Sequence determinants of a microtubule tip localization signal (MtLS).

Microtubule plus-end-tracking proteins (+TIPs) specifically localize to the growing plus-ends of microtubules to regulate microtubule dynamics and functions. A large group of +TIPs contain a short linear motif, SXIP, which is essential for them to bind to end-binding proteins (EBs) and target microtubule ends. The SXIP sequence site thus acts as a widespread microtubule tip localization signal (MtLS). Here we have analyzed the sequence-function relationship of a canonical MtLS. Using synthetic peptide arrays on membrane supports, we identified the residue preferences at each amino acid position of the SXIP motif and its surrounding sequence with respect to EB binding. We further developed an assay based on fluorescence polarization to assess the mechanism of the EB-SXIP interaction and to correlate EB binding and microtubule tip tracking of MtLS sequences from different +TIPs. Finally, we investigated the role of phosphorylation in regulating the EB-SXIP interaction. Together, our results define the sequence determinants of a canonical MtLS and provide the experimental data for bioinformatics approaches to carry out genome-wide predictions of novel +TIPs in multiple organisms.

Metabolic compartmentalization in the human cortex and hippocampus: evidence for a cell- and region-specific localization of lactate dehydrogenase 5 and pyruvate dehydrogenase.

BACKGROUND: For a long time now, glucose has been thought to be the main, if not the sole substrate for brain energy metabolism. Recent data nevertheless suggest that other molecules, such as monocarboxylates (lactate and pyruvate mainly) could be suitable substrates. Although monocarboxylates poorly cross the blood brain barrier (BBB), such substrates could replace glucose if produced locally.The two key enzymatiques systems required for the production of these monocarboxylates are lactate dehydrogenase (LDH; EC1.1.1.27) that catalyses the interconversion of lactate and pyruvate and the pyruvate dehydrogenase complex that irreversibly funnels pyruvate towards the mitochondrial TCA and oxydative phosphorylation. RESULTS: In this article, we show, with monoclonal antibodies applied to post-mortem human brain tissues, that the typically glycolytic isoenzyme of lactate dehydrogenase (LDH-5; also called LDHA or LDHM) is selectively present in astrocytes, and not in neurons, whereas pyruvate dehydrogenase (PDH) is mainly detected in neurons and barely in astrocytes. At the regional level, the distribution of the LDH-5 immunoreactive astrocytes is laminar and corresponds to regions of maximal 2-deoxyglucose uptake in the occipital cortex and hippocampus. In hippocampus, we observed that the distribution of the oxidative enzyme PDH was enriched in the neurons of the stratum pyramidale and stratum granulosum of CA1 through CA4, whereas the glycolytic enzyme LDH-5 was enriched in astrocytes of the stratum moleculare, the alveus and the white matter, revealing not only cellular, but also regional, selective distributions. The fact that LDH-5 immunoreactivity was high in astrocytes and occurred in regions where the highest uptake of 2-deoxyglucose was observed suggests that glucose uptake followed by lactate production may principally occur in these regions. CONCLUSION: These observations reveal a metabolic segregation, not only at the cellular but also at the regional level, that support the notion of metabolic compartmentalization between astrocytes and neurons, whereby lactate produced by astrocytes could be oxidized by neurons.

MRI visualized neo-intimal dissection and co-localization of novel apoptotic markers apolipoprotein C-1, ceramide and caspase-3 in a Watanabe hyperlipidemic rabbit model

PURPOSE: Apoptotic arterial wall vascular smooth muscle cell death is known to contribute to plaque vulnerability and rupture. Novel apoptotic markers like apolipoprotein C-I have been implicated in apoptotic human vascular smooth muscle cell death via recruiting a neutral sphingomyelinase (N-SMase)-ceramide pathway. In vivo relevance of these observations in an animal model of plaque rupture has not been shown. METHODS AND RESULTS: Using Watanabe rabbits, we investigated three different groups (group 1, three normal Watanabe rabbits; group 2, six Watanabe rabbits fed with high cholesterol diet for 3 months; group 3, five Watanabe rabbits with similar diet but additional endothelial denudation). We followed progression of atherosclerosis to pharmacologically induced plaque rupture non-invasively using novel 3D magnetic resonance Fast-Field-Echo angiography (TR=7.2, TE=3.6 ms, matrix=512 x 512) and Fast-Spin-Echo vessel wall imaging methods (TR=3 heart beats, TE=10.5 ms, matrix=304 x 304) on 1.5 T MRI. MRI provided excellent image quality with good MRI versus histology vessel wall thickness correlation (r=0.8). In six animals of group 2/3 MRI detected neo-intimal dissection in the abdominal aorta which was accompanied by immuno-histochemical demonstration of concomitant aforementioned novel apoptotic markers, previously implicated in the apoptotic smooth muscle cell death in vitro. CONCLUSIONS: Our studies suggest a potential role for the signal transduction pathway involving apolipoprotein C-I for in vivo apoptosis and atherosclerotic plaque rupture visualized by MRI.

