Elemental microanalysis of fibroblasts by a scanning proton microprobe and application to Menkes’ disease

A scanning proton microprobe has been used for the elemental microanalysis of individual fibroblast cells. Both normal fibroblasts and fibroblasts cultured from patients with Menkes' disease, an X-linked genetic disorder known to be associated with defective copper metabolism, were examined by the probe. The cells were cultured on a thin ultra-clean nylon foil and retained on that surface for analysis. The focused high-energy proton beam was used to irradiate selected individual cells and elemental information was derived from X-ray and backscattered proton data. The sensitivity of the scanning proton microprobe to trace concentrations of heavy elements has allowed this elemental information to be used to identify individual cells as being either normal or a Menkes' mutant. The cell identification was based on the application of discriminate analysis to a data set formed from the ratios of copper to each of the macroelements present in the cell. This method of cell identification offers the promise of rapid diagnosis of Menkes' disease.     

Gravitropically-stimulated seedlings show autotropism in weightlessness

In a spaceflight experiment, autotropism by oat ( Avena sativa L.) coleoptiles following gravitropic responses was prominent in weightlessness: counter-reactions led to the straightening of the curved coleoptiles. This was not the case during clinorotation on earth. The autotropic reactions appeared to be related to the stimulus received during the stimulus period, i.e. the greater the response the greater the autotropic counter-reaction. Previous models of the gravitropic system which predicted that coleoptiles would not straighten in weightlessness are disproved. A modification to one of the models is proposed which includes the autotropic response observed in spaceflight. The nature of the counter-reactions in the absence of gravitropic stimulation is discussed.     

Direct assessment of symbiotically fixed nitrogen in the rhizosphere of alfalfa

Rhizodeposition has been proposed as one mechanism for the accumulation of significant amounts of N in soil during legume growth. The objective of this experiment was to directly quantify losses of symbiotically fixed N from living alfalfa (Medicago sativa L.) roots to the rhizosphere. We used ¹⁵N-labeled N₂ gas to tag recently fixed N in three alfalfa lines [cv. Saranac, Ineffective Saranac (an ineffectively nodulated line), and an unnamed line in early stages of selection for apparent N excretion] growing in 1-m long polyvinylchloride drainage lysimeters in loamy sand soil in a greenhouse. Plants were in the late vegetative to flowering growth stage during the 2-day labelling period. We determined the fate of this fixed N in various plant organs and soil after a short equilibration period (2 to 4 days) and after one regrowth period (35 to 37 days). Extrapolated N₂ fixation rates (46 to 77g plant^–1 h^–1) were similar to rates others have measured in the field. Although there was significant accretion of total N in rhizosphere compared to bulk soil, less than 1% was derived from newly fixed N and there were no differences between the excreting line and Saranac. Loss of N in percolate water was small. These results provide the first direct evidence that little net loss of symbiotically-fixed N occurs from living alfalfa roots into surrounding soil. In addition, these results confirm our earlier findings, which depended on indirect ¹⁵N labelling techniques. Net N accumulation in soil during alfalfa growth is likely due to other processes, such as decomposition of roots, nodules, and above ground litter, rather than to N excretion from living roots and nodules.     

Potato lectin: a modular protein sharing sequence similarities with the extensin family, the hevein lectin family, and snake venom disintegrins (platelet aggregation inhibitors)

Potato (Solanum tuberosum) lectin, is a chimeric chitin-binding protein comprised of a lectin domain fused to a hydroxyproline-rich glycoprotein domain. Here peptide sequence information from both domains is presented. A partial sequence of a major tryptic peptide T2: Leu-Pro-Ser-Hyp-Hyp-Hyp-Hyp-Hyp-Hyp-(His)-Hyp-Ser-Hyp-Hyp-Hyp-Hyp-Ser-Hyp-Hyp-Ser-Hyp-Hyp-Hyp-Hyp-Ser-Hyp-Hyp- was similar to the ‘P3’ type extensin major repetitive sequence: Ser-Hyp-Hyp-Hyp-Hyp-Ser-Hyp-Ser-Hyp-Hyp-Hyp-Hyp-suggesting common evolutionary origins for the extensins and the hydroxyproline-rich glycoprotein (HRGP) domain of potato lectin. Furthermore, alignment of three chymotryptic peptides from potato lectin, C1: Cys-Gly-Thr-Thr-Ser-Asp-Tyr, C2: Cys-Ser-Pro-Gly-Tyr, and C8: Thr-Gly-Glu-Cys-Cys-Ser-Ile with similar sequences from the hevein lectin family indicates that they have homologous chitin-binding domains, and hence have common evolutionary origins. Finally, all plant chitin-binding domains examined bore a remarkable sequence similarity, particularly in the spacing of Cys residues, to the disintegrins (platelet aggregation inhibitors) which occur in crotalid and viperid snake venoms. As such, sequence similarities not only identify potato lectin as a member of both the hevein and extensin families of plant proteins, but also suggest that an archetypal polypeptide module gave rise to both the plant chitin-binding domain and the reptile disintegrins.     

