http://www.recercat.cat:80/handle/2072/48762 www.ub.edu Sat, 18 May 2013 08:02:25 GMT 2013-05-18T08:02:25Z http://www.recercat.cat:80/bitstream/id/26319/ http://www.recercat.cat:80/handle/2072/48762 http://www.recercat.cat:80/handle/2072/211230 Fine-Tuning Translation Kinetics Selection as the Driving Force of Codon Usage Bias in the Hepatitis A Virus Capsid Aragonès, Ll.; Guix Arnau, Susana; Ribes Mora, Enric; Bosch, Albert; Pintó Solé, Rosa María Hepatitis A virus (HAV), the prototype of genus Hepatovirus, has several unique biological characteristics that distinguish it from other members of the Picornaviridae family. Among these, the need for an intact eIF4G factor for the initiation of translation results in an inability to shut down host protein synthesis by a mechanism similar to that of other picornaviruses. Consequently, HAV must inefficiently compete for the cellular translational machinery and this may explain its poor growth in cell culture. In this context of virus/cell competition, HAV has strategically adopted a naturally highly deoptimized codon usage with respect to that of its cellular host. With the aim to optimize its codon usage the virus was adapted to propagate in cells with impaired protein synthesis, in order to make tRNA pools more available for the virus. A significant loss of fitness was the immediate response to the adaptation process that was, however, later on recovered and more associated to a re-deoptimization rather than to an optimization of the codon usage specifically in the capsid coding region. These results exclude translation selection and instead suggest fine-tuning translation kinetics selection as the underlying mechanism of the codon usage bias in this specific genome region. Additionally, the results provide clear evidence of the Red Queen dynamics of evolution since the virus has very much evolved to re-adapt its codon usage to the environmental cellular changing conditions in order to recover the original fitness. http://www.recercat.cat:80/handle/2072/211230 http://www.recercat.cat:80/handle/2072/211079 In cortical neurons HDAC3 suppresses RD4-dependent SMRT export Soriano Zaragoza, Francesc X.(Francesc Xavier); Hardingham, G.E. The transcriptional corepressor SMRT controls neuronal responsiveness of several transcription factors and can regulate neuroprotective and neurogenic pathways. SMRT is a multi-domain protein that complexes with HDAC3 as well as being capable of interactions with HDACs 1, 4, 5 and 7. We previously showed that in rat cortical neurons, nuclear localisation of SMRT requires histone deacetylase activity: Inhibition of class I/II HDACs by treatment with trichostatin A (TSA) causes redistribution of SMRT to the cytoplasm, and potentiates the activation of SMRT-repressed nuclear receptors. Here we have sought to identify the HDAC(s) and region(s) of SMRT responsible for anchoring it in the nucleus under normal circumstances and for mediating nuclear export following HDAC inhibition. We show that in rat cortical neurons SMRT export can be triggered by treatment with the class I-preferring HDAC inhibitor valproate and the HDAC2/3-selective inhibitor apicidin, and by HDAC3 knockdown, implicating HDAC3 activity as being required to maintain SMRT in the nucleus. HDAC3 interaction with SMRT's deacetylation activation domain (DAD) is known to be important for activation of HDAC3 deacetylase function. Consistent with a role for HDAC3 activity in promoting SMRT nuclear localization, we found that inactivation of SMRT's DAD by deletion or point mutation triggered partial redistribution of SMRT to the cytoplasm. We also investigated whether other regions of SMRT were involved in mediating nuclear export following HDAC inhibition. TSA- and valproate-induced SMRT export was strongly impaired by deletion of its repression domain-4 (RD4). Furthermore, over-expression of a region of SMRT containing the RD4 region suppressed TSA-induced export of full-length SMRT. Collectively these data support a model whereby SMRT's RD4 region can recruit factors capable of mediating nuclear export of SMRT, but whose function and/or recruitment is suppressed by HDAC3 activity. Furthermore, they underline the fact that HDAC inhibitors can cause reorganization and redistribution of corepressor complexes. http://www.recercat.cat:80/handle/2072/211079 http://www.recercat.cat:80/handle/2072/211024 Podocalyxin is a novel polysialylated neural adhesion protein with multiple roles in neural development and synapse formation Vitureira, N.; Andrés, R.; Perez Martinez, E.; Martínez García, Albert; Bribián Arruego,Ana; Blasi Cabús, Joan; Chelliah, S.; López Domenech, G.; De Castro, F.; Burgaya i Márquez, Ferran; McNagny, K.; Soriano García, Eduardo Neural development and plasticity are regulated by neural adhesion proteins, including the polysialylated form of NCAM (PSA-NCAM). Podocalyxin (PC) is a renal PSA-containing protein that has been reported to function as an anti-adhesin in kidney podocytes. Here we show that PC is widely expressed in neurons during neural development. Neural PC interacts with the ERM protein family, and with NHERF1/2 and RhoA/G. Experiments in vitro and phenotypic analyses of podxl-deficient mice indicate that PC is involved in neurite growth, branching and axonal fasciculation, and that PC loss-of-function reduces the number of synapses in the CNS and in the neuromuscular system. We also show that whereas some of the brain PC functions require PSA, others depend on PC per se. Our results show that PC, the second highly sialylated neural adhesion protein, plays multiple roles in neural development. http://www.recercat.cat:80/handle/2072/211024 http://www.recercat.cat:80/handle/2072/210802 Reelin controls progenitor cell migration in the healty and pathological adult brain Courtès, S.; Vernerey, J.; Pujadas Puigdomènech, Lluís; Magalon, K.; Cremer, H.; Soriano García, Eduardo; Durbec, P.; Cayre, M. Understanding the signals that control migration of neural progenitor cells in the adult brain may provide new therapeutic opportunities. Reelin is best known for its role in regulating cell migration during brain development, but we now demonstrate a novel function for reelin in the injured adult brain. First, we show