Spacial debris collector robot : An AI-based and autonomous debris collector satellite simulation

In this paper the core functions of an artificial intelligence (AI) for controlling a debris collector robot are designed and implemented. Using the robot operating system (ROS) as the base of this work a multi-agent system is built with abilities for task planning.

Integració d’ una unitat inercial i d’ un giroscopi de fibra òptica en un robot submarí i desenvolupament d’ un sistema de posicionament dins una piscina

El Grup de Visió per Computador i Robòtica (VICOROB) del departament d'Electrònica, Informàtica i Automàtica de la Universitat de Girona investiga en el camp de la robòtica submarina. Al CIRS (Centre d’Investigació en Robòtica Submarina), laboratori que forma part del grup VICOROB, el robot submarí Ictineu és la principal eina utilitzada per a desenvolupar els projectes de recerca. Recentment, el CIRS ha adquirit un nou sistema de sensors d' orientació basat en una unitat inercial i un giroscopi de fibra òptica. Aquest projecte pretén realitzar un estudi d' aquests dispositius i integrar-los al robot Ictineu. D' altra banda, aprofitant les característiques d’aquests sensors giroscopics i les mesures d' un sonar ja integrat al robot, es vol desenvolupar un sistema de localització capaç de determinar la posició del robot en el pla horitzontal de la piscina en temps real

Entorn de programació pel robot Stäubli mitjantçant MatLab, aplicació a un sistema de recol·lecció de peces basat en visió artificial

Els objectius del projecte són: realitzar un intèrpret de comandes en VAL3 que rebi les ordres a través d’una connexió TCP/IP; realitzar una toolbox de Matlab per enviar diferents ordres mitjançant una connexió TCP/IP; adquirir i processar mitjançant Matlab imatges de la càmera en temps real i detectar la posició d’objectes artificials mitjançant la segmentació per color i dissenyar i realitzar una aplicació amb Matlab que reculli peces detectades amb la càmera. L’abast del projecte inclou: l’estudi del llenguatge de programació VAL3 i disseny de l’ intèrpret de comandes, l’estudi de les llibreries de Matlab per comunicació mitjançant TCP/IP, per l’adquisició d’imatges, pel processament d’imatges i per la programació en C; el disseny de la aplicació recol·lectora de peces i la implementació de: un intèrpret de comandes en VAL3, la toolbox pel control del robot STAUBLI en Matlab i la aplicació recol·lectora de peces mitjançant el processament d’imatges en temps real també en Matlab

Disseny i construcció d’ un braç de robot de 5 graus de llibertat governat des de PC via USB.

El braç robot es va crear com a resposta a una necessitat de fabricació d’elements mitjançant la producció en cadena i en tasques que necessiten precisió. Hi ha, però, altres tipus de tasques les quals no són repetitives, ni poden ésser programades, que necessiten però ser controlades en tot moment per un ésser humà. Són activitats que han d’estar realitzades per un ésser humà, però que requereixen molta precisió, és per això que es creu necessari el disseny d’un prototipus de control d’un braç robot estàndard, que permeti a una persona el control total sobre aquest en temps real per a la realització d’una tasca no repetitiva i no programable prèviament.Pretenem, en el present projecte, dissenyar i construir un braç robot de 5 graus de llibertat, controlat des d’un PC mitjançant un microcontrolador PIC amb comunicació a través d’un bus USB. El robot serà governat des d’un PC a través d’un software de control específic

