Lateglacial and Holocene vegetation history in the mountain range of central Pyrenees

We present the pollen analysis of a new sedimentary sequence taken at La Pouretère ( 1720 m), in the mountain vegetation zone of the Marcadau valley (central Pyrenees). The Lateglacial and Holocene chronology is supported by six 14C-dating results. The complementary analysis of some vegetal macroremains, stomata, pollen-clusters and the use of pollen influx allows us to elucidate the dynamic of mountain species such as Pinus and specially Abies but also to infer the unusual part played by Betula at the beginning of the Postglacial period.     

Modulating HIV-1 replication by RNA interference directed against human transcription elongation factor SPT5

Background

Dendritic cell (DC) transmission of human immunodeficiency virus (HIV) to CD4+ T cells occurs across a point of cell-cell contact referred to as the infectious synapse. The relationship between the infectious synapse and the classically defined immunological synapse is not currently understood. We have recently demonstrated that human B cells expressing exogenous DC-SIGN, DC-specific intercellular adhesion molecule-3 (ICAM-3)-grabbing nonintegrin, efficiently transmit captured HIV type 1 (HIV-1) to CD4+ T cells. K562, another human cell line of hematopoietic origin that has been extensively used in functional analyses of DC-SIGN and related molecules, lacks the principal molecules involved in the formation of immunological synaptic junctions, namely major histocompatibility complex (MHC) class II molecules and leukocyte function-associated antigen-1 (LFA-1). We thus examined whether K562 erythroleukemic cells could recapitulate efficient DC-SIGN-mediated HIV-1 transmission (DMHT).

Results

Here we demonstrate that DMHT requires cell-cell contact. Despite similar expression of functional DC-SIGN, K562/DC-SIGN cells were inefficient in the transmission of HIV-1 to CD4+ T cells when compared with Raji/DC-SIGN cells. Expression of MHC class II molecules or LFA-1 on K562/DC-SIGN cells was insufficient to rescue HIV-1 transmission efficiency. Strikingly, we observed that co-culture of K562 cells with Raji/DC-SIGN cells impaired DMHT to CD4+ T cells. The K562 cell inhibition of transmission was not directly exerted on the CD4+ T cell targets and required contact between K562 and Raji/DC-SIGN cells.

Conclusions

DMHT is cell type dependent and requires cell-cell contact. We also find that the cellular milieu can negatively regulate DC-SIGN transmission of HIV-1 in trans.     

Temporal and spatial genetic structure in Vitellaria paradoxa (shea tree) in an agroforestry system in southern Mali

Ten microsatellite loci were used to investigate the impact of human activity on the spatial and temporal genetic structure of Vitellaria paradoxa (Sapotaceae), a parkland tree species in agroforestry systems in southern Mali. Two stands (forest and fallow) and three cohorts (adults, juveniles and natural regeneration) in each stand were studied to: (i) compare their levels of genetic diversity (gene diversity, HE; allelic richness, Rs; and inbreeding, FIS); (ii) assess their genetic differentiation (FST); and (iii) compare their levels of spatial genetic structuring. Gene diversity parameters did not vary substantially among stands or cohorts, and tests for bottleneck events were nonsignificant. The inbreeding coefficients were not significantly different from zero in most cases (FIS = -0.025 in forest and 0.045 in fallow), suggesting that the species is probably outbreeding. There was a weak decrease in F(IS) with age, suggesting inbreeding depression. Differentiation of stands within each cohort was weak (FST = 0.026, 0.0005, 0.010 for adults, juveniles and regeneration, respectively), suggesting extensive gene flow. Cohorts within each stand were little differentiated (FST = -0.001 and 0.001 in forest and fallow, respectively). The spatial genetic structure was more pronounced in fallow than in forest where adults showed no spatial structuring. In conclusion, despite the huge influence of human activity on the life cycle of Vitellaria paradoxa growing in parkland systems, the impact on the pattern of genetic variation at microsatellite loci appears rather limited, possibly due to the buffering effect of extensive gene flow between unmanaged and managed populations.     

Effect of changes in pH on wall tension in isolated rat pulmonary artery: role of the RhoA/Rho-kinase pathway

Pulmonary arteries (PA) are resistant to the vasodilator effects of extracellular acidosis in systemic vessels; the mechanism underlying this difference between systemic and pulmonary circulations has not been elucidated. We hypothesized that RhoA/Rho-kinase-mediated Ca2+ sensitization pathway played a greater role in tension development in pulmonary than in systemic vascular smooth muscle and that this pathway was insensitive to acidosis. In arterial rings contracted with the alpha1-agonist phenylephrine (PE), the Rho-kinase inhibitor Y-27632 (< or =3 microM) induced greater relaxation in precontracted PA rings than in aortic rings. In PA rings stimulated by PE, the activation of RhoA was greater than in aorta. Normocapnic acidosis (NA) induced a smaller relaxation in precontracted PA than in aorta. However, in the presence of nifedipine and thapsigargin, when PE-induced contraction was predominantly mediated by Rho-kinase, the relaxant effect of NA was reduced and similar in both vessel types. Furthermore, in the presence of Y-27632, NA induced a greater relaxation in both PA and aorta, which was similar in both vessels. Finally, in alpha-toxin-permeabilized smooth muscle, PE-induced contraction at constant Ca2+ activity was inhibited by Y-27632 and unaffected by acidosis. These results indicate that Ca2+ sensitization induced by the RhoA/Rho-kinase pathway played a greater role in agonist-induced vascular smooth muscle contraction in PA than in aorta and that tension mediated by this pathway was insensitive to acidosis. The predominant role of the RhoA/Rho-kinase pathway in the pulmonary vasculature may account for the resistance of this circulation to the vasodilator effect of acidosis observed in the systemic circulation.     

