS/RAF signaling and display unprecedented efficacy are characterized by resistance mechanisms. The targeting of survival pathways would be an attractive alternative strategy, if tumor-specific cell death can be achieved. Bcl-2 proteins play a central role in regulating survival of tumor cells. In this study, we systematically investigated the relevance of antiapoptotic Bcl-2 proteins, i.e., Bcl-2, Bcl-xL, Bcl-w, Mcl-1, and A1, in melanoma cell lines and non-malignant cells using RNAi. We found that melanoma cells required the presence of specific antiapoptotic Bcl-2 proteins: Inhibition of Mcl-1 and A1 strongly induced cell death in some melanoma cell lines, whereas non-malignant cells, i.e., primary human fibroblasts or keratinocytes were not affected. This specific sensitivity of melanoma cells was further enhanced by the combined inhibition of Mcl-1 and A1 and resulted in 60% to 80% cell death in all melanoma cell lines tested. This treatment was successfully combined with chemotherapy, which killed a substantial proportion of cells that survived Mcl-1 and A1 inhibition. Together, these results identify antiapoptotic proteins on which specifically melanoma cells rely on and, thus, provide a basis for the development of new Bcl-2 protein-targeting therapies
The Long-Baseline Neutrino Experiment: Exploring Fundamental Symmetries of the Universe
The preponderance of matter over antimatter in the early Universe, the
dynamics of the supernova bursts that produced the heavy elements necessary for
life and whether protons eventually decay --- these mysteries at the forefront
of particle physics and astrophysics are key to understanding the early
evolution of our Universe, its current state and its eventual fate. The
Long-Baseline Neutrino Experiment (LBNE) represents an extensively developed
plan for a world-class experiment dedicated to addressing these questions. LBNE
is conceived around three central components: (1) a new, high-intensity
neutrino source generated from a megawatt-class proton accelerator at Fermi
National Accelerator Laboratory, (2) a near neutrino detector just downstream
of the source, and (3) a massive liquid argon time-projection chamber deployed
as a far detector deep underground at the Sanford Underground Research
Facility. This facility, located at the site of the former Homestake Mine in
Lead, South Dakota, is approximately 1,300 km from the neutrino source at
Fermilab -- a distance (baseline) that delivers optimal sensitivity to neutrino
charge-parity symmetry violation and mass ordering effects. This ambitious yet
cost-effective design incorporates scalability and flexibility and can
accommodate a variety of upgrades and contributions. With its exceptional
combination of experimental configuration, technical capabilities, and
potential for transformative discoveries, LBNE promises to be a vital facility
for the field of particle physics worldwide, providing physicists from around
the globe with opportunities to collaborate in a twenty to thirty year program
of exciting science. In this document we provide a comprehensive overview of
LBNE's scientific objectives, its place in the landscape of neutrino physics
worldwide, the technologies it will incorporate and the capabilities it will
possess.Comment: Major update of previous version. This is the reference document for
LBNE science program and current status. Chapters 1, 3, and 9 provide a
comprehensive overview of LBNE's scientific objectives, its place in the
landscape of neutrino physics worldwide, the technologies it will incorporate
and the capabilities it will possess. 288 pages, 116 figure
Longitudinal river zonation in the tropics: examples of fish and caddisflies from endorheic Awash river, Ethiopia
Primary Research PaperSpecific concepts of fluvial ecology are
well studied in riverine ecosystems of the temperate
zone but poorly investigated in the Afrotropical
region. Hence, we examined the longitudinal zonation
of fish and adult caddisfly (Trichoptera) assemblages
in the endorheic Awash River (1,250 km in length),
Ethiopia. We expected that species assemblages are
structured along environmental gradients, reflecting
the pattern of large-scale freshwater ecoregions. We
applied multivariate statistical methods to test for differences in spatial species assemblage structure and
identified characteristic taxa of the observed biocoenoses
by indicator species analyses. Fish and
caddisfly assemblages were clustered into highland
and lowland communities, following the freshwater
ecoregions, but separated by an ecotone with highest
biodiversity. Moreover, the caddisfly results suggest
separating the heterogeneous highlands into a forested
and a deforested zone. Surprisingly, the Awash
drainage is rather species-poor: only 11 fish (1
endemic, 2 introduced) and 28 caddisfly species (8
new records for Ethiopia) were recorded from the
mainstem and its major tributaries. Nevertheless,
specialized species characterize the highland forests, whereas the lowlands primarily host geographically
widely distributed species. This study showed that a
combined approach of fish and caddisflies is a
suitable method for assessing regional characteristics
of fluvial ecosystems in the tropicsinfo:eu-repo/semantics/publishedVersio
Bmp7 Regulates the Survival, Proliferation, and Neurogenic Properties of Neural Progenitor Cells during Corticogenesis in the Mouse