that Reelin is upregulated around lesions. Second, experimentally increasing Reelin expression levels in healthy mouse brain leads to a change in the migratory behavior of subventricular zone-derived progenitors, triggering them to leave the rostral migratory stream (RMS) to which they are normally restricted during their migration to the olfactory bulb. Third, we reveal that Reelin increases endogenous progenitor cell dispersal in periventricular structures independently of any chemoattraction but via cell detachment and chemokinetic action, and thereby potentiates spontaneous cell recruitment to demyelination lesions in the corpus callosum. Conversely, animals lacking Reelin signaling exhibit reduced endogenous progenitor recruitment at the lesion site. Altogether, these results demonstrate that beyond its known role during brain development, Reelin is a key player in post-lesional cell migration in the adult brain. Finally our findings provide proof of concept that allowing progenitors to escape from the RMS is a potential therapeutic approach to promote myelin repair. http://www.recercat.cat:80/handle/2072/210802 http://www.recercat.cat:80/handle/2072/210803 GSK3 is involved in the relief of mitochondria pausing in a tau-dependent manner. Llorens Martín, M.; López Domenech, G.; Soriano García, Eduardo; Avila, J. Mitochondrial trafficking deficits have been implicated in the pathogenesis of several neurological diseases, including Alzheimer's disease (AD). The Ser/Thre kinase GSK3β is believed to play a fundamental role in AD pathogenesis. Given that GSK3β substrates include Tau protein, here we studied the impact of GSK3β on mitochondrial trafficking and its dependence on Tau protein. Overexpression of GSK3β in neurons resulted in an increase in motile mitochondria, whereas a decrease in the activity of this kinase produced an increase in mitochondria pausing. These effects were dependent on Tau proteins, as Tau (−/−) neurons did not respond to distinct GSK3β levels. Furthermore, differences in GSK3β expression did not affect other parameters like mitochondria velocity or mitochondria run length. We conclude that GSK3B activity regulates mitochondrial axonal trafficking largely in a Tau-dependent manner. http://www.recercat.cat:80/handle/2072/210803 http://www.recercat.cat:80/handle/2072/210801 A signaling mechanism coupling netrin-1/deleted in colorectal cancer chemoattraction to SNARE-mediated exocytosis in axonal growth cones Cotrufo, Tiziana; Pérez-Brangulí, F.; Muhaisen, A.; Ros, O.; Andrés, R.; Baeriswyl, T.; Fuschini, G.; Tarragó i Clua, Teresa; Pascual Sánchez, Marta; Ureña Bares, Jesús Mariano; Blasi Cabús, Joan; Giralt Lledó, Ernest; Stoeckli, E.T.; Soriano García, Eduardo Directed cell migration and axonal guidance are essential steps in neural development. Both processes are controlled by specific guidance cues that activate the signaling cascades that ultimately control cytoskeletal dynamics. Another essential step in migration and axonal guidance is the regulation of plasmalemma turnover and exocytosis in leading edges and growth cones. However, the cross talk mechanisms linking guidance receptors and membrane exocytosis are not understood. Netrin-1 is a chemoattractive cue required for the formation of commissural pathways. Here, we show that the Netrin-1 receptor deleted in colorectal cancer (DCC) forms a protein complex with the t-SNARE (target SNARE) protein Syntaxin-1 (Sytx1). This interaction is Netrin-1 dependent both in vitro and in vivo, and requires specific Sytx1 and DCC domains. Blockade of Sytx1 function by using botulinum toxins abolished Netrin-1-dependent chemoattraction of axons in mouse neuronal cultures. Similar loss-of-function experiments in the chicken spinal cord in vivo using dominant-negative Sytx1 constructs or RNAi led to defects in commissural axon pathfinding reminiscent to those described in Netrin-1 and DCC loss-of-function models. We also show that Netrin-1 elicits exocytosis at growth cones in a Sytx1-dependent manner. Moreover, we demonstrate that the Sytx1/DCC complex associates with the v-SNARE (vesicle SNARE) tetanus neurotoxin-insensitive vesicle-associated membrane protein (TI-VAMP) and that knockdown of TI-VAMP in the commissural pathway in the spinal cord results in aberrant axonal guidance phenotypes. Our data provide evidence of a new signaling mechanism that couples chemotropic Netrin-1/DCC axonal guidance and Sytx1/TI-VAMP SNARE proteins regulating membrane turnover and exocytosis. http://www.recercat.cat:80/handle/2072/210801 http://www.recercat.cat:80/handle/2072/210800 Effects of neurotrophins on synaptic protein expression in the visual cortex of dark reared rats.      Cotrufo, Tiziana; Viegi, A.; Mascia, L.; Berardi, N.; Maffei, L. Total lack of visual experience [dark rearing (DR)] is known to prolong the critical period and delay development of sensory functions in mammalian visual cortex. Recent results show that neurotrophins (NTs) counteract the effects of DR on functional properties of visual cortical cells and exert a strong control on critical period duration. NTs are known to modulate the development and synaptic efficacy of neurotransmitter systems that are affected by DR. However, it is still unknown whether the actions of NTs in dark-reared animals involve interaction with neurotransmitter systems. We have studied the effects of DR on the expression of key molecules in the glutamatergic and GABAergic systems in control and NT-treated animals. We have found that DR reduced the expression of the NMDA receptor 2A subunit and its associated protein PSD-95 (postsynaptic density-95), of GRIP (AMPA glutamate receptor interacting protein), and of the biosynthetic enzyme GAD (glutamic acid decarboxylase). Returning dark-reared animals to light for 2 hr restored normal expression of the above-mentioned proteins almost completely. NT treatment specifically counteracts DR effects; NGF acts primarily on the NMDA system, whereas BDNF acts primarily on the GABAergic system. Finally, the action of NT4 seems to involve both excitatory and inhibitory systems. These data demonstrate that different NTs counteract DR effects by modulating the expression of key molecules of the excitatory and inhibitory