Control de posició del robot wifibot

Aquest projecte pretén presentar de forma clara i detallada l’estructura i el funcionament del robot així com dels components que el conformen. Aquesta informació és de vital importància a l’hora de desenvolupar aplicacions per al robot. Un cop descrites les característiques del robot s’analitzaran les eines necessàries i/o disponibles per poder desenvolupar programari per cada nivell de la forma més senzilla i eficient possible. Posteriorment s’analitzaran els diferents nivells de programació i se’n contrastaran els avantatges i els inconvenients de cada un. Aquest anàlisi es començarà fent pel nivell més alt i anirà baixant amb la intenció de no entrar en nivells més baixos del necessari. Baixar un nivell en la programació suposa haver de crear aplicacions sempre compatibles amb els nivells superiors de forma que com més es baixa més augmenta la complexitat. A partir d’aquest anàlisi s’ha arribat a la conclusió que per tal d’aprofitar totes les prestacions del robot és precís arribar a programar en el nivell més baix del robot. Finalment l’objectiu és obtenir una sèrie de programes per cada nivell que permetin controlar el robot i fer-lo seguir senzilles trajectòries

Selective tumor localization of radiolabeled anti-human melanoma monoclonal antibody fragment demonstrated in the nude mouse model.

Monoclonal antibodies (Mab) directed against distinct epitopes of the human 240 kD melanoma-associated antigen have been evaluated for their capacity to localize in human melanoma grafted into nude mice. A favorable tumor to normal tissue ratio of 13 was obtained with intact 131I-labeled MAb Me1-14. This ratio was further increased to 43 and 23 by the use of F(ab')2 and Fab fragments, respectively. The specificity of tumor localization was demonstrated by the simultaneous injection of F(ab')2 fragments from MAb Me1-14 and anti-CEA MAb 35, each labeled with a different iodine isotope, into nude mice grafted with a melanoma and colon carcinoma. The fragments from both MAb localized with perfect selectivity in their relevant tumor as shown by differential whole body scanning and by direct measurement of the two isotopes in tumors and normal tissues. These in vivo experimental results suggest that the F(ab')2 fragment from MAb Me1-14 is suitable for melanoma detection by immunoscintigraphy in patients.

Expression of T cell receptor genes in the thymus: localization of transcripts in situ and comparison of mature and immature subsets.

We have analyzed the expression of T cell receptor (TcR) genes in the thymus using in situ RNA hybridizations with probes to the constant regions of the TcR alpha, beta, gamma and delta chains. Localization of transcripts revealed low TcR alpha mRNA levels in the thymus cortex and very low levels in the subcapsular region. In contrast, TcR beta message was very abundant in the cortex. TcR gamma or delta mRNA+ thymocytes showed a scattered, predominantly cortical localization. In contrast to gamma, TcR delta transcripts were abundant in the subcapsular region. Control experiments with sorted TcR alpha/beta or gamma/delta cells revealed a detection efficiency of 75%-85% for the respective TcR mRNA and data on TcR gene expression in mature, CD3+ thymocytes were consistent with previous reports. The analysis of immature, CD3- thymocyte subsets, however, revealed a virtual absence of TcR alpha transcripts and an unexpectedly high proportion of cells (14%-29%) expressing the gene for the TcR delta chain. The data are discussed in view of current models of lineage relationships in the thymus.

Dimeric recombinant IgA directed against carcino-embryonic antigen, a novel tool for carcinoma localization.

Carcinoembryonic antigen (CEA) has been shown to be one of the best markers for in vivo tumor targeting of radiolabeled antibodies, despite the fact that it is localized predominantly at the apical side of human colon carcinoma cells within the fairly closed pseudolumen structures formed by these tumors. Due to this particular histological localization, a large proportion of the CEA molecules may remain inaccessible to the intravenously injected radiolabeled anti-CEA antibodies of IgG isotype, which are widely used in the clinic. In order to improve targeting, we made a recombinant dimeric IgA, which should have the capacity to translocate from the basolateral to the apical side of the pseudolumen formed by colon carcinoma cells after binding to the polyIg receptor (pIgR). A genomic chimeric mouse-human IgA2 construct was made using one of our most specific anti-CEA hybridomas, CE-25. The chimeric IgA (chIgA) was expressed in the Sp2/0 myeloma cell line. The secreted recombinant antibody was found to consist mostly of a dimeric form of IgA with a molecular weight of about 350 kDa. The dimeric chIgA was shown to translocate efficiently in vitro across a monolayer of epithelial cells expressing the pIgR and to retain full CEA binding activity.

