Studies of the trafficking of the insulin-responsive glucose transporter, GLUT4, in 3T3-L1 adipocytes

The translocation of GLUT4 from intracellular stores to the plasma membrane in response to insulin accounts for the large insulin-mediated glucose uptake in muscle and fat tissue. A defect of this translocation mechanism is evident in individuals with insulin resistance and type 2 diabetes. Hence, understanding the molecular basis of GLUT4 localisation and recycling is important in order to assist the design of rational therapies for the treatment of this disease.Here, we have used iodixanol gradient analysis to examine the intracellular distribution of GLUT4. By this method intracellular GLUT4 could be separated into two pools one of which is highly insulin sensitive, and corresponds to "GLUT4 storage vesicles" (GSV), while the other is less insulin sensitive and is of endosomal origin. We further show that during differentiation of 3T3-L1 fibroblasts into adipocytes, the formation of the GSV compartment appears to be driven by the expression of GLUT4.SNARE proteins are involved in the fidelity of GLUT4 translocation, but little is known about how these proteins may be altered in insulin resistance. Using insulin-resistant 3T3-L1 adipocytes, we show that SNARE protein levels are increased. The potential importance of this observation is discussed. Using iodixanol gradient analysis we also found that the downregulation of GLUT4 known to occur in insulin-resistant 3T3-L1 adipocytes is predominantly from the GSV compartment. The implications of these data for the aetiology of insulin resistance are discussed

PUBA: Privacy-Preserving User-Data Bookkeeping and Analytics

In this paper we propose Privacy-preserving User-data Bookkeeping & Analytics (PUBA), a building block destined to enable the implementation of business models (e.g., targeted advertising) and regulations (e.g., fraud detection) requiring user-data analysis in a privacy-preserving way. In PUBA, users keep an unlinkable but authenticated cryptographic logbook containing their historic data on their device. This logbook can only be updated by the operator while its content is not revealed. Users can take part in a privacy-preserving analytics computation, where it is ensured that their logbook is up-to-date and authentic while the potentially secret analytics function is verified to be privacy-friendly. Taking constrained devices into account, users may also outsource analytic computations (to a potentially malicious proxy not colluding with the operator).We model our novel building block in the Universal Composability framework and provide a practical protocol instantiation. To demonstrate the flexibility of PUBA, we sketch instantiations of privacy-preserving fraud detection and targeted advertising, although it could be used in many more scenarios, e.g. data analytics for multi-modal transportation systems. We implemented our bookkeeping protocols and an exemplary outsourced analytics computation based on logistic regression using the MP-SPDZ MPC framework. Performance evaluations using a smartphone as user device and more powerful hardware for operator and proxy suggest that PUBA for smaller logbooks can indeed be practical

Differentiation at the MHCIIα and Cath2 Loci in Sympatric Salvelinus alpinus Resource Morphs in Lake Thingvallavatn

Publisher's versionNorthern freshwater fish may be suitable for the genetic dissection of ecological traits because they invaded new habitats after the last ice age (∼10.000 years ago). Arctic charr (Salvelinus alpinus) colonizing streams and lakes in Iceland gave rise to multiple populations of small benthic morphotypes, often in sympatry with a pelagic morphotype. Earlier studies have revealed significant, but subtle, genetic differentiation between the three most common morphs in Lake Thingvallavatn. We conducted a population genetic screen on four immunological candidate genes Cathelicidin 2 (Cath2), Hepcidin (Hamp), Liver expressed antimicrobial peptide 2a (Leap-2a), and Major Histocompatibility Complex IIα (MHCIIα) and a mitochondrial marker (D-loop) among the three most common Lake Thingvallavatn charr morphs. Significant differences in allele frequencies were found between morphs at the Cath2 and MHCIIα loci. No such signal was detected in the D-loop nor in the other two immunological genes. In Cath2 the small benthic morph deviated from the other two (FST = 0.13), one of the substitutions detected constituting an amino acid replacement polymorphism in the antimicrobial peptide. A more striking difference was found in the MHCIIα. Two haplotypes were very common in the lake, and their frequency differed greatly between the morphotypes (from 22% to 93.5%, FST = 0.67). We then expanded our study by surveying the variation in Cath2 and MHCIIα in 9 Arctic charr populations from around Iceland. The populations varied greatly in terms of allele frequencies at Cath2, but the variation did not correlate with morphotype. At the MHCIIα locus, the variation was nearly identical to the variation in the two benthic morphs of Lake Thingvallavatn. The results are consistent with a scenario where parts of the immune systems have diverged substantially among Arctic charr populations in Iceland, after colonizing the island ∼10.000 years ago.The Palsson laboratory is supported by Icelandic Research foundation and the University of Iceland research fund. Icelandic research foundation (grant of excellence: nr 100204011) to S.S. Sigurdsson, A. Palsson, B.K. Kristjansson, Zophonias O. Jonsson and Ian A. Johnston paid for part of this work. Kalina H. Kapralova and Johannes Gudbrandsson were supported by the University of Iceland doctoral fund. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Peer ReviewedRitrýnt tímari