neurotransmitter systems http://www.recercat.cat:80/handle/2072/210800 http://www.recercat.cat:80/handle/2072/210539 A highly expressed miR-101 isomiR is a functional silencing small RNA Llorens, Franc; Bañez-Coronel, M.; Pantano, L.; Río Fernández, José Antonio del; Ferrer, Isidro (Ferrer Abizanda); Estivill, Xavier; Martí, Eulàlia Background MicroRNAs (miRNAs) are short non-coding regulatory RNAs that control gene expression usually producing translational repression and gene silencing. High-throughput sequencing technologies have revealed heterogeneity at length and sequence level for the majority of mature miRNAs (IsomiRs). Most isomiRs can be explained by variability in either Dicer1 or Drosha cleavage during miRNA biogenesis at 5" or 3" of the miRNA (trimming variants). Although isomiRs have been described in different tissues and organisms, their functional validation as modulators of gene expression remains elusive. Here we have characterized the expression and function of a highly abundant miR-101 5"-trimming variant (5"-isomiR-101). Results The analysis of small RNA sequencing data in several human tissues and cell lines indicates that 5"-isomiR-101 is ubiquitously detected and a highly abundant, especially in the brain. 5"- isomiR-101 was found in Ago-2 immunocomplexes and complementary approaches showed that 5"-isomiR-101 interacted with different members of the silencing (RISC) complex. In addition, 5"-isomiR-101 decreased the expression of five validated miR-101 targets, suggesting that it is a functional variant. Both the binding to RISC members and the degree of silencing were less efficient for 5"-isomiR-101 compared with miR-101. For some targets, both miR-101 and 5"-isomiR-101 significantly decreased protein expression with no changes in the respective mRNA levels. Although a high number of overlapping predicted targets suggest similar targeted biological pathways, a correlation analysis of the expression profiles of miR-101 variants and predicted mRNA targets in human brains at different ages, suggest specific functions for miR-101- and 5"-isomiR-101. Conclusions These results suggest that isomiRs are functional variants and further indicate that for a given miRNA, the different isomiRs may contribute to the overall effect as quantitative and qualitative fine-tuners of gene expression. http://www.recercat.cat:80/handle/2072/210539 http://www.recercat.cat:80/handle/2072/210537 Multiple platform assessmant of the EGF dependent transcritpome by microarrays and deep TAG sequencing analysis Llorens, Franc; Hummel, Manuela; Pastor Durán, Xavier; Ferrer, Anna; Pluvinet, Raquel; Vivancos, Ama; Castillo, Ester; Iraola Guzmán, Susana; Mosquera, A.M.; Gonzalez, Eva; Lozano, J.; Ingham, M.; Dohm, J.C.; Noguera, Marc; Kofler, R.; Río Fernández, José Antonio del; Bayes, M.; Himmelbauer, H.; Sumoy, Lauro Abstract Background: Epidermal Growth Factor (EGF) is a key regulatory growth factor activating many processes relevant to normal development and disease, affecting cell proliferation and survival. Here we use a combined approach to study the EGF dependent transcriptome of HeLa cells by using multiple long oligonucleotide based microarray platforms (from Agilent, Operon, and Illumina) in combination with digital gene expression profiling (DGE) with the Illumina Genome Analyzer. Results: By applying a procedure for cross-platform data meta-analysis based on RankProd and GlobalAncova tests, we establish a well validated gene set with transcript levels altered after EGF treatment. We use this robust gene list to build higher order networks of gene interaction by interconnecting associated networks, supporting and extending the important role of the EGF signaling pathway in cancer. In addition, we find an entirely new set of genes previously unrelated to the currently accepted EGF associated cellular functions. Conclusions: We propose that the use of global genomic cross-validation derived from high content technologies (microarrays or deep sequencing) can be used to generate more reliable datasets. This approach should help to improve the confidence of downstream in silico functional inference analyses based on high content data. http://www.recercat.cat:80/handle/2072/210537 http://www.recercat.cat:80/handle/2072/210536 Effects of Enriched Physical and Social Environments on Motor Performance, Associative Learning, and Hippocampal Neurogenesis in Mice. Madroñal, N.; Lopez-Aracil, C.; Rangel, A.; Río Fernández, José Antonio del; Delgado-Garcia, J.M.; Gruart, A. We have studied the motor abilities and associative learning capabilities of adult mice placed in different enriched environments. Three-month-old animals were maintained for a month alone (AL), alone in a physically enriched environment (PHY), and, finally, in groups in the absence (SO) or presence (SOPHY) of an enriched environment. The animals' capabilities were subsequently checked in the rotarod test, and for classical and instrumental learning. The PHY and SOPHY groups presented better performances in the rotarod test and in the acquisition of the instrumental learning task. In contrast, no significant differences between groups were observed for classical eyeblink conditioning. The four groups presented similar increases in the strength of field EPSPs (fEPSPs) evoked at the hippocampal CA3-CA1 synapse across classical conditioning sessions, with no significant differences between groups. These trained animals were pulse-injected with bromodeoxyuridine (BrdU) to determine hippocampal neurogenesis. No significant differences were found in the number of NeuN/BrdU double-labeled neurons. We repeated the same BrdU study in one-month-old mice raised for an additional month in the above-mentioned four different environments. These animals were not submitted to rotarod or conditioned tests. Non-trained PHY and SOPHY groups presented more neurogenesis than the other two groups. Thus, neurogenesis seems to be related to physical enrichment at early ages, but not to learning acquisition in adult mice. http://www.recercat.cat:80/handle/2072/210536 http://www.recercat.cat:80/handle/2072/210538 Myelin-associated proteins block the migration of olfactory ensheathing cells: an in vitro study using single cell tracking