Identification, purification, and characterization of truncated forms of the human nerve growth factor receptor

We report the presence of truncated forms of the nerve growth factor receptor (NGFRt) in the conditioned medium of the human melanoma cell line A875 and in human urine and amniotic fluid. Radioiodinated nerve growth factor (125I-NGF) specifically bound to NGFRt was chemically cross-linked. After immunoprecipitation, labeled receptor species were visualized by autoradiography following sodium dodecyl sulfate-polyacrylamide gel electrophoresis. NGFRts were purified from human adult male urine or a mixture of human amniotic fluid and infant urine by using a combination of either ion exchange chromatography (adult) or ammonium sulfate precipitation (infant) and immunoaffinity chromatography. Typical yields were about 1 microgram/liter of adult urine and 75 micrograms/liter of amniotic fluid/infant urine. The purified proteins, with molecular masses of 45, 40, and 35 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (12%), were confirmed to be NGFRts by amino-terminal sequencing and were designated NGFRt-1, NGFRt-2, and NGFRt-3, respectively. The isoelectric points of these three species ranged from 3.3 to 3.95 and displayed intraspecies heterogeneity; subsequently, amino acid residues covalently modified with sialic acid-containing carbohydrates were documented. The binding affinities of these species for nerve growth factor were comparable to that of the low affinity cell surface receptor. The potential to isolate milligram quantities of human NGFRts allows for model studies of the physicochemical structure of the intact receptor and the generation of polyclonal antibodies to study the biological functions of the NGF receptor.     

Enriched populations of rat retinal ganglion cells: Studies using a cell-type specific surface marker

Cells dissociated from adult and neonatal rat retinas were separated by density gradient centrifugation. Previous work had shown that rat retinal cells labelled by an immunofluorescence assay for the Thy-1 antigen were chiefly or exclusively ganglion cells, and so the proportion of Thy-1 positive cells in the density gradient fractions was used as an index of the enrichment of ganglion cells. The proportion of Thy-1 positive neonatal cells was increased from about 0.4% in the initial dissociate to about 8% in the most enriched fraction of a Percoll step gradient. Amongst adult cells the initial 0.7% Thy-1 positive cells were increased to roughly 2% in the best fraction of a metrizamide step gradient.

The presence of relatively large numbers of Thy-1 positive cells in other fractions suggested that it would be difficult to further increase the proportion of rat ganglion cells by methods based on their sedimentation properties. These results demonstrate the importance of cell-type specific markers in attempts to purify cells from the central nervous system.     

Comparison of the metabolism of benzo[alpha]pyrene and binding to DNA caused by rat liver nuclei and microsomes.

Administration of 3-methylcholanthrene (3MC) to rats greatly enhanced the aryl hydrocarbon hydroxylase (AHH) activity of liver nuclei. However, the binding in vitro [3H]benzo[alpha]pyrene (BP) to DNA within the nuclei which occurred at the same time as hydroxylation of BP was much less enhanced. Thin layer chromatography of the metabolites of BP produced by these nuclei revealed the same metabolites in similar relative amounts as were produced by rat liver microsomes prepared from rats which had received 3MC. The binding to DNA was further analysed by hydrolysis of the DNA and fractionation on a Sephadex column. This analysis revealed that the binding to DAN in nuclei was very similar in nature to that which occurred when calf-thymus DNA was added to microsomes metabolising BP.     

Ethylene in plant growth and development, storage, soil microbiology and plant pathology

In field experiments on a sandy loam at Wellesbourne, England in 1972 and on a silt loam at Agassiz, British Columbia in 1973, combinations of herbicides and insecticides were applied at sowing to determine their effects on weeds and invertebrate populations and on the growth and yield of cauliflowers grown at high density There was good agreement between the results from the two locations. The two combinations of herbicides, 0.6 kg trifluralin/ha incorporated pre-drilling plus 2.2 kg propachlor/ha pre-emergence and 2.2 kg nitrofen/ha plus 2.2 kg propachlor/ha both applied pre-emergence, gave good weed control, their relative effectiveness depending on the species composition of the weed population. The insecticides isophenphos, carbofuran, chlorfenvinphos and fensulfothion were applied as bow-wave treatments. None of them, whether in combination with herbicides or not, adversely affected crop stand or yield. Yield was reduced when either weeds or root-fly maggots (Hylemya brassicae (Bouché)) were not controlled. Only in one experiment was there any evidence of any herbicide-insecticide interactions. Where trifluralin and carbofuran were applied together at Agassiz, the control of both weeds and maggots was less than that with the other combinations. None of the treatments affected the populations of predatory beetles, but the numbers of earthworms were greatly reduced by carbofuran and to a lesser extent by chlorfenvinphos. Except for carbofuran in one experiment, the treatments had no effects on the numbers of aphids, lepidopterous larvae or leaf miners present at harvest.     

Influence of Meloidogyne incognita on the Content of Amino Acids and Nicotine in Tobacco Grown Under Gnotobiotic Conditions

Seedlings of Meloidogyne incognita-resistant (N.C. 95) and -susceptible (McNair 30) tobacco cultivars were grown aseptically for 55 days inside isolator chambers in autoclaved soil infested with 0 or 3,000 axenized eggs of M. im ognita per 500 cc of soil. Healthy and infected plants were compared. Dry root weights of infected plants of resistant and susceptible cultivars were 16% and 84%, respectively, less than the controls. Sixteen amino acids, including those precursors for nicotine, and nicotine, increased significantly in infected roots of both cultivars. Increases in amino acids in infected roots ranged from 28% for valine to 103% for tyrosine in the resistant N.C. 95, and from 30% for leucine to 148% for tyrosine in lhe susceptible McNair 30. Nicotine content (dry weight basis) increased 42% and 62% in infected roots of resistant and susceptible cultivars, respectively. Nematode infection increased nicotine by 112% in leaves of N.C. 95, and decreased it by 56% in leaves of McNair 30. Root damage by M. incognita probably decreased nicotine movement into leaves of McNair 30. In N.C. 95, nicotine movement into leaves apparently was not adversel b affected due to lack of significant root damage.