Molecular genetic approaches to defining lipid function.

Lipids fulfill multiple and diverse functions in cells. Establishing the molecular basis for these functions has been challenging due to the lack of catalytic activity of lipids and the pleiotropic effects of mutations that affect lipid composition. By combining molecular genetic manipulation of membrane lipid composition with biochemical characterization of the resulting phenotypes, the molecular details of novel lipid functions have been established. This review summarizes the results of such a combined approach to defining lipid function in bacteria.

Molecular basis for class V beta-tubulin effects on microtubule assembly and paclitaxel resistance.

Vertebrates produce at least seven distinct beta-tubulin isotypes that coassemble into all cellular microtubules. The functional differences among these tubulin isoforms are largely unknown, but recent studies indicate that tubulin composition can affect microtubule properties and cellular microtubule-dependent behavior. One of the isotypes whose incorporation causes the largest change in microtubule assembly is beta5-tubulin. Overexpression of this isotype can almost completely destroy the microtubule network, yet it appears to be required in smaller amounts for normal mitotic progression. Moderate levels of overexpression can also confer paclitaxel resistance. Experiments using chimeric constructs and site-directed mutagenesis now indicate that the hypervariable C-terminal region of beta5 plays no role in these phenotypes. Instead, we demonstrate that two residues found in beta5 (Ser-239 and Ser-365) are each sufficient to inhibit microtubule assembly and confer paclitaxel resistance when introduced into beta1-tubulin; yet the single mutation of residue Ser-239 in beta5 eliminates its ability to confer these phenotypes. Despite the high degree of conservation among beta-tubulin isotypes, mutations affecting residue 365 demonstrate that amino acid substitutions can be context sensitive; i.e. an amino acid change in one isotype will not necessarily produce the same phenotype when introduced into a different isotype. Modeling studies indicate that residue Cys-239 of beta1-tubulin is close to a highly conserved Cys-354 residue suggesting the possibility that disulfide formation could play a significant role in the stability of microtubules formed with beta1- but not with beta5-tubulin.

Forest structure and composition of previously selectively logged and non-logged montane forests at Mt. Kilimanjaro

The montane forests of Mount Kilimanjaro in Tanzania have been subjected to a long history of selective logging. However, since 1984 logging of indigenous trees is prohibited. Today, these forests allow us to evaluate the long-term effects of selective logging. We mapped the height and diameter at breast height (DBH) of all trees >10 cm DBH on 10 sites of 0.25 ha. Five sites represent non-logged forests, another five selectively logged forests. We tested whether forests were still visibly affected 30–40 years after selective logging in terms of their forest structure and tree diversity. Additionally we compared tree densities of different species guilds, including disturbance-indicator species, late-successional species and main timber species. Furthermore, we specifically compared the community size distributions of selectively logged and non-logged forests, first across all species and then for the most important timber species, Ocotea usambarensis, alone. 30–40 years after selective logging forests still showed a higher overall stem density, mainly due to higher relative abundances of small trees (<50 cm DBH) in general, and higher densities of small size class stems of late-successional species specifically. For O. usambarensis, the selectively logged sites harboured higher relative abundances of small trees and lower relative abundances of harvestable trees. The higher relative abundance of small O. usambarensis-stems in selectively logged forests appears promising for future forest recovery. Thus, outside protected areas, selective logging may be a sustainable management option if logging cycles are considerably longer than 40 years, enough large source trees remain, and the recruiting O. usambarensis individuals find open space for their establishment.

Phospholipid oxidation generates potent anti-inflammatory lipid mediators that mimic structurally related pro-resolving eicosanoids by activating Nrf2.