and traction force microscopy Nocentini, Sara; Reginensi, D.; Garcia, S.; Carulla, Patricia; Moreno-Flores, M.; Wandosell, F.; Trepat Guixer, Xavier; Bribián Arruego,Ana; Río Fernández, José Antonio del Newly generated olfactory receptor axons grow from the peripheral to the central nervous system aided by olfactory ensheathing cells (OECs). Thus, OEC transplantation has emerged as a promising therapy for spinal cord injuries and for other neural diseases. However, these cells do not present a uniform population, but, instead, a functionally heterogeneous population that exhibits a variety of responses including adhesion, repulsion and crossover during cell-cell and cell-matrix interactions. Some studies report that the migratory properties of OECs are compromised by inhibitory molecules and potentiated by chemical gradients. Here, we demonstrated that rodent OECs express all the components of the Nogo Receptor complex and that their migration is blocked by Myelin. Next, we used cell tracking and traction force microscopy to analyze OEC migration and its mechanical properties over Myelin. Our data relate the absence of traction force of OEC with lower migratory capacity, which correlates with changes in the F-Actin cytoskeleton and focal adhesion distribution. Lastly, OEC traction force and migratory capacity is enhanced after cell incubation with the Nogo Receptor inhibitor NEP1-40. http://www.recercat.cat:80/handle/2072/210538 http://www.recercat.cat:80/handle/2072/210534 Involvement of Dab1 in APP processing and [beta]-amyloid deposition in sporadic Creutzfeldt-Jakob patients. Gavín Marín, Rosalina; Ferrer, Isidro (Ferrer Abizanda); Río Fernández, José Antonio del Alzheimer"s disease and prion pathologies (e.g., Creutzfeldt-Jakob disease) display profound neural lesions associated with aberrant protein processing and extracellular amyloid deposits. For APP processing, emerging data suggest that the adaptor protein Dab1 plays a relevant role in regulating its intracellular trafficking and secretase-mediated proteolysis. Although some data have been presented, a putative relationship between human prion diseases and Dab1/APP interactions is lacking. Therefore, we have studied the putative relation between Dab1, APP processing and Aβ deposition, targets in sCJD cases. Our biochemical results categorized two groups of sCJD cases, which also correlated with PrPsc types 1 and 2 respectively. One group, with PrPsc type 1 showed increased Dab1 phosphorylation, and lower βCTF production with an absence of Aβ deposition. The second sCJD group, which carried PrPsc type 2, showed lower levels of Dab1 phosphorylation and βCTF production, similar to control cases. Relevant Aβ deposition in the second sCJD group was measured. Thus, a direct correlation between Dab1 phosphorylation, Aβ deposition and PrPsc type in human sCJD is presented for the first time. http://www.recercat.cat:80/handle/2072/210534 http://www.recercat.cat:80/handle/2072/210535 Developmental expression of the oligodendrocyte myelin glycoprotein in the mouse telencephalon Gil, V.; Bichler, Z.; Lee, J.K.; Seira Oriach, Oscar; Llorens, Franc; Bribián Arruego,Ana; Morales, R.; Claverol-Tinture, E.; Soriano García, Eduardo; Sumoy, Lauro; Zheng, B.; Río Fernández, José Antonio del The oligodendrocyte myelin glycoprotein is a glycosylphosphatidylinositol-anchored protein expressed by neurons and oligodendrocytes in the CNS. Attempts have been made to identify the functions of the myelin-associated inhibitory proteins (MAIPs) after axonal lesion or in neurodegeneration. However, the developmental roles of some of these proteins and their receptors remain elusive. Recent studies indicate that NgR1 and the recently discovered receptor PirB restrict cortical synaptic plasticity. However, the putative factors that trigger these effects are unknown. Since Nogo-A is mostly associated with the endoplasmic reticulum and MAG appears late during development, the putative participation of OMgp should be considered. Here we examine the pattern of development of OMgp immunoreactive elements during mouse telencephalic development. OMgp immunoreactivity in the developing cortex follows the establishment of the thalamo-cortical barrel-field. At cellular level, we located OMgp neuronal membranes in dendrites and axons as well as in brain synaptosome fractions and axon varicosities. Lastly, the analysis of the barrel-field in OMgp-deficient mice revealed that although thalamo-cortical connections were formed, their targeting in layer IV was altered and numerous axons ectopically invaded layer II-III. Our data support the idea that early-expressed MAIPs play an active role during development and point to OMgp participating in thalamo-cortical connections. http://www.recercat.cat:80/handle/2072/210535 http://www.recercat.cat:80/handle/2072/210533 New insights into cellular prion protein (PrPc) functions: the 'ying and yang' of a relevant protein. Nicolas, O.; Gavín Marín, Rosalina; Río Fernández, José Antonio del The conversion of cellular prion protein (PrPc), a GPI-anchored protein, into a protease-K-resistant and infective form (generally termed PrPsc) is mainly responsible for Transmissible Spongiform Encephalopathies (TSEs), characterized by neuronal degeneration and progressive loss of basic brain functions. Although PrPc is expressed by a wide range of tissues throughout the body, the complete repertoire of its functions has not been fully determined. Recent studies have confirmed its participation in basic physiological processes such as cell proliferation and the regulation of cellular homeostasis. Other studies indicate that PrPc interacts with several molecules to activate signaling cascades with a high number of cellular effects. To determine PrPc functions, transgenic mouse models have been generated in the last decade. In particular, mice lacking specific domains of the PrPc protein have revealed the contribution of these domains to neurodegenerative processes. A dual role of PrPc has been shown, since most authors report protective roles for this protein while others describe pro-apoptotic functions. In this review, we summarize new findings on PrPc functions, especially those related to neural degeneration and cell