Growth patterns and food habits of Baetis rhodani, Capnia pygmaea and Diura nanseni in a West Norwegian river

As a part of weir-pond-ecosystem project, the growth patterns and food habits of Baetis rhodani (Ephemeroptera), Capnia pygmaea (Plecoptera), and Diura nanseni (Plecoptera) were investigated. B. rhodani was bivoltine. The winter generation consisted of three cohorts. C. pygmaea and D. nanseni accomplished their nymphal growth in one year. The predominate food consumed by B. rhodani and C. pygmaea was detritus. Variations in the diet of these detritivores are discussed in relation to microdistribution and possible competition. The most common prey items of the carnivorous D. nanseni were chironomids and nymphs of B. rhodani and C. pygmaea . Prey shortage is suggested to be the reason for small D. nanseni eating detritus.     

Physiological potential for survival of propagules of crassulacean Acid metabolism species

Terminal stem joints from three opuntias were detached and maintained for 160 days under natural climatic conditions in the winter and summer. Neither Crassulacean acid metabolism (CAM) nor CAM-idling, as evidenced by a diurnal malate flux, was maintained throughout the two periods; ceasing earlier in the summer period. A 13 to 20% fresh weight loss occurred over the winter period, as opposed to a 30 to 40% loss over the summer period, although tissue water potentials remained above −1.5 megapascals. Chlorophyll and protein contents remained essentially constant in the winter but decreased in the summer. Starch content decreased slightly over the winter but more significantly over the summer. Mucilage content increased slightly in winter and declined slightly in summer. The initiation of rooting was found to be inversely related to spine density and dependent upon orientation and season. Comparison of these data suggest rooting coincided with the cessation of CAM-idling in both climatic periods and was uncoupled from the occurrence of precipitation. The physiological limit for survival of these propagules after detachment was lower than anticipated being of only a few months' duration.     

Amyloid-β Protofibrils: Size,Morphology and Synaptotoxicity of an Engineered Mimic

Structural and biochemical studies of the aggregation of the amyloid-β peptide (Aβ) are important to understand the mechanisms of Alzheimer''s disease, but research is complicated by aggregate inhomogeneity and instability. We previously engineered a hairpin form of Aβ called Aβcc, which forms stable protofibrils that do not convert into amyloid fibrils. Here we provide a detailed characterization of Aβ₄₂ cc protofibrils. Like wild type Aβ they appear as smooth rod-like particles with a diameter of 3.1 (±0.2) nm and typical lengths in the range 60 to 220 nm when observed by atomic force microscopy. Non-perturbing analytical ultracentrifugation and nanoparticle tracking analyses are consistent with such rod-like protofibrils. Aβ₄₂ cc protofibrils bind the ANS dye indicating that they, like other toxic protein aggregates, expose hydrophobic surface. Assays with the OC/A11 pair of oligomer specific antibodies put Aβ₄₂ cc protofibrils into the same class of species as fibrillar oligomers of wild type Aβ. Aβ₄₂ cc protofibrils may be used to extract binding proteins in biological fluids and apolipoprotein E is readily detected as a binder in human serum. Finally, Aβ₄₂ cc protofibrils act to attenuate spontaneous synaptic activity in mouse hippocampal neurons. The experiments indicate considerable structural and chemical similarities between protofibrils formed by Aβ₄₂ cc and aggregates of wild type Aβ₄₂. We suggest that Aβ₄₂ cc protofibrils may be used in research and applications that require stable preparations of protofibrillar Aβ.     

Sequestration of the Aβ Peptide Prevents Toxicity and Promotes Degradation In Vivo

Protein aggregation, arising from the failure of the cell to regulate the synthesis or degradation of aggregation-prone proteins, underlies many neurodegenerative disorders. However, the balance between the synthesis, clearance, and assembly of misfolded proteins into neurotoxic aggregates remains poorly understood. Here we study the effects of modulating this balance for the amyloid-beta (Aβ) peptide by using a small engineered binding protein (Z_Aβ3) that binds with nanomolar affinity to Aβ, completely sequestering the aggregation-prone regions of the peptide and preventing its aggregation. Co-expression of Z_Aβ3 in the brains of Drosophila melanogaster expressing either Aβ₄₂ or the aggressive familial associated E22G variant of Aβ₄₂ abolishes their neurotoxic effects. Biochemical analysis indicates that monomer Aβ binding results in degradation of the peptide in vivo. Complementary biophysical studies emphasize the dynamic nature of Aβ aggregation and reveal that Z_Aβ3 not only inhibits the initial association of Aβ monomers into oligomers or fibrils, but also dissociates pre-formed oligomeric aggregates and, although very slowly, amyloid fibrils. Toxic effects of peptide aggregation in vivo can therefore be eliminated by sequestration of hydrophobic regions in monomeric peptides, even when these are extremely aggregation prone. Our studies also underline how a combination of in vivo and in vitro experiments provide mechanistic insight with regard to the relationship between protein aggregation and clearance and show that engineered binding proteins may provide powerful tools with which to address the physiological and pathological consequences of protein aggregation.     

