The 39 untranslated region (39UTR) of the dengue virus (DENV) genome contain several sequences required for translation, replication and cyclization processes. This region also binds cellular proteins such as La, polypyrimidine tract-binding protein (PTB), Y box-binding protein 1, poly(A)-binding protein and the translation initiation factor eEF-1a. PTB is a cellular protein that interacts with the regulatory sequences of positive-strand RNA viruses such as several picornaviruses and hepatitis C virus. In the present report, it was demonstrated that PTB translocates from the nucleus to the cytoplasm during DENV infection. At 48 h post-infection, PTB, as well as the DENV proteins NS1 and NS3, were found to co-localize with the endoplasmic reticulum marker calnexin. Silencing of PTB expression inhibited virus translation and replication, whilst overexpression of PTB augmented these processes. Thus, these results provide evidence that, during infection, PTB moves from the nucleus to the cytoplasm and plays an important role in the DENV replicative cycle. INTRODUCTIONDengue virus (DENV), a member of the family Flaviviridae, is the causative agent of dengue fever, dengue haemorrhagic fever and dengue shock syndrome. The single-stranded, positive-polarity RNA genome of approximately 11 kb contains a type I cap at the 59 end and lacks a poly(A) tail at the 39 end. The single open reading frame (ORF) encodes a polyprotein that generates three structural proteins -envelope (E), membrane and capsid (C) -and seven non-structural proteins -NS1, NS2A, NS2B, NS3, NS4A, NS4B and NS5. Flanking the ORF, the viral RNA contains two untranslated regions (UTRs) involved in various functions such as initiation and regulation of virus translation, replication and assembly (Lindenbach & Rice, 2001;Proutski et al., 1999). The 39UTR of DENV and other mosquito-borne flaviviruses contains a conserved stemloop structure within the last~96 nt (Brinton & Dispoto, 1988;Brinton et al., 1986;Grange et al., 1985;Mohan & Padmanabhan, 1991). Additionally, there are two pairs of conserved sequences (59CS1, 39CS1, 59UAR and 39UAR) that together induce DENV cyclization (Alvarez et al., 2005a, b; Hahn et al., 1987). These motifs are essential for negativestrand RNA synthesis of DENV (Ackermann & Padmanabhan, 2001; Alvarez et al., 2005a, b;Villordo & Gamarnik, 2009;You & Padmanabhan, 1999) and other flaviviruses (Bredenbeek et al., 2003;Corver et al., 2003;Jones et al., 2005;Khromykh et al., 2001;Lo et al., 2003;Nomaguchi et al., 2004). On the other hand, sequences present within a large stem-loop structure located at the 59 end as well as a conserved oligo(U) track function as the promoter for viral polymerase activity (Lodeiro et al., 2009). Moreover, an RNA secondary structure present in the coding region of DENV type 2 (DENV-2) directs translation, start-codon selection and replication of the viral genome (Clyde & Harris, 2006;Clyde et al., 2008). Although it has been shown that the cyclization process does not require the presence of cellular or v...

The exchange of substances between higher organisms and the environment occurs across transporting epithelia whose basic features are tight junctions (TJs) that seal the intercellular space, and polarity, which enables cells to transport substances vectorially. In a previous study, we demonstrated that 10 nM ouabain modulates TJs, and we now show that it controls polarity as well. We gauge polarity through the development of a cilium at the apical domain of Madin-Darby canine kidney cells (MDCK, epithelial dog kidney). Ouabain accelerates ciliogenesis in an ERK1/2-dependent manner. Claudin-2, a molecule responsible for the Na + and H 2 O permeability of the TJs, is also present at the cilium, as it colocalizes and coprecipitates with acetylated α-tubulin. Ouabain modulates claudin-2 localization at the cilium through ERK1/2. Comparing wild-type and ouabain-resistant MDCK cells, we show that ouabain acts through Na + ,K + -ATPase. Taken together, our previous and present results support the possibility that ouabain constitutes a hormone that modulates the transporting epithelial phenotype, thereby playing a crucial role in metazoan life.E-cadherin | occludin | cell adhesion | cardiotonic steroids

Summary Upon activation with physiological stimuli, human platelets undergo morphological changes, centralizing their organelles and secreting effector molecules at the site of vascular injury. Previous studies have indicated that the actin filaments and microtubules of suspension‐activated platelets play a critical role in granule movement and exocytosis; however, the participation of these cytoskeleton elements in adhered platelets remains unexplored. α‐ and β‐dystrobrevin members of the dystrophin‐associated protein complex in muscle and non‐muscle cells have been described as motor protein receptors that might participate in the transport of cellular components in neurons. Recently, we characterized the expression of dystrobrevins in platelets; however, their functional diversity within this cellular model had not been elucidated. The present study examined the contribution of actin filaments and microtubules in granule trafficking during the platelet adhesion process using cytoskeleton‐disrupting drugs, quantification of soluble P‐selectin, fluorescence resonance transfer energy analysis and immunoprecipitation assays. Likewise, we assessed the interaction of α‐dystrobrevins with the ubiquitous kinesin heavy chain. Our results strongly suggest that microtubules and actin filaments participate in the transport of alpha and dense granules in the platelet adhesion process, during which α‐dystrobrevins play the role of regulatory and adaptor proteins that govern trafficking events.

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To maintain the continuity of an injured blood vessel, platelets change shape, secrete granule contents, adhere, aggregate, and retract in a haemostatic plug. Ordered arrays of microtubules, microfilaments, and associated proteins are responsible for these platelet responses. In full-spread platelets, microfilament bundles in association with other cytoskeleton proteins are anchored in focal contacts. Recent studies in migrating cells suggest that co-ordination and direct physical interaction of microtubules and actin network modulate adhesion development. In platelets, we have proposed a feasible association between these two cytoskeletal systems, as well as the participation of the dystrophin-associated protein complex, as part of the focal adhesion complex. The present study analysed the participation of microtubules and actin during the platelet adhesion process. Confocal microscopy, fluorescence resonance transfer energy and immunoprecipitation assays were used to provide evidence of a cross-talk between these two cytoskeletal systems. Interestingly, beta-dystroglycan was found to act as an interplay protein between actin and microtubules and an additional communication between these two cytoskeleton networks was maintained through proteins of focal adhesion complex. Altogether our data are indicative of a dynamic co-participation of actin filaments and microtubules in modulating focal contacts to achieve platelet function.

This fMRI work studies brain activity of healthy volunteers who manipulated a virtual object in the context of a digital game by applying two different control methods: using their right hand or using their gaze. The results show extended activations in sensorimotor areas, not only when participants played in the traditional way (using their hand) but also when they used their gaze to control the virtual object. Furthermore, with the exception of the primary motor cortex, regional motor activity was similar regardless of what the effector was: the arm or the eye. These results have a potential application in the field of the neurorehabilitation as a new approach to generate activation of the sensorimotor system to support the recovery of the motor functions.

The role of platelets in coagulation and the haemostatic process was initially suggested two centuries ago, and under appropriate physiological stimuli, these undergo abrupt morphological changes, attaching and spreading on damaged endothelium, preventing bleeding. During the adhesion process, platelet cytoskeleton reorganizes generating compartments in which actin filaments, microtubules, and associated proteins are arranged in characteristic patterns mediating crucial events, such as centralization of their organelles, secretion of granule contents, aggregation with one another to form a haemostatic plug, and retraction of these aggregates. However, the role of Intermediate filaments during the platelet adhesion process has not been explored. J. Cell. Biochem. 114: 2050-2060, 2013. © 2013 Wiley Periodicals, Inc.