signaling. http://www.recercat.cat:80/handle/2072/210533 http://www.recercat.cat:80/handle/2072/210419 Neurites regrowth of cortical neurons by GSK3b inhibition independently of Nogo Receptor 1 Seira Oriach, Oscar; Gavín Marín, Rosalina; Gil, V.; Llorens, Franc; Rangel, A.; Soriano García, Eduardo; Río Fernández, José Antonio del Lesioned axons do not regenerate in the adult mammalian central nervous system, owing to the overexpression of inhibitory molecules such as myelin-derived proteins or chondroitin sulphate proteoglycans. In order to overcome axon inhibition, strategies based on extrinsic and intrinsic treatments have been developed. For myelin-associated inhibition, blockage with NEP1-40, receptor bodies or IN-1 antibodies has been used. In addition, endogenous blockage of cell signalling mechanisms induced by myelin-associated proteins is a potential tool for overcoming axon inhibitory signals. We examined the participation of glycogen synthase kinase 3 (GSK3) and ERK1/2 in axon regeneration failure in lesioned cortical neurons. We also investigated whether pharmacological blockage of GSK3 and ERK1/2 activities facilitates regeneration after myelin-directed inhibition in two models: i) cerebellar granule cells and ii) lesioned entorhino-hippocampal pathway in slice cultures, and whether the regenerative effects are mediated by Nogo Receptor 1 (NgR1). We demonstrate that, in contrast to ERK1/2 inhibition, the pharmacological treatment of GSK3 inhibition strongly facilitated regrowth of cerebellar granule neurons over myelin independently of NgR1. Lastly these regenerative effects were corroborated in the lesioned EHP in NgR1 -/- mutant mice. These results provide new findings for the development of new assays and strategies to enhance axon regeneration in injured cortical connections. http://www.recercat.cat:80/handle/2072/210419 http://www.recercat.cat:80/handle/2072/210295 APP processing and b-amyloid deposition in sporadic Creutzfeldt-Jakob patients is dependent on Dab1. Gavín Marín, Rosalina; Ferrer, I.; del Río, José A. Alzheimer"s disease and prion pathologies (e.g., Creutzfeldt-Jakob disease (CJD)) display profound neural lesions associated with aberrant protein processing and extracellular amyloid deposits. Dab1 has been implicated in the regulation of Amyloid Precursor Protein (APP), but a direct link between human prion diseases and Dab1/APP interactions has not been published. Here we examined this putative relationship in seventeen cases of sporadic CJD (sCJD) post mortem. Biochemical analyses of brain tissue revealed two groups, which also correlated with PrPsc types 1 and 2. One group, with PrPsc type 1 showed increased Dab1 phosphorylation, and lower CTF production with an absence of A deposition. The second sCJD group, which carried PrPsc type 2, showed lower levels of Dab1 phosphorylation and CTF production, and A deposition. Thus, the present observations suggest a correlation between Dab1-phosphorylation, A deposition and PrPsc type in sCJD. http://www.recercat.cat:80/handle/2072/210295 http://www.recercat.cat:80/handle/2072/210294 New insights into cellular prion protein (PrPc) functions: the 'ying and yang' of a relevant protein. Nicolas, Oriol; Gavín Marín, Rosalina; del Río, José A. The conversion of cellular prion protein (PrPc), a GPI-anchored protein, into a protease-K-resistant and infective form (generally termed PrPsc) is mainly responsible for Transmissible Spongiform Encephalopathies (TSEs), characterized by neuronal degeneration and progressive loss of basic brain functions. Although PrPc is expressed by a wide range of tissues throughout the body, the complete repertoire of its functions has not been fully determined. Recent studies have confirmed its participation in basic physiological processes such as cell proliferation and the regulation of cellular homeostasis. Other studies indicate that PrPc interacts with several molecules to activate signaling cascades with a high number of cellular effects. To determine PrPc functions, transgenic mouse models have been generated in the last decade. In particular, mice lacking specific domains of the PrPc protein have revealed the contribution of these domains to neurodegenerative processes. A dual role of PrPc has been shown, since most authors report protective roles for this protein while others describe pro-apoptotic functions. In this review, we summarize new findings on PrPc functions, especially those related to neural degeneration and cell signaling. http://www.recercat.cat:80/handle/2072/210294 http://www.recercat.cat:80/handle/2072/209453 Role of the cellular prion protein in oligodendrocyte precursor cell proliferation and differentiation in the developing and adult mouse CNS Bribián Arruego,Ana; Fontana, X.; Llorens, Franc; Gavín Marín, Rosalina; Reina del Pozo, Manuel; García-Verdugo, J.M.; Torres, Juan María; de Castro, F.; Río Fernández, José Antonio del There are numerous studies describing the signaling mechanisms that mediate oligodendrocyte precursor cell (OPC) proliferation and differentiation, although the contribution of the cellular prion protein (PrPc) to this process remains unclear. PrPc is a glycosyl-phosphatidylinositol (GPI)-anchored glycoprotein involved in diverse cellular processes during the development and maturation of the mammalian central nervous system (CNS). Here we describe how PrPc influences oligodendrocyte proliferation in the developing and adult CNS. OPCs that lack PrPc proliferate more vigorously at the expense of a delay in differentiation, which correlates with changes in the expression of oligodendrocyte lineage markers. In addition, numerous NG2-positive cells were observed in cortical regions of adult PrPc knockout mice, although no significant changes in myelination can be seen, probably due to the death of surplus cells. http://www.recercat.cat:80/handle/2072/209453 http://www.recercat.cat:80/handle/2072/206524 La función activadora de un apósito bioactivo con carga iónica sobre los fibroblastos humanos Castellarnau Castella, Conxita; Martín, Mercè; Marcos, Anna; Casaroli Marano, Ricardo Pedro; Reina del Pozo, Manuel; Vilaró, Senén, 1956-2005 La cicatrización de las heridas es un proceso complejo que implica diferentes fases que