213 research outputs found
Decision criteria for the selection of analytical instruments used in clinical chemistry: V The interaction of new instrumentation with laboratory infra-structure: modelling and simulation for planning of laboratory functions
Beskattning av idrottsutövare - intern internationell personbeskattning ur ett elitidrottsperspektiv
Idrottsmarknaden har gÄtt igenom stora förÀndringar de senaste Ärtiondena, vilket har lett till en pÄ mÄnga sÀtt ökad internationell konkurrens. FrÄn en tid nÀr Àven mÄnga av de stora stjÀrnorna sÄgs som glada amatörer har professionalismen inom idrotten ökat och allt fler utövare försörjer sig pÄ sitt idrottande. PÄ lagidrottsomrÄdet spelar lÀndernas interna regler stor roll för klubbarnas konkurrenskraft dÄ de pÄverkar möjligheten att erbjuda löner och villkor som kan locka de stora stjÀrnorna. Med hÀnsyn till att harmoniseringen pÄ skatteomrÄdet Àr högst begrÀnsad inom EU kan de regionala skillnaderna i skattesystemen variera avsevÀrt, varför det finns anledning att se nÀrmare pÄ hur dessa samspelar med idrotten.
Utövare inom de individuella idrotterna har större möjlighet att sjÀlva vÀlja sin bosÀttning Àn vad lagidrottarna har, eftersom de förstnÀmndas idrottande inte pÄ samma sÀtt Àr bundet till en viss plats eller land. För dessa utövare Àr praktiska skÀl ofta huvudorsaken vid val av bosÀttning, men Àven ett lands skatteregler Àr nÄgot som mÄnga gÄnger tas hÀnsyn till. Skattereglerna Àr inte minst avgörande i de fall idrottsstjÀrnor vÀljer att flytta ut frÄn Sverige efter att karriÀren har avslutats.
Den hÀr uppsatsen undersöker, med det svenska interna regelverket som utgÄngspunkt, skatterÀttens förhÄllande till elitidrottsutövare. Utöver att beskriva det teoretiska ramverket syftar uppsatsen ocksÄ till att tydliggöra och behandla de praktiska skatterÀttsliga frÄgor som elitidrottsutövare i Sverige har att brottas med
Tubulin Dimers Oligomerize before Their Incorporation into Microtubules
In the presence of GTP, purified dimers of α- and ÎČ-tubulin will interact longitudinally and laterally to self-assemble into microtubules (MTs). This property provides a powerful in vitro experimental system to describe MT dynamic behavior at the micrometer scale and to study effects and functioning of a large variety of microtubule associated proteins (MAPs). Despite the plethora of such data produced, the molecular mechanisms of MT assembly remain disputed. Electron microscopy (EM) studies suggested that tubulin dimers interact longitudinally to form short oligomers which form a tube by lateral interaction and which contribute to MT elongation. This idea is however challenged: Based on estimated association constants it was proposed that single dimers represent the major fraction of free tubulin. This view was recently supported by measurements suggesting that MTs elongate by addition of single tubulin dimers. To solve this discrepancy, we performed a direct measurement of the longitudinal interaction energy for tubulin dimers. We quantified the size distribution of tubulin oligomers using EM and fluorescence correlation spectroscopy (FCS). From the distribution we derived the longitudinal interaction energy in the presence of GDP and the non-hydrolysable GTP analog GMPCPP. Our data suggest that MT elongation and nucleation involves interactions of short tubulin oligomers rather than dimers. Our approach provides a solid experimental framework to better understand the role of MAPs in MT nucleation and growth
Students envisioning the future.