Disseny i control d'un robot equilibrista sobre dues rodes

Dissenyar un robot equilibrista i construir-lo. A més implementar les funcions per poder fer que s'aguanti dret. El robot ens permetrà transportar documents i ens detectarà obstacles mentres fa el transport. Estarà controlat per una aplicació java construida especialment per poder fer el control per bluetooth i també podrem monitoritzar informació i fer canvis en la configuració.

Radiolabeled fragments of monoclonal antibodies against carcinoembryonic antigen for localization of human colon carcinoma grafted into nude mice.

Four monoclonal antibodies against carcinoembryonic antigen (CEA) have been selected from 32 hybrids that produce antibodies against this antigen, by the criteria of high affinity for CEA and low cross-reactivity with granulocyte glycoprotein(s). The specificity of tumor localization in vivo of the four MAb, and their F(ab')2 and Fab fragments was compared in nude mice bearing grafts of a serially transplanted, CEA-producing, human colon carcinoma. The distribution of radiolabeled MAb and their fragments after intravenous injection was analyzed by direct measurement of radioactivity in tumor and normal organs, as well as by whole-body scanning and by autoradiography of tumor sections. Paired labeling experiments, in which 131I-labeled antibody or fragments and 125I-labeled control IgG are injected simultaneously, were undertaken to determine the relative tumor uptakes of each labeled protein. The tumor antibody uptake divided by that of control IgG defines the specificity index of localization. Tumor antibody uptakes (as compared with the whole mouse), ranging between 7 and 15, and specificity indices ranging between 3.4 and 6.8, were obtained with the four intact MAb at day 4-5 after injection. With F(ab')2 fragments of the four MAb, at day 3, the tumor antibody uptakes ranged between 12 and 24 and the specificity indices between 5.3 and 8.2. With the Fab fragments prepared from the two most promising MAb, the antibody uptakes reached values of 34 and 82 at day 2-3 and the specificity indices were as high as 12 and 19. The scanning results paralleled those obtained by direct measurement of radioactivity. With intact MAb, tumor grafts of 0.5-1 g gave very contrasted positive scans 3 d after injection. Using MAb fragments, tumors of smaller size were detectable earlier. The best results were obtained with Fab fragments of MAb 35, which gave clear detections of tumors weighing only 0.1 g as early as 48 h after injection. Autoradiographs of tumor sections from mice injected with 125I-labeled MAb demonstrated that the radioactivity was localized in the tumor tissues and not in the stromal connective tissue of mouse origin. The highest radioactivity concentration was localized in areas known to contain CEA such as the pseudolumen of glands and the apical side of carcinoma cells. The penetration of radioactivity in the central part of tumor nodules and the pseudolumen appeared to be increased with the use of MAb fragments.

The murine interleukin-2 receptor gamma chain gene: organization, chromosomal localization and expression in the adult thymus.

Defects in the interleukin-2 receptor gamma (IL-2R gamma) chain in the man result in an X-linked severe combined immunodeficiency, SCIDX1, characterized by an absence of T-cell differentiation. This phenotype may result from pertubations in IL-2, IL-4-, IL-7- or IL-15-mediated signaling, as the IL-2R gamma chain forms an integral component of these receptor systems. We have isolated and characterized cDNA and genomic clones for the murine IL-2R gamma. The gene (Il2rg) is well conserved between mouse and man with respect to overall structure and size, and contains regions of high conservation in the promoter region as well. Il2rg maps to mouse X chromosome region 40, in a region of synteny with human Xq12-13.1. We have also explored the expression of the IL-2R gamma during thymocyte development. IL-2R gamma transcripts are detected in the earliest thymocyte precursor cells and persist throughout intrathymic development into the mature peripheral compartment. Genomic clones for the murine IL-2R gamma will allow for further studies on the regulation and function of this gene in vivo.

Small scale underwater change detection

In this paper, we present a method to deal with the constraints of the underwater medium for finding changes between sequences of underwater images. One of the main problems of underwater medium for automatically detecting changes is the low altitude of the camera when taking pictures. This emphasise the parallax effect between the images as they are not taken exactly at the same position. In order to solve this problem, we are geometrically registering the images together taking into account the relief of the scene