se solapan entre sí, incluyendo la inflamación, la epitelización, la angiognesis y la síntesis y deposición de matriz extracelular. Los fibroblastos dérmicos tienen una función esencial en la formación del tejido de granulación. Migran hasta la lesión en respuesta a citoquinas, proliferan y sintetizan las proteínas de la matriz extracelular, las cuales son la base del proceso de reparación futuro. Los oligoelementos tales como el zinc y el manganeso son necesarios para muchas funciones celulares y, por consiguiente, pueden potencialmente estimular los procesos de reparación de las heridas. En el presente estudio hemos investigado el efecto de un aposito que contiene zinc, calcio y manganeso (Trionic®) sobre la proliferación, el crecimiento, la síntesis de colágeno I y III y la migración de los fibroblastos. Los resultados obtenidos indican que los oligoelementos solubles presentes en Trionic® actúan estimulando la proliferación, el crecimiento, la biosíntesis de colágeno y la migración de los fibroblastos. Dada la participación crucial que estas funciones celulares tienen sobre el comportamiento de los fibroblastos durante el proceso de formación del tejido de granulación, concluimos que los iones Ca2+, Zn2+, y Mn2+ contenidos en Trionic® pueden proporcionar potenciales beneficios en el tratamiento de las heridas crónicas y durante la fase reparativa del proceso de cicatrización. http://www.recercat.cat:80/handle/2072/206524 http://www.recercat.cat:80/handle/2072/203108 Ultrastructural studies of oogenesis in Bolinus brandaris(Gastropoda: Muricidae). Amor Pérez, Ma. José (María José); Ramón Herrero, Montserrat; Durfort i Coll, Mercè Ultrastructural studies of oogenesis in Bolinus brandaris are described. Although the initial phase of oogenesis is common to most animal species, vitellogenesis can be considered a species-specific characteristic. In the vitellogenesis of B.brandaris, mitochondria and endoplasmic reticula play a relevant role in the formation of myelinised membranous systems. Nuclear envelope, Golgi body and the oocyte plasma membrane invaginations are three possible origins for annulate lamellae. The latter can be considered membranous reservoirs. There are two sources for the vitellum: exogenous (from follicular cells) and endogenous (from the endoplasmic reticulum of the same oocyte). http://www.recercat.cat:80/handle/2072/203108 http://www.recercat.cat:80/handle/2072/203107 Ultrastructural data on the life cycle of the parasite, Perkinsus atlanticus (Apicomplexa), on the clam, Ruditapes phillipinarum, in the Mediterranean Sagristà i Mateo, Elena; Durfort i Coll, Mercè; Azevedo, Carlos An Apicomplexan Perkinsus species has been found parasitizing the clam Ruditapes philippinarum (= Tapes semidecussatus) collected on the Mediterranean coast in the region of the Ebro Delta (Tarragona, Spain). Light and transmission electron microscopy were used to study different stages of this parasite during zoosporulation induced by incubation in thioglycollate medium and seawater. During incubation the trophozoites began zoosporulation, which originated prezoosporangia and zoosporangia at different developmental stages. Successive cytokinesis and nucleokinesis gave rise to prezoospores, which became elongate and differentiated in biflagellated zoospores. The latter presented large mitochondria and an apical complex formed by a conoid, polar ring, micronemes, rhophtries and subpellicular microtubules. The zoosporangium wall showed some typical lamosomes and a discharge tube developed in early phases of incubation. Ultrastructural data were compared with the only four species of the genus Perkinsus previously described. The morphological data, the host and the geographic proximity suggest that the species located on the Mediterranean coast was Perkinsus atlanticus. http://www.recercat.cat:80/handle/2072/203107 http://www.recercat.cat:80/handle/2072/196363 Neuroprotective role of PrPC against kainate-induced epileptic seizures and cell death depends on the modulation of JNK3 activation by GluR6/7-PSD-95 binding Carulla, Patricia; Bribián Arruego,Ana; Rangel Rincones, Alejandra Helena; Gavín Marín, Rosalina; Ferrer, Isidro (Ferrer Abizanda); Caelles Franch, Carme; Río Fernández, José Antonio del; Llorens, Franc Cellular prion protein (PrPC) is a glycosyl-phosphatidylinositol¿anchored glycoprotein. When mutated or misfolded, the pathogenic form (PrPSC) induces transmissible spongiform encephalopathies. In contrast, PrPC has a number of physiological functions in several neural processes. Several lines of evidence implicate PrPC in synaptic transmission and neuroprotection since its absence results in an increase in neuronal excitability and enhanced excitotoxicity in vitro and in vivo. Furthermore, PrPC has been implicated in the inhibition of N-methyl-D-aspartic acid (NMDA)¿mediated neurotransmission, and prion protein gene (Prnp) knockout mice show enhanced neuronal death in response to NMDA and kainate (KA). In this study, we demonstrate that neurotoxicity induced by KA in Prnp knockout mice depends on the c-Jun N-terminal kinase 3 (JNK3) pathway since Prnpo/oJnk3o/o mice were not affected by KA. Pharmacological blockage of JNK3 activity impaired PrPC-dependent neurotoxicity. Furthermore, our results indicate that JNK3 activation depends on the interaction of PrPC with postsynaptic density 95 protein (PSD-95) and glutamate receptor 6/7 (GluR6/7). Indeed, GluR6¿PSD-95 interaction after KA injections was favored by the absence of PrPC. Finally, neurotoxicity in Prnp knockout mice was reversed by an AMPA/KA inhibitor (6,7-dinitroquinoxaline-2,3-dione) and the GluR6 antagonist NS-102. We conclude that the protection afforded by PrPC against KA is due to its ability to modulate GluR6/7-mediated neurotransmission and hence JNK3 activation. http://www.recercat.cat:80/handle/2072/196363 http://www.recercat.cat:80/handle/2072/196362 ß-Catenin Regulation during the Cell Cycle: Implications in G2/M and Apoptosis Olmeda, David; Castel i Gil, Susanna; Vilaró, Senén, 1956-2005; Cano, Amparo ß-catenin is a multifunctional protein involved in cell-cell adhesion and Wnt signal transduction. ß-Catenin signaling has been proposed to act as inducer of cell proliferation in different