Exposure of biological membranes to reactive oxygen species creates a complex mixture of distinct oxidized phospholipid (OxPL) species, which contribute to the development of chronic inflammatory diseases and metabolic disorders. While the ability of OxPL to modulate biological processes is increasingly recognized, the nature of the biologically active OxPL species and the molecular mechanisms underlying their signaling remain largely unknown. We have employed a combination of mass spectrometry, synthetic chemistry, and immunobiology approaches to characterize the OxPL generated from the abundant phospholipid 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (PAPC) and investigated their bioactivities and signaling pathways in vitro and in vivo. Our study defines epoxycyclopentenones as potent anti-inflammatory lipid mediators that mimic the signaling of endogenous, pro-resolving prostanoids by activating the transcription factor nuclear factor E2-related factor 2 (Nrf2). Using a library of OxPL variants, we identified a synthetic OxPL derivative, which alleviated endotoxin-induced lung injury and inhibited development of pro-inflammatory T helper (Th) 1 cells. These findings provide a molecular basis for the negative regulation of inflammation by lipid peroxidation products and propose a novel class of highly bioactive compounds for the treatment of inflammatory diseases.

Decreased phosphatidylcholine plasmalogens - A putative novel lipid signature in patients with stable coronary artery disease and acute myocardial infarction.

OBJECTIVE Glycerophospholipids and sphingolipids are structurally heterogeneous due to differences in the O- and N-linked fatty acids and head groups. Sphingolipids also show a heterogeneity in their sphingoid base composition which up to now has been little appreciated. The aim of this study was to investigate the association of certain glycerophospholipid and sphingolipid species with stable coronary artery disease (CAD) and acute myocardial infarction (AMI). METHODS The lipid profile in plasma from patients with stable CAD (n = 18) or AMI (n = 17) was compared to healthy subjects (n = 14). Sixty five glycerophospholipid and sphingolipid species were quantified by LC-MS. The relative distribution of these lipids into lipoprotein fractions was analyzed. RESULTS In the CAD cohort, 45 glycerophospholipid and sphingolipid species were significantly lower compared to healthy controls. In the AMI group, 42 glycerophospholipid and sphingolipid species were reduced. Four PC plasmalogens (PC33:1, PC33:2, PC33:3 and PC35:3) showed the most significant difference. Out of eleven analyzed sphingoid bases, four were lower in the CAD and six in the AMI group. Sphingosine-1-phosphate (S1P) levels were reduced in the AMI group whereas an atypical C16:1 S1P was lower in both groups. Phosphatidylcholine and sphingomyelin species were exclusively present in lipoprotein particles, whereas lysophosphatidylcholines were mainly found in the lipoprotein-free fraction. The observed differences were not explained by the use of statins as confirmed in a second, independent cohort. CONCLUSIONS Reduced levels of four PC plasmalogens (PC33:1, PC33:2, PC33:3 and PC35:3) were identified as a putatively novel lipid signature for CAD and AMI.

Different-sized grazers have distinctive effects on plant functional composition of an African savannah

Grazing ungulates play a key role in many ecosystems worldwide and can form diverse assemblages, such as in African savannahs. In many of these ecosystems, present-day ungulate communities are impoverished subsets of once diverse assemblages. While we know that excluding all ungulates from grasslands can exert major effects on both the structure and composition of the vegetation, how different individual ungulate species may have contrasting effects on grassland communities remains poorly understood. Here, we performed a long-term ‘Russian doll’ grazing exclosure experiment in an African savannah to test for the effects of different size classes of grazers on grassland structure and composition. At five sites, grazer species of decreasing size class (ranging from white rhino to scrub hare) were excluded using four fence types, to experimentally create different realized grazer assemblages. The vegetation structure and the grass functional community composition were characterized in 6 different years over a 10-year period. Additionally, animal footprints were counted to quantify the abundance of different ungulate species in each treatment. We found that while vegetation height was mostly driven by total grazing pressure of all species together, ungulate community composition best explained the functional community composition of grasses. In the short term, smaller ungulate species (‘mesoherbivores’) had strongest effects on vegetation composition, by shifting communities towards dominance by species with low specific leaf area and low nutritional value. In the long term, large grazers had stronger but similar effects on the functional composition of the system. Surprisingly, the largest ‘mega-herbivore’, the white rhinoceros, did not have strong effects on the vegetation structure or composition. Synthesis. Our results support the idea that different size classes of grazers have varying effects on the functional composition of grassland plant communities. Therefore, the worldwide decline in the diversity of ungulate species is expected to have (had) major impacts on community composition and functioning of grassland ecosystems, even if total grazing pressure has remained constant, for example, due to replacement by livestock.

