Pollination ecology of Arum italicum (Araceae)

The pollination ecology of Arum italicum was studied in south-western France. This plant attracts olfactory dung-breeding flies through deceit. These insects are principally represented by Diptera, all belonging to saprophyte families. The volatilization of the odouriferous compounds, responsible for their attraction, is achieved through the production of heat by the appendix. The insects are trapped for 24 h in order to participate in both sexual phases of the protogynous inflorescence. The male flowers produce three heat events during flowering. These peaks of heat seem to be involved in the spathe movements, since they occur during the opening of the inflorescence and the liberation of the insects. The last male heat event may be linked with the liberation of pollen and its dispersion by stimulating trapped flies. According to their frequency and pollen-load, two Psychoda species appear to be the most efficient pollinators ( P. crassipenis and P. pusilla ). Nevertheless, each of the other attracted species could play a significant role under different spatio-temporal conditions. Experiments on self-pollination have shown that obligate cross-pollination is necessary for A. italicum to set seeds. Moreover, hand- and natural-pollinated plants showed similarly high abortion frequencies suggesting that seed set may be more constrained by resources rather than by pollination limitation.  © 2003 The Linnean Society of London, Botanical Journal of the Linnean Society , 2003, 141 , 205–214.     

Theory and practice of combining coagulant fixation and microwave histoprocessing

The German, F. Blum, introduced formalin as a fixative in 1893. Formalin rapidly became popular for hardening and preserving gross human and animal specimens. As a result, microscopy for diagnostic pathology by combining paraffin embedding and formalin fixation was developed. Alcohol-based fixatives have coagulation of proteins as their main preservative effect. Because there is no cross-linking, immunostaining is not compromised, and DNA and RNA is not damaged. Ethyl alcohol was used by Dutch scientists of the 18th century, but was replaced by the cheaper formalin. Addition of low molecular weight polyethylene glycol (PEG) optimized the coagulant fixative, Kryofix. The polyethylene glycol prevents excessive hardening and enhances the speed of coagulation of proteins. Kryofix was used on a large scale for skin biopsies in Leiden between 1987 and 2001. DNA preservation by the formulated coagulant fixative, BoonFix, is related to the concentration of ethyl alcohol, PEG and acetic acid. BoonFix has been used since 2004 in Leiden for over 40,000 diagnostic skin biopsies and more than 100,000 cervical samples. A literature review and three decades of experience with coagulant, formalin-free fixatives in pathology suggest that when health authorities realize that formalin invalidates expensive tests, it might eventually be eliminated legislatively from diagnostic pathology. Finally, coagulant fixation is optimal for microwave histoprocessing where ethyl alcohol is followed by isopropanol.     

Parallel changes in genetic diversity and species diversity following a natural disturbance

We examined the spatial and temporal variation of species diversity and genetic diversity in a metacommunity comprising 16 species of freshwater gastropods. We monitored species abundance at five localities of the Ain river floodplain in southeastern France, over a period of four years. Using 190 AFLP loci, we monitored the genetic diversity of Radix balthica , one of the most abundant gastropod species of the metacommunity, twice during that period. An exceptionally intense drought occurred during the last two years and differentially affected the study sites. This allowed us to test the effect of natural disturbances on changes in both genetic and species diversity. Overall, local (alpha) diversity declined as reflected by lower values of gene diversity H _S and evenness. In parallel, the among-sites (beta) diversity increased at both the genetic ( F _ST) and species ( F _STC) levels. These results suggest that disturbances can lead to similar changes in genetic and community structure through the combined effects of selective and neutral processes.     

Interactions of neural glycosaminoglycans and proteoglycans with protein ligands: assessment of selectivity, heterogeneity and the participation of core proteins in binding

The method of affinity coelectrophoresis was used to study the binding of nine representative glycosaminoglycan (GAG)-binding proteins, all thought to play roles in nervous system development, to GAGs and proteoglycans isolated from developing rat brain. Binding to heparin and non-neural heparan and chondroitin sulfates was also measured. All nine proteins-laminin-1, fibronectin, thrombospondin-1, NCAM, L1, protease nexin-1, urokinase plasminogen activator, thrombin, and fibroblast growth factor-2-bound brain heparan sulfate less strongly than heparin, but the degree of difference in affinity varied considerably. Protease nexin-1 bound brain heparan sulfate only 1.8- fold less tightly than heparin (Kdvalues of 35 vs. 20 nM, respectively), whereas NCAM and L1 bound heparin well (Kd approximately 140 nM) but failed to bind detectably to brain heparan sulfate (Kd>3 microM). Four proteins bound brain chondroitin sulfate, with affinities equal to or a few fold stronger than the same proteins displayed toward cartilage chondroitin sulfate. Overall, the highest affinities were observed with intact heparan sulfate proteoglycans: laminin-1's affinities for the proteoglycans cerebroglycan (glypican-2), glypican-1 and syndecan-3 were 300- to 1800-fold stronger than its affinity for brain heparan sulfate. In contrast, the affinities of fibroblast growth factor-2 for cerebroglycan and for brain heparan sulfate were similar. Interestingly, partial proteolysis of cerebroglycan resulted in a >400- fold loss of laminin affinity. These data support the views that (1) GAG-binding proteins can be differentially sensitive to variations in GAG structure, and (2) core proteins can have dramatic, ligand-specific influences on protein-proteoglycan interactions.      