Is plasma membrane lipid composition defined in the exocytic or the endocytic pathway?

Compared with intracellular membranes, the plasma membrane is rich in cholesterol and sphingomyelin. How does this distinct composition arise? Here David Allan and Karl-Josef Kallen take a critical view of the belief that these lipids arrive at the plasma membrane via vesicular traffic from the Golgi complex and propose instead that they may be accreted in the endocytic recycling pathway.     

Effect of endotoxin on lipid order and motion in erythrocyte membranes

Electron paramagnetic resonance employing a lipid-specific spin label has been used to investigate the molecular effects of endotoxin on the physical state of bilayer lipids in rat erythrocyte membranes. When added at a concentration as low as 40 μg/ml to whole blood (plasma plus leukocytes present), decreased membrane lipid motion was found in subsequently washed and spin-labeled intact erythrocytes (P < 0.02). However, if endotoxin were added to washed, plasma plus leukocyte-free intact erythrocytes, no change in the motion of the spin label was found, suggesting that plasma-soluble substances and/or leukocytes are required to produce the change in the physical state of lipids. The decreased lipid motion found in these studies is discussed with reference to the known decreased deformability of endotoxin-treated red cells and to the pathogenesis of sepsis.     

Interactions between size and temperature influence fecundity and longevity of a tortricid moth,Zeiraphera canadensis

Females of Zeiraphera canadensis Mut. & Free., the spruce bud moth, were reared in the laboratory at constant and alternating temperatures, and in an outdoor insectary, to (1) determine the effects of temperature, age and size on several reproductive parameters and, (2) to test the hypothesis that body size-temperature interactions influence longevity and realized fecundity. Egg maturation was linearly related to age and large moths developed eggs at a higher rate than small ones. Mcan lifetime oviposition rate reached a maximum and remained stable at temperatures 20° C while the mean lifetime rate of egg maturation increased linearly with temperature, indicating that higher temperatures adversely affect oviposition. The production of nonviable eggs increased with age but also with temperature, suggesting high temperature (25° C) reduces egg quality and/or hinders fertilization. The realized fecundity and longevity of females reared under an alternating temperature regime (mean 20° C) was significantly less than that of females reared at constant 20° C. Similar realized fecundity, longevity and mean lifetime oviposition rates for females reared at temperatures alternating between 10 and 25° C (mean 20° C) and those at constant 25° C reflected the inability of females to recover from elevated diurnal temperatures. Longevity was positively related to female body size at constant 15 and 20° C but the relationships were negative for moths exposed to diurnal temperatures equal to or exceeding 25° C. Due to the reduced longevity of large moths at high temperatures, linear regressions between size and realized fecundity were only significant at constant temperatures 20° C. At higher temperatures, the size-fecundity relationship became curvilinear as a result of the diminished reproductive output of large individuals. Reduced fecundity and longevity of large females at high temperatures may have been due to elevated internal temperatures of large-bodied moths. Large females in a controlled-environment chamber maintained at 25° C developed an internal temperature excess (i.e. temperature above ambient) of nearly 2° C while small-bodied females exceeded ambient by only 0.3° C. However, when held at 20° C, the temperature excess of large-bodied moths was much less than 1° C and small-bodied females did not differ from ambient. Such interactions between temperature and body size suggest that there should be stabilizing selection toward moderate-sized individuals and may explain the absence of size-related effects on fecundity and longevity previously reported for several other lepidopterans.