tumors. However, in some developmental contexts and cell systems ß-catenin also acts as a positive modulator of apoptosis. To get additional insights into the role of ß-Catenin in the regulation of the cell cycle and apoptosis, we have analyzed the levels and subcellular localization of endogenous ß-catenin and its relation with adenomatous polyposis coli (APC) during the cell cycle in S-phase¿synchronized epithelial cells. ß-Catenin levels increase in S phase, reaching maximum accumulation at late G2/M and then abruptly decreasing as the cells enter into a new G1 phase. In parallel, an increased cytoplasmic and nuclear localization of ß-catenin and APC is observed during S and G2 phases. In addition, strong colocalization of APC with centrosomes, but not ß-catenin, is detected in M phase. Interestingly, overexpression of a stable form of ß-catenin, or inhibition of endogenous ß-catenin degradation, in epidermal keratinocyte cells induces a G2 cell cycle arrest and leads to apoptosis. These results support a role for ß-catenin in the control of cell cycle and apoptosis at G2/M in normal and transformed epidermal keratinocytes. http://www.recercat.cat:80/handle/2072/196362 http://www.recercat.cat:80/handle/2072/195768 Assembly and disassembly of the Golgi Complex: two prodesses arranged in a cis to trans direction Alcalde, José; Bonay, Pedro; Roa, Ana; Vilaró, Senén, 1956-2005; Sandoval, Ignacio V. We have studied the disassembly and assembly of two morphologically and functionally distinct parts of the Golgi complex, the cis/middle and trans cisterna/trans network compartments. For this purpose we have followed the redistribution of three cis/middle- (GMPc-1, GMPc-2, MG 160) and two trans- (GMPt-1 and GMPt-2) Golgi membrane proteins during and after treatment of normal rat kidney (NRK) cells with brefeldin A (BFA). BFA induced complete disassembly of the cis/middle- and trans-Golgi complex and translocation of GMPc and GMPt to the ER. Cells treated for short times (3 min) with BFA showed extensive disorganization of both cis/middle- and trans-Golgi complexes. However, complete disorganization of the trans part required much longer incubations with the drug. Upon removal of BFA the Golgi complex was reassembled by a process consisting of three steps: (a) exist of cis/middle proteins from the ER and their accumulation into vesicular structures scattered throughout the cytoplasm; (b) gradual relocation and accumulation of the trans proteins in the vesicles containing the cis/middle proteins; and (c) assembly of the cisternae, and reconstruction of the Golgi complex within an area located in the vicinity of the centrosome from which the ER was excluded. Reconstruction of the cis/middle-Golgi complex occurred under temperature conditions inhibitory of the reorganization of the trans-Golgi complex, and was dependent on microtubules. Reconstruction of the trans-Golgi complex, disrupted with nocodazole after selective fusion of the cis/middle-Golgi complex with the ER, occurred after the release of cis/middle-Golgi proteins from the ER and the assembly of the cis/middle cisternae. http://www.recercat.cat:80/handle/2072/195768 http://www.recercat.cat:80/handle/2072/182329 Nogo, myelin and axonal regeneration Mingorance Jiménez, Alfredo; Soriano García, Eduardo; Río Fernández, José Antonio del Adult mammalian central nervous system (CNS) axons have very limited capacity of regrowth after injury. In recent years, advances in the field of axonal regeneration have proved that neurons do not regenerate, mainly because of the presence of inhibitory molecules. Myelin-associated proteins limit axonal outgrowth and their blockage improves the regeneration of damaged fiber tracts. Three of these proteins, Nogo, MAG and OMgp, share a common neuronal receptor (NgR), and together represent one of the main hindrances to neuronal regeneration. The recent molecular cloning of Nogo and its receptors opened a new door to the study of axon regeneration. However, many of the elements involved in the myelin inhibitory pathway are still unknown, and the preliminary experiments with knockout mice are rather contradictory. Because of this complexity, Nogo and NgR need to be characterized before precise strategies to promote axon regeneration in the CNS can be designed. http://www.recercat.cat:80/handle/2072/182329 http://www.recercat.cat:80/handle/2072/169197 Changes in the enterocyte cytoskeleton in newborn rats exposed to ethanol in utero Montes Castillo, Juan Francisco; Estrada i Bertran, Gemma; López Tejero, M. Dolores; García Valero, Josep BACKGROUND: Cytoskeletal changes after longterm exposure to ethanol have been described in a number of cell types in adult rat and humans. These changes can play a key part in the impairment of nutrient assimilation and postnatal growth retardation after prenatal damage of the intestinal epithelium produced by ethanol intake. AIMS: To determine, in the newborn rat, which cytoskeletal proteins are affected by longterm ethanol exposure in utero and to what extent. ANIMALS: The offspring of two experimental groups of female Wistar rats: ethanol treated group receiving up to 25% (w/v) of ethanol in the drinking fluid and control group receiving water as drinking fluid. METHODS: Single and double electron microscopy immunolocalisation and label density estimation of cytoskeletal proteins on sections of proximal small intestine incubated with monoclonal antibodies against actin, alpha-tubulin, cytokeratin (polypeptides 1, 5, 6, 7, 8, 10, 11, and 18), and with a polyclonal antibody anti-beta 1,4-galactosyl transferase as trans golgi (TG) or trans golgi network (TGN) marker, or both. SDS-PAGE technique was also performed on cytoskeletal enriched fractions from small intestine. Western blotting analysis was carried out by incubation with the same antibodies used for immunolocalisation. RESULTS: Intestinal epithelium of newborn rats from the ethanol treated group showed an overexpression of cytoskeletal polypeptides ranging from 39 to 54 kDa, affecting actin and some cytokeratins, but not tubulin. Furthermore, a cytokeratin related polypeptide of 28-29 kDa was identified together with an increase in free ubiquitin in the same group. It was noteworthy that actin and cytokeratin were abnormally located in the TG or the TGN, or both. CONCLUSIONS: Longterm exposure to ethanol in utero causes severe dysfunction in the cytoskeleton of the developing intestinal epithelium. Actin and cytokeratins, which are involved in cytoskeleton anchoring to plasma membrane and cell adhesion, are particularly affected, showing overexpression, impaired proteolysis, and mislocalisation. http://www.recercat.cat:80/handle/2072/169197 http://www.recercat.cat:80/handle/2072/117300 Les comunitats segetals de la Cerdanya. Consideracions generals sobre la vegetació medioeuropea de la classe Secalietea a Catalunya Carreras i Raurell, Jordi; Carrillo, Empar; Font i Castell, Xavier; Masalles i Saumell, Ramon M., 1948-; Ninot i Sugrañes, Josep Maria; Soriano i Tomàs, Ignasi; Vigo, Josep, 1937- Aquest article estudia la vegetació dels sembrats (classe Secalietea Br.