How can students be included as critical stakeholders in the systems and services provided by a university? To address the whole student experience, we engaged students and employees at a large Swedish university in a vision seminar process to elicit how these groups envisioned an ideal future university, and the necessary changes to technology and organisational structures required to achieve this ideal version. The process entailed six four-hour workshops with four groups consisting of six participants each. A survey instrument was used to follow up on the participants' experiences of participating in the vision seminar process and their thoughts on the future of the university. The results show that the participating students were more positive compared to the university employees. The students envisioned harmonized interdepartmental systems, seamlessly integrating a variety of services into one university-provided solution. The employees envisioned their future work as flexible, enabled by technology providing excellent support without hindering pedagogical and organisational development. Using technological frames, these visions of the future are identified, analysed and discussed in relation to the quality of university education and a holistic view on students' university experience. Finally we discuss the broader implications of the visions on the future of university education
Template-free 13-protofilament microtubuleâMAP assembly visualized at 8 A resolution
Microtubule-associated proteins (MAPs) are essential for regulating and organizing cellular microtubules (MTs). However, our mechanistic understanding of MAP function is limited by a lack of detailed structural information. Using cryo-electron microscopy and single particle algorithms, we solved the 8 Ă
structure of doublecortin (DCX)-stabilized MTs. Because of DCXâs unusual ability to specifically nucleate and stabilize 13-protofilament MTs, our reconstruction provides unprecedented insight into the structure of MTs with an in vivo architecture, and in the absence of a stabilizing drug. DCX specifically recognizes the corner of four tubulin dimers, a binding mode ideally suited to stabilizing both lateral and longitudinal lattice contacts. A striking consequence of this is that DCX does not bind the MT seam. DCX binding on the MT surface indirectly stabilizes conserved tubulinâtubulin lateral contacts in the MT lumen, operating independently of the nucleotide bound to tubulin. DCXâs exquisite binding selectivity uncovers important insights into regulation of cellular MTs
Ectopic A-lattice seams destabilize microtubules
Natural microtubules typically include one A-lattice seam within an otherwise helically symmetric B-lattice tube. It is currently unclear how A-lattice seams influence microtubule dynamic instability. Here we find that including extra A-lattice seams in GMPCPP microtubules, structural analogues of the GTP caps of dynamic microtubules, destabilizes them, enhancing their median shrinkage rate by >20-fold. Dynamic microtubules nucleated by seeds containing extra A-lattice seams have growth rates similar to microtubules nucleated by B-lattice seeds, yet have increased catastrophe frequencies at both ends. Furthermore, binding B-lattice GDP microtubules to a rigor kinesin surface stabilizes them against shrinkage, whereas microtubules with extra A-lattice seams are stabilized only slightly. Our data suggest that introducing extra A-lattice seams into dynamic microtubules destabilizes them by destabilizing their GTP caps. On this basis, we propose that the single A-lattice seam of natural B-lattice MTs may act as a trigger point, and potentially a regulation point, for catastrophe
Enteric Species F Human Adenoviruses use Laminin-Binding Integrins as Co-Receptors for Infection of Ht-29 Cells
The enteric species F human adenovirus types 40 and 41 (HAdV-40 and -41) are the third most common cause of infantile gastroenteritis in the world. Knowledge about HAdV-40 and -41 cellular infection is assumed to be fundamentally different from that of other HAdVs since HAdV-40 and -41 penton bases lack the aV-integrin-interacting RGD motif. This motif is used by other HAdVs mainly for internalization and endosomal escape. We hypothesised that the penton bases of HAdV-40 and -41 interact with integrins independently of the RGD motif. HAdV-41 transduction of a library of rodent cells expressing specific human integrin subunits pointed to the use of laminin-binding alpha 2-, alpha 3- and alpha 6- containing integrins as well as other integrins as candidate co-receptors. Specific laminins prevented internalisation and infection, and recombinant, soluble HAdV-41 penton base proteins prevented infection of human intestinal HT-29 cells. Surface plasmon resonance analysis demonstrated that HAdV-40 and -41 penton base proteins bind to alpha 6-containing integrins with an affinity similar to that of previously characterised penton base:integrin interactions. With these results, we propose that laminin-binding integrins are co-receptors for HAdV-40 and -41