Simultaneous stretching and contraction of stress fibers in vivo

To study the dynamics of stress fiber components in cultured fibroblasts, we expressed alpha-actinin and the myosin II regulatory myosin light chain (MLC) as fusion proteins with green fluorescent protein. Myosin activation was stimulated by treatment with calyculin A, a serine/threonine phosphatase inhibitor that elevates MLC phosphorylation, or with LPA, another agent that ultimately stimulates phosphorylation of MLC via a RhoA-mediated pathway. The resulting contraction caused stress fiber shortening and allowed observation of changes in the spacing of stress fiber components. We have observed that stress fibers, unlike muscle myofibrils, do not contract uniformly along their lengths. Although peripheral regions shortened, more central regions stretched. We detected higher levels of MLC and phosphorylated MLC in the peripheral region of stress fibers. Fluorescence recovery after photobleaching revealed more rapid exchange of myosin and alpha-actinin in the middle of stress fibers, compared with the periphery. Surprisingly, the widths of the myosin and alpha-actinin bands in stress fibers also varied in different regions. In the periphery, the banding patterns for both proteins were shorter, whereas in central regions, where stretching occurred, the bands were wider.     

金属框架结构材料MOF-199对漆酶的固定化及其性质

以金属框架结构材料MOF-199为载体对漆酶进行固定化,并对固定化酶的性质进行初步研究。首先,以3-氨基丙基三乙氧基硅烷对载体MOF-199进行表面氨基化修饰,再用戊二醛对载体进行活化,最后对漆酶进行固定化。固定化条件优化结果表明:在漆酶质量浓度0.3 g/L,戊二醛用量1%(体积分数),pH 4.8下固定7 h,制得固定化酶活性最高。对固定化酶的研究发现:最适反应温度为40℃,最适pH为5.2,在连续操作7次后,固定化酶的活力仍能保持在51%。固定化漆酶热稳定性,pH耐受性,贮存稳定性均明显高于游离漆酶。     

Evolving strategies for enzyme engineering

Directed evolution is a common technique to engineer enzymes for a diverse set of applications. Structural information and an understanding of how proteins respond to mutation and recombination are being used to develop improved directed evolution strategies by increasing the probability that mutant sequences have the desired properties. Strategies that target mutagenesis to particular regions of a protein or use recombination to introduce large sequence changes can complement full-gene random mutagenesis and pave the way to achieving ever more ambitious enzyme engineering goals.     

Structure and Function of Palladin's Actin Binding Domain

Here, we report the NMR structure of the actin-binding domain contained in the cell adhesion protein palladin. Previously, we demonstrated that one of the immunoglobulin domains of palladin (Ig3) is both necessary and sufficient for direct filamentous actin binding in vitro. In this study, we identify two basic patches on opposite faces of Ig3 that are critical for actin binding and cross-linking. Sedimentation equilibrium assays indicate that the Ig3 domain of palladin does not self-associate. These combined data are consistent with an actin cross-linking mechanism that involves concurrent attachment of two actin filaments by a single palladin molecule by an electrostatic mechanism. Palladin mutations that disrupt actin binding show altered cellular distributions and morphology of actin in cells, revealing a functional requirement for the interaction between palladin and actin in vivo.     

A new species of Goniothalamus (Annonaceae) from New Caledonia, representing a significant range extension for the genus

A previously unknown Annonaceae species from the South Pacific island of New Caledonia is described as Goniothalamus dumontetii . This is the first Goniothalamus species reported from the island, and the easternmost record for the genus. It is easily distinguished from its congeners by the shape of the monocarp (flattened elongate with lateral triangular projections), which reflects the shape of the seeds (flattened rhombohedral). The conservation status of the species is evaluated as endangered (EN) using World Conservation Union (IUCN) red list categories, as it is known from only one relatively small population. The interpretation of geological and molecular data suggests that Goniothalamus dispersed to New Caledonia relatively recently, and does not represent a relict of the break-up of Gondwana.  © 2007 The Linnean Society of London, Botanical Journal of the Linnean Society , 2007, 155 , 497–503.