Primary lowland dipterocarp forest at Danum Valley, Sabah, Malaysia. Species composition and patterns in the understorey

The spatial distributions of species of tree ≥10 cm gbh were examined in two 4 ha plots and related to the local variation in topography and soil chemistry. The plots were similar in their species composition, particularly in terms of the densities of small trees, and they showed very similar edaphic characteristics. Size class distributions varied little within and between plots. Ordination of 0.25 ha subplots highlighted parallel gradients in the vegetation of both plots when the densities of trees ≥10 cm gbh were considered. Focusing on understorey trees in the 10-<50 cm gbh class at the 0.04 ha subplot scale showed a similar vegetation gradient in both plots closely associated with change from lower slope to ridge. No relationship with soil chemistry was found. On the ridges a special group of understorey species formed clumps and these species contributed importantly to the ordinations. Borneo has a regional history of occasionally severe droughts. It is suggested here that the observed patterns in the understorey are due to differential responses to low soil water supply, the ridges probably tending to dryness more than the lower slopes. Within the large and diverse family Euphorbiaceae, which dominates the understorey at Danum, there may be ecophysiological groupings of species. The long-term effects of disturbance interacting with local edaphic factors on forest structure and composition are discussed.

Membrane Lipid Integrity Relies on a Threshold of ATP Production Rate in Potato Cell Cultures Submitted to Anoxia

In this paper we report on our study of the changes in biomass, lipid composition, and fermentation end products, as well as in the ATP level and synthesis rate in cultivated potato (Solanum tuberosum) cells submitted to anoxia stress. During the first phase of about 12 h, cells coped with the reduced energy supply brought about by fermentation and their membrane lipids remained intact. The second phase (12–24 h), during which the energy supply dropped down to 1% to 2% of its maximal theoretical normoxic value, was characterized by an extensive hydrolysis of membrane lipids to free fatty acids. This autolytic process was ascribed to the activation of a lipolytic acyl hydrolase. Cells were also treated under normoxia with inhibitors known to interfere with energy metabolism. Carbonyl-cyanide-4-trifluoromethoxyphenylhydrazone did not induce lipid hydrolysis, which was also the case when sodium azide or salicylhydroxamic acid were fed separately. However, the simultaneous use of sodium azide plus salicylhydroxamic acid or 2-deoxy-D-glucose plus iodoacetate with normoxic cells promoted a lipid hydrolysis pattern similar to that seen in anoxic cells. Therefore, a threshold exists in the rate of ATP synthesis (approximately 10 μmol g−1 fresh weight h−1), below which the integrity of the membranes in anoxic potato cells cannot be preserved.

Compilation of scientific results of the OASIS project

Seamounts are of great interest to science, industry and conservation because of their potential role as 'stirring rods' of the oceans, their enhanced productivity, their high local biodiversity, and the growing exploitation of their natural resources. This is accompanied by rising concern about the threats to seamount ecosystems, e.g. through over-fishing and the impact of trawling. OASIS described the functioning characteristics of seamount ecosystems. OASIS' integrated hydrographic, biogeochemical and biological information. Based on two case studies. The scientific results, condensed in conceptual and mass balanced ecosystem models, were applied to outline a model management plan as well as site-specific management plans for the seamounts investigated. OASIS addressed five main objectives: Objective 1: To identify and describe the physical forcing mechanisms effecting seamount systems Objective 2: To assess the origin, quality and dynamics of particulate organic material within the water column and surface sediment at seamounts. Objective 3: To describe aspects of the biodiversity and the ecology of seamount biota, to assess their dynamics and the maintenance of their production. Objective 4: Modelling the trophic ecology of seamount ecosystems. Objective 5: Application of scientific knowledge to practical conservation.

Structural group composition of low-polarity organic matter dissolved in waters of the Northwest Indian Ocean and the Red Sea

Data on amounts of various functional groups, i.e. aldehyde, acid, ester, alcohol, thiol and aromatic groups in several fractions of low-polarity dissolved organic matter are presented. An assumption that this organic matter is part of the lipid fraction is not confirmed. Amount of aromatic compounds in waters of the Northwest Indian Ocean is estimated to be about 1000 times higher than quantity of aromatic hydrocarbons discharged into the ocean each year in petroleum and petroleum products.