BPAG1a and b Associate with EB1 and EB3 and Modulate Vesicular Transport, Golgi Apparatus Structure, and Cell Migration in C2.7 Myoblasts

BPAG1a and BPAG1b (BPAG1a/b) constitute two major isoforms encoded by the dystonin (Dst) gene and show homology with MACF1a and MACF1b. These proteins are members of the plakin family, giant multi-modular proteins able to connect the intermediate filament, microtubule and microfilament cytoskeletal networks with each other and to distinct cell membrane sites. They also serve as scaffolds for signaling proteins that modulate cytoskeletal dynamics. To gain better insights into the functions of BPAG1a/b, we further characterized their C-terminal region important for their interaction with microtubules and assessed the role of these isoforms in the cytoskeletal organization of C2.7 myoblast cells. Our results show that alternative splicing does not only occur at the 5′ end of Dst and Macf1 pre-mRNAs, as previously reported, but also at their 3′ end, resulting in expression of additional four mRNA variants of BPAG1 and MACF1. These isoform-specific C-tails were able to bundle microtubules and bound to both EB1 and EB3, two microtubule plus end proteins. In the C2.7 cell line, knockdown of BPAG1a/b had no major effect on the organization of the microtubule and microfilament networks, but negatively affected endocytosis and maintenance of the Golgi apparatus structure, which became dispersed. Finally, knockdown of BPAG1a/b caused a specific decrease in the directness of cell migration, but did not impair initial cell adhesion. These data provide novel insights into the complexity of alternative splicing of Dst pre-mRNAs and into the role of BPAG1a/b in vesicular transport, Golgi apparatus structure as well as in migration in C2.7 myoblasts

Cryo Electron Tomography of Herpes Simplex Virus during Axonal Transport and Secondary Envelopment in Primary Neurons

During herpes simplex virus 1 (HSV1) egress in neurons, viral particles travel from the neuronal cell body along the axon towards the synapse. Whether HSV1 particles are transported as enveloped virions as proposed by the ‘married’ model or as non-enveloped capsids suggested by the ‘separate’ model is controversial. Specific viral proteins may form a recruitment platform for microtubule motors that catalyze such transport. However, their subviral location has remained elusive. Here we established a system to analyze herpesvirus egress by cryo electron tomography. At 16 h post infection, we observed intra-axonal transport of progeny HSV1 viral particles in dissociated hippocampal neurons by live-cell fluorescence microscopy. Cryo electron tomography of frozen-hydrated neurons revealed that most egressing capsids were transported independently of the viral envelope. Unexpectedly, we found not only DNA-containing capsids (cytosolic C-capsids), but also capsids lacking DNA (cytosolic A-/B-capsids) in mid-axon regions. Subvolume averaging revealed lower amounts of tegument on cytosolic A-/B-capsids than on C-capsids. Nevertheless, all capsid types underwent active axonal transport. Therefore, even few tegument proteins on the capsid vertices seemed to suffice for transport. Secondary envelopment of capsids was observed at axon terminals. On their luminal face, the enveloping vesicles were studded with typical glycoprotein-like spikes. Furthermore, we noted an accretion of tegument density at the concave cytosolic face of the vesicle membrane in close proximity to the capsids. Three-dimensional analysis revealed that these assembly sites lacked cytoskeletal elements, but that filamentous actin surrounded them and formed an assembly compartment. Our data support the ‘separate model’ for HSV1 egress, i.e. progeny herpes viruses being transported along axons as subassemblies and not as complete virions within transport vesicles

Etude et mise en place d'une plate-forme de télé rééducation orthoptiste

International audienceLa plate forme TéOdevie, développée conjointement entre le service d'ophtalmologie du CHU de Brest, le Laboratoire de Traitement de l'Information Médicale et l'Ecole Nationale Supérieure des Télécommunications de Bretagne se compose de deux postes reliés par un réseau haut débit (2 Mbits symétrique). Le poste patient se compose d'un ordinateur de type PC équipé d'un écran tactile, d'une caméra (filmant le regard) et d'un jeu de lumières. Le poste orthoptiste dispose d'une caméra. L'interface graphique du poste orthoptiste permet la création et la sauvegarde d'exercices dédiés au patient, la gestion de la position de la caméra ainsi que la gestion des lumières du poste patient. Il permet aussi en cours de rééducation de visualiser les yeux du patient ainsi que l'examen en temps réel de l'exercice se déroulant sur l'écran du poste patien

Platform for low vision reeducation: elaboration and assessment

International audienc

Magma storage and degassing beneath the youngest volcanoes of the Massif Central (France): Lessons for the monitoring of a dormant volcanic province

