Plasma membrane H+-ATPase regulation is required for auxin gradient formation preceding phototropic growth.

Phototropism is a growth response allowing plants to align their photosynthetic organs toward incoming light and thereby to optimize photosynthetic activity. Formation of a lateral gradient of the phytohormone auxin is a key step to trigger asymmetric growth of the shoot leading to phototropic reorientation. To identify important regulators of auxin gradient formation, we developed an auxin flux model that enabled us to test in silico the impact of different morphological and biophysical parameters on gradient formation, including the contribution of the extracellular space (cell wall) or apoplast. Our model indicates that cell size, cell distributions, and apoplast thickness are all important factors affecting gradient formation. Among all tested variables, regulation of apoplastic pH was the most important to enable the formation of a lateral auxin gradient. To test this prediction, we interfered with the activity of plasma membrane H(+)-ATPases that are required to control apoplastic pH. Our results show that H(+)-ATPases are indeed important for the establishment of a lateral auxin gradient and phototropism. Moreover, we show that during phototropism, H(+)-ATPase activity is regulated by the phototropin photoreceptors, providing a mechanism by which light influences apoplastic pH.

Podocalyxin is a novel polysialylated neural adhesion protein with multiple roles in neural development and synapse formation

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.

Myosin motors and not actin comets are mediators of the actin-based Golgi-to-endoplasmic reticulum protein transport

We have previously reported that actin filaments are involved in protein transport from the Golgi complex to the endoplasmic reticulum. Herein, we examined whether myosin motors or actin comets mediate this transport. To address this issue we have used, on one hand, a combination of specific inhibitors such as 2,3-butanedione monoxime (BDM) and 1-[5-isoquinoline sulfonyl]-2-methyl piperazine (ML7), which inhibit myosin and the phosphorylation of myosin II by the myosin light chain kinase, respectively; and a mutant of the nonmuscle myosin II regulatory light chain, which cannot be phosphorylated (MRLC2AA). On the other hand, actin comet tails were induced by the overexpression of phosphatidylinositol phosphate 5-kinase. Cells treated with BDM/ML7 or those that express the MRLC2AA mutant revealed a significant reduction in the brefeldin A (BFA)-induced fusion of Golgi enzymes with the endoplasmic reticulum (ER). This delay was not caused by an alteration in the formation of the BFA-induced tubules from the Golgi complex. In addition, the Shiga toxin fragment B transport from the Golgi complex to the ER was also altered. This impairment in the retrograde protein transport was not due to depletion of intracellular calcium stores or to the activation of Rho kinase. Neither the reassembly of the Golgi complex after BFA removal nor VSV-G transport from ER to the Golgi was altered in cells treated with BDM/ML7 or expressing MRLC2AA. Finally, transport carriers containing Shiga toxin did not move into the cytosol at the tips of comet tails of polymerizing actin. Collectively, the results indicate that 1) myosin motors move to transport carriers from the Golgi complex to the ER along actin filaments; 2) nonmuscle myosin II mediates in this process; and 3) actin comets are not involved in retrograde transport.

Mutant Huntingtin impairs post-Golgi trafficking to lysosomes by delocalizing optineurin/Rab8 complex from the Golgi apparatus

Huntingtin regulates post-Golgi trafficking of secreted proteins. Here, we studied the mechanism by which mutant huntingtin impairs this process. Colocalization studies and Western blot analysis of isolated Golgi membranes showed a reduction of huntingtin in the Golgi apparatus of cells expressing mutant huntingtin. These findings correlated with a decrease in the levels of optineurin and Rab8 in the Golgi apparatus that can be reverted by overexpression of full-length wild-type huntingtin. In addition, immunoprecipitation studies showed reduced interaction between mutant huntingtin and optineurin/Rab8. Cells expressing mutant huntingtin produced both an accumulation of clathrin adaptor complex 1 at the Golgi and an increase of clathrin-coated vesicles in the vicinity of Golgi cisternae as revealed by electron microscopy. Furthermore, inverse fluorescence recovery after photobleaching analysis for lysosomal-associated membrane protein-1 and mannose-6-phosphate receptor showed that the optineurin/Rab8-dependent post-Golgi trafficking to lysosomes was impaired in cells expressing mutant huntingtin or reducing huntingtin levels by small interfering RNA. Accordingly, these cells showed a lower content of cathepsin D in lysosomes, which led to an overall reduction of lysosomal activity. Together, our results indicate that mutant huntingtin perturbs post-Golgi trafficking to lysosomal compartments by delocalizing the optineurin/Rab8 complex, which, in turn, affects the lysosomal function.

Formation of aluminosilicate-bearing quartz veins in the Simano nappe (Central Alps): structural, thermobarometric and oxygen isotope constraints