-Bl. 1952) de la Baixa Cerdanya i zones adjacents mitjançant el mètode fitocenològic i fa un repàs d'aquest tipus de comunitats a la Catalunya extramediterrània. Els sembrats dels terrenys àcids són referits a l'associació Scleranthetum anuui Br.-Bl. 1915, de l'aliança Scleranthion annui, força rara a la Cerdanya. Pel que fa a les comunitats neutròfiles, adscrites a l'aliança Caucalidion lappulae, discutim de primer el tractament sintaxonòmic rebut per aquest tipus de vegetació als Pirineus i altres zones peninsulars en treballs precedents. Hi reconeixem tres associacions: Kickxio-Nigelletum gallicae Fanlo 1988, coneguda només dels Pirineus centrals; Violo-Legousietum Hybridae O. Bolòs 1959, estesa pels Prepirineus calcaris i la Catalunya central; i Biforo radiantis-Centaureetum cyani Vigo, Carreras, Carrillo et I. Soriano, ass nova, dins de la qual proposem d'incloure les comunitats cerdanes http://www.recercat.cat:80/handle/2072/117300 http://www.recercat.cat:80/handle/2072/117299 L'anatomia microscòpica de les plantes Durfort i Coll, Mercè The microscopic anatomy of plants. A review of the main treatises on Plant Microscopic Anatomy and their translations published along the XX century is followed by a discussion on various aspects of the structure in plant cells, methodological questions and teminology. Problems related to using dry specimens from herbaria for microscopic studies are considered. As an example, a study has been made on a species named in memory of Prof. Dr. Oriol de Bolòs, Delphinium bolosii (BLANCHÉ and MOLERO, 1983).; Unes consideracions prèvies de caire bibliogràfic sobre els principals tractats d'anatomia microscòpica vegetal i llurs traduccions en el decurs del segle XX. deixen pas a unes reflexions sobre determinats aspectes de l'estructura cel·lular, sobre qüestions metodològiques així com sobre matisos de tipus terminològic. Es planteja la problemàtica de les plantes de l'herbari per a fer-ne estudis microscòpics de Qüalitat, i com a model il·lustratiu d'aquesta dificultat s'ha escollit una espècie que va ser dedicada al professor Dr. Oriol de Bolòs, el Delphinium bolosii (BLANCHË and MOLERO, 1983). http://www.recercat.cat:80/handle/2072/117299 http://www.recercat.cat:80/handle/2072/117298 Some reflections on Geobotany and Vegetation mapping Vigo, Josep, 1937- uivalent Spanish and Catalan terms. http://www.recercat.cat:80/handle/2072/117298 http://www.recercat.cat:80/handle/2072/49667 Regulation of GABA(A) and glutamate receptor expression, synaptic facilitation and long-term potentiation in the hippocampus of prion mutant mice Rangel Rincones, Alejandra Helena; Madronal, Noelia; Gruart i Massó, Agnès; Gavín Marín, Rosalina; Llorens, Franc; Sumoy, Lauro; Torres, Juan María; Delgado-García, José María; Río Fernández, José Antonio del Background: Prionopathies are characterized by spongiform brain degeneration, myoclonia, dementia, and periodic electroencephalographic (EEG) disturbances. The hallmark of prioniopathies is the presence of an abnormal conformational isoform (PrP(sc)) of the natural cellular prion protein (PrP(c)) encoded by the Prnp gene. Although several roles have been attributed to PrP(c), its putative functions in neuronal excitability are unknown. Although early studies of the behavior of Prnp knockout mice described minor changes, later studies report altered behavior. To date, most functional PrP(c) studies on synaptic plasticity have been performed in vitro. To our knowledge, only one electrophysiological study has been performed in vivo in anesthetized mice, by Curtis and coworkers. They reported no significant differences in paired-pulse facilitation or LTP in the CA1 region after Schaffer collateral/commissural pathway stimulation. Principal Findings: Here we explore the role of PrP(c) expression in neurotransmission and neural excitability using wild-type, Prnp -/- and PrP(c)-overexpressing mice (Tg20 strain). By correlating histopathology with electrophysiology in living behaving mice, we demonstrate that both Prnp -/- mice but, more relevantly Tg20 mice show increased susceptibility to KA, leading to significant cell death in the hippocampus. This finding correlates with enhanced synaptic facilitation in paired-pulse experiments and hippocampal LTP in living behaving mutant mice. Gene expression profiling using Illumina microarrays and Ingenuity pathways analysis showed that 129 genes involved in canonical pathways such as Ubiquitination or Neurotransmission were co-regulated in Prnp -/- and Tg20 mice. Lastly, RT-qPCR of neurotransmission-related genes indicated that subunits of GABA(A) and AMPA-kainate receptors are co-regulated in both Prnp -/- and Tg20 mice. Conclusions/Significance: Present results demonstrate that PrP(c) is necessary for the proper homeostatic functioning of hippocampal circuits, because of its relationships with GABA(A) and AMPA-Kainate neurotransmission. New PrP(c) functions have recently been described, which point to PrP(c) as a target for putative therapies in Alzheimer's disease. However, our results indicate that a "gain of function" strategy in Alzheimer's disease, or a "loss of function" in prionopathies, may impair PrP(c) function, with devastating effects. In conclusion, we believe that present data should be taken into account in the development of future therapies. http://www.recercat.cat:80/handle/2072/49667