Antagonistic roles of ESCRT and Vps class C/HOPS complexes in the recycling of yeast membrane proteins

In Saccharomyces cerevisiae, deficiencies in the ESCRT machinery trigger the mistargeting of endocytic and biosynthetic ubiquitinated cargoes to the limiting membrane of the vacuole. Surprisingly, impairment of this machinery also leads to the accumulation of various receptors and transporters at the plasma membrane in both yeast and higher eukaryotes. Using the well-characterized yeast endocytic cargo uracil permease (Fur4p), we show here that the apparent stabilization of the permease at the plasma membrane in ESCRT mutants results from an efficient recycling of the protein. Whereas several proteins as well as internalized dyes are known to be recycled in yeast, little is known about the machinery and molecular mechanisms involved. The SNARE protein Snc1p is the only cargo for which the recycling pathway is well characterized. Unlike Snc1p, endocytosed Fur4p did not pass through the Golgi apparatus en route to the plasma membrane. Although ubiquitination of Fur4p is required for its internalization, deubiquitination is not required for its recycling. In an attempt to identify actors in this new recycling pathway, we found an unexpected phenotype associated with loss of function of the Vps class C complex: cells defective for this complex are impaired for recycling of Fur4p, Snc1p, and the lipophilic dye FM4-64. Genetic analyses indicated that these phenotypes were due to the functioning of the Vps class C complex in trafficking both to and from the late endosomal compartment.     

Silencing of Toll pathway components by direct injection of double-stranded RNA into Drosophila adult flies

Double-stranded RNA (dsRNA) gene interference is an efficient method to silence gene expression in a sequence-specific manner. Here we show that the direct injection of dsRNA can be used in adult Drosophila flies to disrupt function of endogenous genes in vivo. As a proof of principle, we have used this method to silence components of a major signaling cascade, the Toll pathway, which controls fruit fly resistance to fungal and Gram-positive bacterial infections. We demonstrate that the knockout is efficient only if dsRNA is injected in 4- or more day-old flies and that it lasts for at least 1 week. Furthermore, we report dsRNA-based epistatic gene analysis via injection of a mixture of two dsRNAs and propose that injection of dsRNA represents a powerful method for rapid functional analysis of genes in Drosophila melanogaster adults, particularly of those whose mutations are lethal during development.     

Enzymatic Browning and Biochemical Alterations in Black Spots of Pineapple [Ananas comosus (L.) Merr.]

Penicillium funiculosum Thom. was consistently isolated from pineapple-infected fruitlet (black spots). Polyphenol oxidase, peroxidase, and laccase activities were determined in extracts from contiguous and infected fruitlets. Healthy fruitlets showed a rather high level of polyphenol oxidase (optimum pH 7.0), and this activity was tremendously increased (×10) in contiguous infected fruitlets. Furthermore, infected fruitlets also exhibited laccase activity (optimum pH 4.0), while peroxidase was rather constant in both fruitlets. Browning reactions were attributed to qualitative and quantitative modifications of the enzymatic equipment (polyphenol oxidase and laccase) (p<0.0001). In infected fruiltets, sucrose and L-malic acid were present at significantly lower amounts than in healthy ones, likely owing to fungal metabolism (p<0.0001), whereas cell wall material was three times higher, which could be viewed as a defense mechanism to limit expansion of the mycelium. RID= ID= <E5>Correspondence to: </E5>S. Avallone; <E5>email:</E5> avallone&commat;siarc.cnearc.fr Received: 3 September 2002 / Accepted: 25 September 2002     

Enzymatic browning and biochemical alterations in black spots of pineapple [Ananas comosus (L.) Merr.

Penicillium funiculosum Thom. was consistently isolated from pineapple-infected fruitlet (black spots). Polyphenol oxidase, peroxidase, and laccase activities were determined in extracts from contiguous and infected fruitlets. Healthy fruitlets showed a rather high level of polyphenol oxidase (optimum pH 7.0), and this activity was tremendously increased (X 10) in contiguous infected fruitlets. Furthermore, infected fruitlets also exhibited laccase activity (optimum pH 4.0), while peroxidase was rather constant in both fruitlets. Browning reactions were attributed to qualitative and quantitative modifications of the enzymatic equipment (polyphenol oxidase and laccase) (p < 0.0001). In infected fruiltets, sucrose and L-malic acid were present at significantly lower amounts than in healthy ones, likely owing to fungal metabolism (p < 0.0001), whereas cell wall material was three times higher, which could be viewed as a defense mechanism to limit expansion of the mycelium.     

Changes of action potentials and force at lowered [Na+]o in mouse skeletal muscle: implications for fatigue

We examined 1) whether the effects of lowered trans-sarcolemmal Na⁺ gradient on force differed between nonfatigued fast- and slow-twitch muscles of mice and 2) whether effects on action potentials could explain the decrease of force. The Na⁺ gradient was reduced by lowering the extracellular [Na⁺] ([Na⁺]_o). The peak force-[Na⁺]_o relationships for the twitch and tetanus were the same in nonfatigued extensor digitorum longus and soleus muscles: force was maintained over a large range of [Na⁺]_o and then decreased abruptly over a much smaller range. However, fatigue was significantly exacerbated at a lowered [Na⁺]_o that had little effect in nonfatigued soleus muscle. This finding suggests that substantial differences exist in the Na⁺ effect on force between nonfatigued and fatigued muscle. The reduced contractility in nonfatigued muscles at lowered [Na⁺]_o was largely due to 1) an increased number of inexcitable fibers and threshold for action potentials, 2) a reduction of action potential amplitude, and 3) a reduced capacity to generate action potentials throughout trains. sodium gradient; muscle contraction; action potential train; extensor digitorum longus; soleus