We combined structural analysis, thermobarometry and oxygen isotope geochemistry to constrain the evolution of kyanite and/or andalusite-bearing quartz veins from the amphibolite facies metapelites of the Simano nappe, in the Central Alps of Switzerland. The Simano nappe records a complex polyphase tectonic evolution associated with nappe stacking during Tertiary Alpine collision (D1). The second regional deformation phase (132) is responsible for the main penetrative schistosity and mineral lineation, and formed during top-to-the-north thrusting. During the next stage of deformation (D3) the aluminosilicate-bearing veins formed by crystallization in tension gashes, in tectonic shadows of boudins, as well as along shear bands associated with top-to-the-north shearing. D2 and D3 are coeval with the Early Miocene metamorphic peak, characterised by kyanite + staurolite + garnet + biotite assemblages in metapelites. The peak pressure (P) and temperature (T) conditions recorded are constrained by multiple-equilibrium thermobarometry at 630 +/- 20 degrees C and 8.5 +/- 1 kbar (similar to 27 km depth), which is in agreement with oxygen isotope thermometry indicating isotopic equilibration of quartz-kyanite pairs at 670 +/- 50 degrees C. Quartz-kyanite pairs from the aluminosilicate-bearing quartz veins yield equilibration temperatures of 645 +/- 20 degrees C, confirming that the veins formed under conditions near metamorphic peak. Quartz and kyanite from veins and the surrounding metapelites have comparable isotopic compositions. Local intergranular diffusion in the border of the veins controls the mass-transfer and the growth of the product assemblage, inducing local mobilization of SiO2 and Al2O3. Andalusite is absent from the host rocks, but it is common in quartz veins, where it often pseudomorphs kyanite. For andalusite to be stable at T-max, the pressure in the veins must have been substantially lower than lithostatic. An alternative explanation consistent with structural observations would be inheritance by andalusite of the kyanite isotopic signature during polymorphic transformation after the metamorphic peak.

Total lipid and fatty acid accumulation during basidiospore formation in the ectomycorrhizal fungus Pisolithus sp.

The basidiospores of Pisolithus sp. contain large amounts of lipids, indicating provision for future germination in the host rhizosphere. However, the accumulation, composition, and mobilization of lipids during formation and germination of these spores are largely unknown. In this study, lipid storage and fatty acid composition during basidiosporogenesis were analyzed in fresh basidiocarps using bright-field microscopy and gas chromatography. Abundant lipid bodies are found in the hyphae, basidia, and basidiospores of fungal basidiocarps. This evidences a considerable C transport in the basidiocarp to meet the C demand during basidiospore formation. Fatty acid composition analysis revealed the presence of 24 compounds with chains of 9 to 18 C atoms, either saturated or insaturated, with one or two insaturations. The fatty acid composition and content varied according to the developmental stage of the peridioles. In free basidiospores, the predominant compounds were 16:0, 16:1w5c, 18:1w9c, and 18:2w6,9c/18:0ante, at concentrations of 76, 46, 192, and 51 µg g-1 dry matter, respectively. Our results indicate that oleic acid is the major constituent of lipid reserves in Pisolithus sp. basidiospores. Further studies are being conducted to determine the factors that induce lipid mobilization during spore germination.

Ophiolite-related ultramafic rocks (Serpentinites) in the Caribbean region: a review of their occurrence, composition, origin, emplacements and Ni-Laterite soil formation

Ultramafic rocks, mainly serpentinized peridotites of mantle origin, are mostly associated with the ophiolites of Mesozoic age that occur in belts along three of the margins of the Caribbean plate. The most extensive exposures are in Cuba. The ultramafic-mafic association (ophiolites) were formed and emplaced in several different tectonic environments. Mineralogical studies of the ultramafic rocks and the chemistry of the associated mafic rocks indicate that most of the ultramafic-mafic associations in both the northern and southern margins of the plate were formed in arc-related environments. There is little mantle peridotite exposed in the ophiolitic associations of the west coast of Central America, in the south Caribbean in Curacao and in the Andean belts in Colombia. In these occurrences the chemistry and age of the mafic rocks indicates that this association is mainly part of the 89 Ma Caribbean plateau province. The age of the mantle peridotites and associated ophiolites is probably mainly late Jurassic or Early Cretaceous. Emplacement of the ophiolites possibly began in the Early Cretaceous in Hispaniola and Puerto Rico, but most emplacement took place in the Late Cretaceous to Eocene (e.g. Cuba). Along the northern South America plate margin, in the Caribbean mountain belt, emplacement was by major thrusting and probably was not completed until the Oligocene or even the early Miocene. Caribbean mantle peridotites, before serpentinization, were mainly harzburgites, but dunites and lherzolites are also present. In detail, the mineralogical and chemical composition varies even within one ultramafic body, reflecting melting processes and peridotite/melt interaction in the upper mantle. At least for the northern Caribbean, uplift (postemplacement tectonics) exposed the ultramafic massifs as a land surface to effective laterization in the beginning of the Miocene. Tectonic factors, determining the uplift, exposing the peridotites to weathering varied. In the northern Caribbean, in Guatemala, Jamaica, and Hispaniola, uplift occurred as a result of transpresional movement along pre-existing major faults. In Cuba, uplift occurred on a regional scale, determined by isostatic adjustment. In the south Caribbean, uplift of the Cordillera de la Costa and Serrania del Interior exposing the peridotites, also appears to be related to strike-slip movement along the El Pilar fault system. In the Caribbean, Ni-laterite deposits are currently being mined in the central Dominican Republic, eastern Cuba, northern Venezuela and northwest Colombia. Although apparently formed over ultramafic rocks of similar composition and under similar climatic conditions, the composition of the lateritic soils varies. Factors that probably determined these differences in laterite composition are geomorphology, topography, drainage and tectonics. According to the mineralogy of principal ore-bearing phases, Dominican Ni-laterite deposits are classified as the hydrous silicate-type. The main Ni-bearing minerals are hydrated Mg-Ni silicates (serpentine and ¿garnierite¿) occurring deeper in the profile (saprolite horizon). In contrast, in the deposits of eastern Cuba, the Ni and Cooccurs mainly in the limonite zone composed of Fe hydroxides and oxides as the dominant mineralogy in the upper part of the profile, and are classified as the oxide-type.