Bone morphogenetic proteins (BMPs) are considered important regulators of neural development. However, results mainly from a wide set of in vitro gain-of-function experiments are conflicting since these show that BMPs can act either as inhibitors or promoters of neurogenesis. Here, we report a specific and non-redundant role for BMP7 in cortical neurogenesis in vivo using knockout mice. Bmp7 is produced in regions adjacent to the developing cortex; the hem, meninges, and choroid plexus, and can be detected in the cerebrospinal fluid. Bmp7 deletion results in reduced cortical thickening, impaired neurogenesis, and loss of radial glia attachment to the meninges. Subsequent in vitro analyses of E14.5 cortical cells revealed that lack of Bmp7 affects neural progenitor cells, evidenced by their reduced proliferation, survival and self-renewal capacity. Addition of BMP7 was able to rescue these proliferation and survival defects. In addition, at the developmental stage E14.5 Bmp7 was also required to maintain Ngn2 expression in the subventricular zone. These data demonstrate a novel role for Bmp7 in the embryonic mouse cortex: Bmp7 nurtures radial glia cells and regulates fundamental properties of neural progenitor cells that subsequently affect Ngn2-dependent neurogenesis
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An integrated omics analysis reveals molecular mechanisms that are associated with differences in seed oil content between Glycine max and Brassica napus
Abstract
Background: Rapeseed (Brassica napus L.) and soybean (Glycine max L.) seeds are rich in both protein and oil, which
are major sources of biofuels and nutrition. Although the difference in seed oil content between soybean (~ 20%) and
rapeseed (~ 40%) exists, little is known about its underlying molecular mechanism.
Results: An integrated omics analysis was performed in soybean, rapeseed, Arabidopsis (Arabidopsis thaliana L. Heynh),
and sesame (Sesamum indicum L.), based on Arabidopsis acyl-lipid metabolism- and carbon metabolism-related genes.
As a result, candidate genes and their transcription factors and microRNAs, along with phylogenetic analysis and
co-expression network analysis of the PEPC gene family, were found to be largely associated with the difference
between the two species. First, three soybean genes (Glyma.13G148600, Glyma.13G207900 and Glyma.12G122900)
co-expressed with GmPEPC1 are specifically enriched during seed storage protein accumulation stages, while the
expression of BnPEPC1 is putatively inhibited by bna-miR169, and two genes BnSTKA and BnCKII are co-expressed
with BnPEPC1 and are specifically associated with plant circadian rhythm, which are related to seed oil biosynthesis. Then,
in de novo fatty acid synthesis there are rapeseed-specific genes encoding subunits β-CT (BnaC05g37990D) and BCCP1
(BnaA03g06000D) of heterogeneous ACCase, which could interfere with synthesis rate, and β-CT is positively regulated by
four transcription factors (BnaA01g37250D, BnaA02g26190D, BnaC01g01040D and BnaC07g21470D). In triglyceride synthesis,
GmLPAAT2 is putatively inhibited by three miRNAs (gma-miR171, gma-miR1516 and gma-miR5775). Finally, in rapeseed
there was evidence for the expansion of gene families, CALO, OBO and STERO, related to lipid storage, and
the contraction of gene families, LOX, LAH and HSI2, related to oil degradation.
Conclusions: The molecular mechanisms associated with differences in seed oil content provide the basis for
future breeding efforts to improve seed oil content
A multi-disciplinary perspective on emergent and future innovations in peer review [version 2; referees: 2 approved]
Peer review of research articles is a core part of our scholarly communication system. In spite of its importance, the status and purpose of peer review is often contested. What is its role in our modern digital research and communications infrastructure? Does it perform to the high standards with which it is generally regarded? Studies of peer review have shown that it is prone to bias and abuse in numerous dimensions, frequently unreliable, and can fail to detect even fraudulent research. With the advent of web technologies, we are now witnessing a phase of innovation and experimentation in our approaches to peer review. These developments prompted us to examine emerging models of peer review from a range of disciplines and venues, and to ask how they might address some of the issues with our current systems of peer review. We examine the functionality of a range of social Web platforms, and compare these with the traits underlying a viable peer review system: quality control, quantified performance metrics as engagement incentives, and certification and reputation. Ideally, any new systems will demonstrate that they out-perform and reduce the biases of existing models as much as possible. We conclude that there is considerable scope for new peer review initiatives to be developed, each with their own potential issues and advantages. We also propose a novel hybrid platform model that could, at least partially, resolve many of the socio-technical issues associated with peer review, and potentially disrupt the entire scholarly communication system. Success for any such development relies on reaching a critical threshold of research community engagement with both the process and the platform, and therefore cannot be achieved without a significant change of incentives in research environments
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