Reproduction and recruitment of the seagrass Halophila stipulacea

The small seagrass species, Halophila stipulacea is abundant in the subtidal zone of the Bay of Eilat, Red Sea, southern Israel. Early life history characteristics of this species were investigated in summer 2002 by means of field surveys and outdoor experiments. Monospecific stands were found at depths of between 2 and 20 m. Reproduction began in late May and ripe pericarps were found for 1 month starting from the beginning of August. The ratios of female versus male plants were 0.9 at depths of between 2.5 and 10 m and 0.5 at depths of between 12.5 and 15 m. The proportion of reproductive branches was significantly larger in the shallow (2–5 m) than in the deep (7–15 m) populations, i.e., 20 ± 11% versus 6 ± 10%, respectively. Ripe seeds were predominantly produced at depths of between 2 and 5 m. Experimental studies demonstrated that full sunlight completely inhibited seedling growth at a depth of 30 cm; no macroscopic seedlings could be observed after 40-day exposure to full sunlight. If exposed to 90% photosynthetic active radiation (PAR) but protected from ultraviolet radiation (UVR), the number of macroscopic seedlings increased to 7.4 ± 2.3% of the planted seeds. If protected from both UVR and 80% of the PAR, the number of macroscopic seedlings increased to 22.5 ± 4.0% of the planted seeds. UVR exclusion and 80% PAR reduction also significantly increased the rhizome growth rates of seedlings in the first month after germination (0.14 ± 0.04 mm day⁻¹) compared with only UVR exclusion (0.04 ± 0.02 mm day⁻¹). The absence of H. stipulacea from the uppermost part of the subtidal zone (depths of 0–2 m) may be due to light inhibition of germling growth and uprooting by occasional storms.     

SEA domain autoproteolysis accelerated by conformational strain: mechanistic aspects

A subclass of SEA (sea urchin sperm protein, enterokinase, and agrin) domain proteins undergoes autoproteolysis between glycine and serine in a conserved G^− 1S^+ 1VVV motif to generate stable heterodimers. Autoproteolysis has been suggested to involve only the intramolecular catalytic action of the conserved serine hydroxyl in combination with conformational strain of the glycine-serine peptide bond. We conducted a number of experiments and simulations on the SEA domain from the MUC1 mucin to test this mechanism. Alanine-scanning mutagenesis of polar residues in the vicinity of the cleavage site demonstrates that only the nucleophile at position + 1 is required for efficient proteolysis. Molecular modeling shows that an uncleaved trans peptide is incompatible with the native heterodimeric structure, resulting in disruption of secondary structure elements and distortion of the scissile peptide bond. Insertion of glycine residues (to obtain G_nG^− 1S^+ 1VVV motifs) appears to relieve strain, and autoproteolysis is 100 times slower in a 1G (n = 1) mutant and not measurable in 2G and 4G mutants. Removal of the catalytic serine hydroxyl hampers cleavage considerably, but measurable autoproteolysis of this S1098A mutant still proceeds in the presence of strain alone. The uncleaved SEA precursor populates interconverting partially folded conformations, and autoproteolysis coincides with adoption of proper β-sheet secondary structure and completed folding. Molecular dynamics simulations of the precursor show that the serine hydroxyl and the preceding glycine carbonyl carbon can be in van der Waals contact at the same time as the scissile peptide bond becomes strained. These observations are all consistent with autoproteolysis accelerated by N → O acyl shift and conformational strain imposed upon protein folding in a reaction for which the free-energy barrier is decreased by substrate destabilization rather than by transition-state stabilization. The energetics of this coupled folding and autoproteolysis mechanism is accounted for in an accompanying article.     