Analysis of continuous neuronal activity evoked by natural speech with computational corpus linguistics methods

In the field of neurobiology of language, neuroimaging studies are generally based on stimulation paradigms consisting of at least two different conditions. Designing those paradigms can be very time-consuming and this traditional approach is necessarily data-limited. In contrast, in computational and corpus linguistics, analyses are often based on large text corpora, which allow a vast variety of hypotheses to be tested by repeatedly re-evaluating the data set. Furthermore, text corpora also allow exploratory data analysis in order to generate new hypotheses. By drawing on the advantages of both fields, neuroimaging and computational corpus linguistics, we here present a unified approach combining continuous natural speech and MEG to generate a corpus of speech-evoked neuronal activity

Differential gene expression during early development in recently evolved and sympatric Arctic charr morphs

Phenotypic differences between closely related taxa or populations can arise through genetic variation or be environmentally induced, leading to altered transcription of genes during development. Comparative developmental studies of closely related species or variable populations within species can help to elucidate the molecular mechanisms related to evolutionary divergence and speciation. Studies of Arctic charr (Salvelinus alpinus) and related salmonids have revealed considerable phenotypic variation among populations and in Arctic charr many cases of extensive variation within lakes (resource polymorphism) have been recorded. One example is the four Arctic charr morphs in the ∼10,000 year old Lake Thingvallavatn, which differ in numerous morphological and life history traits. We set out to investigate the molecular and developmental roots of this polymorphism by studying gene expression in embryos of three of the morphs reared in a common garden set-up. We performed RNA-sequencing, de-novo transcriptome assembly and compared gene expression among morphs during an important timeframe in early development, i.e., preceding the formation of key trophic structures. Expectedly, developmental time was the predominant explanatory variable. As the data were affected by some form of RNA-degradation even though all samples passed quality control testing, an estimate of 3′-bias was the second most common explanatory variable. Importantly, morph, both as an independent variable and as interaction with developmental time, affected the expression of numerous transcripts. Transcripts with morph effect, separated the three morphs at the expression level, with the two benthic morphs being more similar. However, Gene Ontology analyses did not reveal clear functional enrichment of transcripts between groups. Verification via qPCR confirmed differential expression of several genes between the morphs, including regulatory genes such as AT-Rich Interaction Domain 4A (arid4a) and translin (tsn). The data are consistent with a scenario where genetic divergence has contributed to differential expression of multiple genes and systems during early development of these sympatric Arctic charr morphs.he project was funded by The Icelandic Center for Research (grant number: 100204011) to Sigurður S. Snorrason, Arnar Pálsson, Zophonías O. Jónsson and Bjarni K. Kristjánsson. The University of Iceland Doctoral Fund to Jóhannes Guðbrandsson and University of Iceland research fund to Arnar Pálsson, Sigurður S. Snorrason and Zophonías O. Jónsson. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.Peer Reviewe

Differential expression of the aryl hydrocarbon receptor pathway associates with craniofacial polymorphism in sympatric Arctic charr