Developing appropriate monitoring strategies in long-quiescent volcanic provinces is challenging due to the rarity of recordable geochemical and geophysical signals and the lack of experienced eruptive phenomenology in living memory. This is the case in the Massif Central (France) where the last eruptive sequence formed the Pavin’s Group of Volcanoes, about 7 ka ago. There, current evidence of a mantle activity reminiscence is suggested by the presence of mineral springwaters, mofettes, and soil degassing. It appears fundamental as a prerequisite to decipher the evolution of the gas phase in the magmatic system at the time of the eruptive activity to understand the meaning of current local gas emissions. In this study, we develop an innovative approach coupling CO2 densimetry and geochemistry of fluid inclusions from products erupted by the Pavin’s Group of Volcanoes. 3D imagery by Raman spectroscopy revealed that carbonate forming in fluid inclusions may lead to underestimation of CO2 density in fluid inclusions by up to 50 % and thus to unreliable barometric estimates. Fortunately, we found that this effect may be limited by focusing on fluid inclusions with a small diameter (<4 m) and where no solid phase is detected on Raman spectra. The time evolution of the eruptions of the Pavin’s Group of Volcanoes shows a progressive decrease of the pressure of magma storage (from more than 9 kbar down to 1.5-2 kbar) in parallel to magma differentiation (from basanites at Montcineyre to benmoreites at Pavin). The analysis of the noble gases entrapped in fluid inclusions yielded two main conclusions: (1) the helium isotope signature (Rc/Ra = 6.5-6.8) is in the range of values obtained in fluid inclusions from mantle xenoliths in the Massif Central (Rc/Ra = 5.6±1.1, on average) suggesting partial melting of the subcontinental lithospheric mantle, and (2) magma degassing (4He/40Ar* from 4.0 to 16.2) mirrors magma differentiation and the progressive rise of the magma ponding zones of the Pavin’s Group of Volcanoes. According to our modelling, about 80 % of the initial gas phase would be already exsolved from these magmas, even if stored at mantle depth. Based on the results obtained from fluid inclusions, we propose a model of the evolution of the signature of noble gases and carbon isotopes from mantle depth to crustal levels. In this frame, gas emissions currently emitted in the area (Rc/Ra = 6.1-6.7 and 4He/40Ar* = 1.7) point to an origin in the lithospheric mantle. This study strongly encourages the establishment of a regular sampling of local gas emissions to detect potential geochemical variations that may reflect a change from current steady-state conditions

Herpes Simplex Virus 1 Protein UL37 Interacts with Viral Glycoprotein gK and Membrane Protein UL20 and Functions in Cytoplasmic Virion Envelopment

We have shown that glycoprotein K (gK) and its interacting partner, the UL20 protein, play crucial roles in virion envelopment. Specifically, virions lacking either gK or UL20 fail to acquire an envelope, thus causing accumulation of capsids in the cytoplasm of infected cells. The herpes simplex virus 1 (HSV-1) UL37 protein has also been implicated in cytoplasmic virion envelopment. To further investigate the role of UL37 in virion envelopment, the recombinant virus DC480 was constructed by insertion of a 12-amino-acid protein C (protC) epitope tag within the UL37 amino acid sequence immediately after amino acid 480. The DC480 mutant virus expressed full-size UL37 as detected by the anti-protC antibody in Western immunoblots, accumulated unenveloped capsids in the cytoplasm of infected cells, and produced very small plaques on African green monkey kidney (Vero) cells that were similar in size to those produced by the UL20-null and UL37-null viruses. The DC480 virus replicated nearly 4 log less efficiently than the parental wild-type virus when grown on Vero cells. However, DC480 mutant virus titers increased nearly 20-fold when the virus was grown on FRT cells engineered to express the UL20 gene in comparison to the titers on Vero cells, while the UL37-null virus replicated approximately 20-fold less efficiently than the DC480 virus on FRT cells. Coimmunoprecipitation experiments and proximity ligation assays showed that gK and UL20 interact with the UL37 protein in infected cells. Collectively, these results indicate that UL37 interacts with the gK-UL20 protein complex to facilitate cytoplasmic virion envelopment. IMPORTANCE Herpes simplex viruses acquire their final envelopes by budding into cytoplasmic membranes derived from the trans-Golgi network (TGN). The tegument proteins UL36 and UL37 are known to be transported to the TGN sites of virus envelopment and to function in virion envelopment, since mutants lacking UL37 accumulate capsids in the cytoplasm that are unable to bud into TGN membranes. Viral glycoprotein K (gK) also functions in cytoplasmic envelopment, in a protein complex with the membrane-associated protein UL20 (UL20mp). This work shows for the first time that the UL37 protein functionally interacts with gK and UL20 to facilitate cytoplasmic virion envelopment. This work may lead to the design of specific drugs that can interrupt UL37 interactions with the gK-UL20 protein complex, providing new ways to combat herpesviral infections