SEA domain autoproteolysis accelerated by conformational strain: energetic aspects

A subclass of proteins with the SEA (sea urchin sperm protein, enterokinase, and agrin) domain fold exists as heterodimers generated by autoproteolytic cleavage within a characteristic G^− 1S^+ 1VVV sequence. Autoproteolysis occurs by a nucleophilic attack of the serine hydroxyl on the vicinal glycine carbonyl followed by an N → O acyl shift and hydrolysis of the resulting ester. The reaction has been suggested to be accelerated by the straining of the scissile peptide bond upon protein folding. In an accompanying article, we report the mechanism; in this article, we provide further key evidence and account for the energetics of coupled protein folding and autoproteolysis. Cleavage of the GPR116 domain and that of the MUC1 SEA domain occur with half-life (t_½) values of 12 and 18 min, respectively, with lowering of the free energy of the activation barrier by ∼ 10 kcal mol^− 1 compared with uncatalyzed hydrolysis. The free energies of unfolding of the GPR116 and MUC1 SEA domains were measured to ∼ 11 and ∼ 15 kcal mol^− 1, respectively, but ∼ 7 kcal mol^− 1 of conformational energy is partitioned as strain over the scissile peptide bond in the precursor to catalyze autoproteolysis by substrate destabilization. A straining energy of ∼ 7 kcal mol^− 1 was measured by using both a pre-equilibrium model to analyze stability and cleavage kinetics data obtained with the GPR116 SEA domain destabilized by core mutations or urea addition, as well as the difference in thermodynamic stabilities of the MUC1 SEA precursor mutant S1098A (with a G^− 1A^+ 1VVV motif) and the wild-type protein. The results imply that cleavage by N → O acyl shift alone would proceed with a t_½ of ∼ 2.3 years, which is too slow to be biochemically effective. A subsequent review of structural data on other self-cleaving proteins suggests that conformational strain of the scissile peptide bond may be a common mechanism of autoproteolysis.     

Interaction between isopod grazing and wave action: a structuring force in macroalgal communities in the southern Baltic Sea

The macroalgal belt in the southern Baltic Sea may be partly structured by the interaction of physical and biological factors. A field study, spanning the 1990s, describes a rapid decline of the Fucus spp. stands along the wave-exposed Swedish southeast coast. During this period, a relative dominance of Fucus vesiculosus L. shifted to a relative dominance of Fucus serratus L. The decline of F. vesiculosus coincided with observations of large numbers of the grazing isopods Idotea baltica (Pallas) and Idotea granulosa Rathke, or with field observations of frequent grazing marks on Fucus fronds. I. baltica, but not I. granulosa, tended to aggregate in the declining Fucus spp. stands, indicating a strong preference for Fucus spp. In a mesocosm experiment I. baltica, when given a choice, grazed both Fucus species at weak water motion. At strong water motion grazing was concentrated on F. vesiculosus. It is hypothesized that one of the reasons I. baltica preferred F. vesiculosus to F. serratus at strong water motion may have been differences in habitat quality, like width of thallus, influencing the ability to cling to the plant. Smaller thallus, as in F. vesiculosus, thus is the preferred habitat for grazing of I. blatica. We postulate that the existence of F. serratus in the area may be favoured by strong wave action and moderate but not strong grazing by I. baltica, relaxing the interspecific competition from F. vesiculosus.     

Riochemical and physiological effects in farmed Baltic salmon fed lipids containing xenobiotics extracted from Baltic herring

During a 2-year experimental period female baltic salmon (Salmo salar) were fed pellets impregnated with oil extracted from Baltic herring (Clupea harengus). This extract contained lipid-soluble xenobiotics present in Baltic herring, which constitute a major part of the natural diet of Baltic salmon. The fish were examined at the time of ovulation in November each year. After 2 years of feeding, the load of polychlorinated dibenzo-paradioxins and furans in the exposed group was about twice that in the control group, but still low compared with concentrations in feral Baltic salmon. In spite of the relatively low exposure level, several vital biochemical functions were disturbed in the treated fish. Organic skeletal variables were affected indicating that the bone metabolism had been altered. Furthermore, the activities of enzymes involved in steroid biosynthesis were affected, which could lead to disturbances in reproductive functions. Splenocytes from exposed fish sampled in November 1990 showed a reduced mitogenic response, indicating that their immune system was suppressed. Feeding the salmon with pollutant-impregnated pellets also resulted in an induction of the hepatic ethoxyresorufin-O-deethylase (EROD) activity after only 6 weeks of exposure. Likewise, morphological abnormalities, i.e. hypertrophic hepatocytes and various stages of hepatic degeneration, were already apparent after 6 weeks of exposure. However, no EROD induction or morphological responses were recorded at the second and third sampling event, i.e. after one and 2 years of exposure, respectively. this could indicate that some physiological functions may adapt to a restricted xenobiotic load.