Background The developmental basis of craniofacial morphology hinges on interactions of numerous signalling systems. Extensive craniofacial variation in the polymorphic Arctic charr, a member of the salmonid family, from Lake Thingvallavatn (Iceland), offers opportunities to find and study such signalling pathways and their key regulators, thereby shedding light on the developmental pathways, and the genetics of trophic divergence. Results To identify genes involved in the craniofacial differences between benthic and limnetic Arctic charr, we used transcriptome data from different morphs, spanning early development, together with data on craniofacial expression patterns and skeletogenesis in model vertebrate species. Out of 20 genes identified, 7 showed lower gene expression in benthic than in limnetic charr morphs. We had previously identified a conserved gene network involved in extracellular matrix (ECM) organization and skeletogenesis, showing higher expression in developing craniofacial elements of benthic than in limnetic Arctic charr morphs. The present study adds a second set of genes constituting an expanded gene network with strong, benthic–limnetic differential expression. To identify putative upstream regulators, we performed knowledge-based motif enrichment analyses on the regulatory sequences of the identified genes which yielded potential binding sites for a set of known transcription factors (TFs). Of the 8 TFs that we examined using qPCR, two (Ahr2b and Ap2) were found to be differentially expressed between benthic and limnetic charr. Expression analysis of several known AhR targets indicated higher activity of the AhR pathway during craniofacial development in benthic charr morphotypes. Conclusion These results suggest a key role of the aryl hydrocarbon receptor (AhR) pathway in the observed craniofacial differences between distinct charr morphotypes.This project was supported by The Icelandic Centre for Research (RANNIS/IRF, Grant 100204), The University of Iceland Research Fund and the Eimskip University Fund.Peer Reviewe

Transcriptional dynamics of a conserved gene expression network associated with craniofacial divergence in Arctic charr

Background Understanding the molecular basis of craniofacial variation can provide insights into key developmental mechanisms of adaptive changes and their role in trophic divergence and speciation. Arctic charr (Salvelinus alpinus) is a polymorphic fish species, and, in Lake Thingvallavatn in Iceland, four sympatric morphs have evolved distinct craniofacial structures. We conducted a gene expression study on candidates from a conserved gene coexpression network, focusing on the development of craniofacial elements in embryos of two contrasting Arctic charr morphotypes (benthic and limnetic). Results Four Arctic charr morphs were studied: one limnetic and two benthic morphs from Lake Thingvallavatn and a limnetic reference aquaculture morph. The presence of morphological differences at developmental stages before the onset of feeding was verified by morphometric analysis. Following up on our previous findings that Mmp2 and Sparc were differentially expressed between morphotypes, we identified a network of genes with conserved coexpression across diverse vertebrate species. A comparative expression study of candidates from this network in developing heads of the four Arctic charr morphs verified the coexpression relationship of these genes and revealed distinct transcriptional dynamics strongly correlated with contrasting craniofacial morphologies (benthic versus limnetic). A literature review and Gene Ontology analysis indicated that a significant proportion of the network genes play a role in extracellular matrix organization and skeletogenesis, and motif enrichment analysis of conserved noncoding regions of network candidates predicted a handful of transcription factors, including Ap1 and Ets2, as potential regulators of the gene network. The expression of Ets2 itself was also found to associate with network gene expression. Genes linked to glucocorticoid signalling were also studied, as both Mmp2 and Sparc are responsive to this pathway. Among those, several transcriptional targets and upstream regulators showed differential expression between the contrasting morphotypes. Interestingly, although selected network genes showed overlapping expression patterns in situ and no morph differences, Timp2 expression patterns differed between morphs. Conclusion Our comparative study of transcriptional dynamics in divergent craniofacial morphologies of Arctic charr revealed a conserved network of coexpressed genes sharing functional roles in structural morphogenesis. We also implicate transcriptional regulators of the network as targets for future functional studies.This project was supported by The Icelandic Centre for Research (RANNIS/IRF, grant 100204) and The University of Iceland Research Fund.Peer Reviewe