Phosphoregulation of the budding yeast EB1 homologue Bim1p by Aurora/Ipl1p
EB1 (end binding 1) proteins have emerged as central regulators of microtubule (MT) plus ends in all eukaryotes, but molecular mechanisms controlling the activity of these proteins are poorly understood. In this study, we show that the budding yeast EB1 protein Bim1p is regulated by Aurora B/Ipl1p-mediated multisite phosphorylation. Bim1p forms a stable complex with Ipl1p and is phosphorylated on a cluster of six Ser residues in the flexible linker connecting the calponin homology (CH) and EB1 domains. Using reconstitution of plus end tracking in vitro and total internal reflection fluorescence microscopy, we show that dimerization of Bim1p and the presence of the linker domain are both required for efficient tip tracking and that linker phosphorylation removes Bim1p from static and dynamic MTs. Bim1 phosphorylation occurs during anaphase in vivo, and it is required for normal spindle elongation kinetics and an efficient disassembly of the spindle midzone. Our results define a mechanism for the use and regulation of CH domains in an EB1 protein
Dynamics of Kv1 Channel Transport in Axons
Concerted actions of various ion channels that are precisely targeted along axons are crucial for action potential initiation and propagation, and neurotransmitter release. However, the dynamics of channel protein transport in axons remain unknown. Here, using time-lapse imaging, we found fluorescently tagged Kv1.2 voltage-gated K+ channels (YFP-Kv1.2) moved bi-directionally in discrete puncta along hippocampal axons. Expressing KvÎČ2, a Kv1 accessory subunit, markedly increased the velocity, the travel distance, and the percentage of moving time of these puncta in both anterograde and retrograde directions. Suppressing the KvÎČ2-associated protein, plus-end binding protein EB1 or kinesin II/KIF3A, by siRNA, significantly decreased the velocity of YFP-Kv1.2 moving puncta in both directions. KvÎČ2 mutants with disrupted either Kv1.2-KvÎČ2 binding or KvÎČ2-EB1 binding failed to increase the velocity of YFP-Kv1.2 puncta, confirming a central role of KvÎČ2. Furthermore, fluorescently tagged Kv1.2 and KvÎČ2 co-moved along axons. Surprisingly, when co-moving with Kv1.2 and KvÎČ2, EB1 appeared to travel markedly faster than its plus-end tracking. Finally, using fission yeast S. pombe expressing YFP-fusion proteins as reference standards to calibrate our microscope, we estimated the numbers of YFP-Kv1.2 tetramers in axonal puncta. Taken together, our results suggest that proper amounts of Kv1 channels and their associated proteins are required for efficient transport of Kv1 channel proteins along axons
Mammalian end binding proteins control persistent microtubule growth
© 2009 Komarova et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License 3.0. The definitive version was published in Journal of Cell Biology 184 (2009): 691-706, doi:10.1083/jcb.200807179.End binding proteins (EBs) are highly conserved core components of microtubule plus-end tracking protein networks. Here we investigated the roles of the three mammalian EBs in controlling microtubule dynamics and analyzed the domains involved. Protein depletion and rescue experiments showed that EB1 and EB3, but not EB2, promote persistent microtubule growth by suppressing catastrophes. Furthermore, we demonstrated in vitro and in cells that the EB plus-end tracking behavior depends on the calponin homology domain but does not require dimer formation. In contrast, dimerization is necessary for the EB anti-catastrophe activity in cells; this explains why the EB1 dimerization domain, which disrupts native EB dimers, exhibits a dominant-negative effect. When microtubule dynamics is reconstituted with purified tubulin, EBs promote rather than inhibit catastrophes, suggesting that in cells EBs prevent catastrophes by counteracting other microtubule regulators. This probably occurs through their action on microtubule ends, because catastrophe suppression does not require the EB domains needed for binding to known EB partners.This work was supported by the Netherlands Organization for Scientifi
c Research grants to A.A., by Funda ç ĂŁ o para a Ci ĂȘ ncia e a Tecnologia
fellowship to S.M. Gouveia, by a FEBS fellowship to R.M. Buey, by the National
Institutes of Health grant GM25062 to G.G. Borisy and by the Swiss
National Science Foundation through grant 3100A0-109423 and by the
National Center of Competence in Research Structural Biology program to
M.O. Steinmetz
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