Comparison of tissue-cultured bovine endothelial cells from aorta and saphenous vein

Endothelial cells were harvested from bovine aorta and saphenous vein with collagenase and cultured in McCoy's 5a medium (modified GIBCO) supplemented with 10% fetal bovine serum. The cells were subcultured through 17 passages over 4 to 5 months. The growth properties in culture of the two cell types were compared. Morphological comparisons included phase microscopy and scanning and transmission electron microscopy. Comparisons with cultured aortic smooth-muscle cells were made using phase and scanning electron microscopy. No differences were found between cultured endothelial cells from aorta and saphenous vein. Differences in growth patterns in culture clearly distinguished both endothelial cell types from smooth-muscle cells. The presence of Weibel-Palade bodies identified the cells from both sources as endothelial.     

Mapping of C2, Bf, and C4 genes to the swine major histocompatibility complex (swine leukocyte antigen)

Three miniature swine lines, inbred for swine leukocyte antigen (SLA) haplotypes, a, c, and d, and a recombinant line, haplotype g, were analyzed for possible restriction fragment length polymorphisms (RFLP) by Southern blot hybridization with human C2, factor B (Bf), and C4 specific probes. The search for RFLP by using a human C2 probe failed to reveal any variants. However, a Taq I polymorphism was identified with the human Bf probe and Bam HI and Pvu II polymorphisms were identified with the human C4 probe. Overlapping restriction fragments were found with the C2 and Bf probes, which strongly suggests close linkage of C2 and Bf genes in swine. Segregation analyses of the Bf and C4 polymorphisms indicated that the polymorphic fragments followed a Mendelian pattern of inheritance. The recombinant haplotype g, which expresses class I genes of haplotype c and class II genes of haplotype d, was shown to produce an identical RFLP pattern, by using the Bf and C4 probes, as haplotype d, but different from that of haplotype c. This indicates that there is a close association of [C4-Bf-C2] and class II genes in miniature swine. Although these data do not show conclusively the location of the [C4-Bf-C2] genes, it is hypothesized that swine [C4-Bf-C2] genes are located between the class II and class I genes, as has been demonstrated in mouse and man.     

Identification of the ATP·Mg-dependent protein phosphatase activator (Fa ) as a synapsin I kinase that inhibits cross-linking of synapsin I with brain microtubules

The ATP·Mg-dependent protein phosphatase activating factor (Fa) has been identified and purified to near homogeneity from brain. In this report, as evidenced on SDS-polyacrylamide gel electrophoresis followed by autoradiography, factorFa has further been identified as a cAMP and Ca²⁺-independent brain kinase that could phosphorylate synapsin I, a neuronal protein that coats synaptic vesicles, binds to cytoskeleton, and is believed to be involved in the modulation of neurotransmission. Kinetic study further indicated that factorFa could phosphorylate synapsin I with a lowK _m value of about 2 µM and with a molar ratio of 1 mol of phosphate per mole of protein. Peptide mapping analysis revealed that factorFa specifically phosphorylated the tail region of synapsin I but on a unique site distinct from those phosphorylated by Ca²⁺/calmodulin-dependent protein kinase II and cAMP-dependent protein kinase, the two well-established synapsin I kinases. Functional study further revealed that factorFa could phosphorylate this unique specific site on the tail region of synapsin I and thereby inhibit cross-linking of synapsin I with microtubules. The results further suggest the possible involvement of factorFa as a synapsin I kinase in the regulation of axonal transport process of synaptic vesicles via the promotion of vesicles motility during neurotransmission.     

Nodulation of lucerne (Medicago sativa L.) in an acid soil: Effects of inoculum size and lime-pelleting

The effects of inoculum level and lime-pelleting were studied in an acid soil with respect to the nodulation and growth of lucerne (Medicago sativa cv Resis) and the population dynamics of Rhizobium meliloti. In small root-boxes (rhizotrons), the in-situ survival of inoculated rhizobia was studied in the micro-environment around the seed for a period of 12 days after sowing. During the initial 24 hours, a strong increase in rhizobial numbers was measured, concomitantly with the development of roots. As a result of lime-pelleting, rhizobial numbers were higher only at 3 days after sowing (P<0.05). Later, this difference diminished steadily. Addition of lime did not increase the adhesion of the rhizobia to the seedling tap root. Plant responses to inoculation were studied in pots. To obtain optimal nodulation, the soil had to be neutralized around the seed with lime and at least 10⁵ cells of R. meliloti were required. With more than 10⁵ rhizobia per seed, lime-pelleting increased the number of crown-nodulated seedlings from 24% to 77%. Higher numbers of rhizobia could not compensate the effect of lime. A strong correlation was found between crown nodulation, nitrogen content and dry weight of the shoots.     

Genotype frequencies associated with incompatibility systems in tristylous plants

A general procedure has been given previously for calculating frequencies of the morphs Long, Mid, and Short in equilibrium populations of tristylous plants. It is now demonstrated that an equilibrium state actually exists at the genotype level if this procedure produces admissible morph frequencies. This result holds in a diploid model, with or without linkage between the two loci involved. It is shown how the genotype frequencies may be determined for any set of mating probabilities. It is also explained how these frequencies may be calculated in a tetraploid model incorporating double reduction. The general theory is applied to a particular situation where the Mid morph is at a selective disadvantage as a seed parent.     

Nonrandomness of point mutation as reflected in nucleotide substitutions in pseudogenes and its evolutionary implications

Summary We have obtained a revised estimate of the pattern of point mutation by considering more pseudogene sequences. Compared with our previous estimate, it agrees better with expectations based on the double-strand structure of DNA. The revised pattern, like the previous one, indicates that mutation occurs nonrandomly among the four nucleotides. In particular, the proportion of transitional mutations (59%) is almost twice as high as the value (33%) expected under random mutation. The same high proportion of transitions is observed in synonymous substitutions in genes. The proportion of transitional changes observed among electrophoretic variants of human hemoglobin is about the same as that predicted by the revised pattern of mutation. We also show that nonrandom mutation increases, by about 15%, the proportion of synonymous mutations due to single-nucleotide changes in the codon table, and increases, from 10% to 50%, the rate of synonymous mutation in the seven genes studied. However, nonrandom mutation reduces (by about 10%) the proportion of polar changes among nonsynonymous mutations in a gene. As far as single-nucleotide changes (in the codon table) are concerned, nonrandom mutation only slightly favors relatively conservative amino acid interchanges, and has virtually no effect on the proportions of radical changes and nonsense mutations.     

Non-photosynthetic effects of red and far-red light on root-nodule formation by leguminous plants

Summary Nodulation of pea and broad bean plants grown in the light was found to be reduced when the roots were exposed to far-red light for 5–15 minutes daily during 5 consecutive days following inoculation with nodule bacteria. Similar results were obtained following a single exposure to far-red light during a period of 15 minutes at the 3rd or 4th day after inoculation. When the roots were exposed to far-red light either before inoculation or during the first two days afterwards there were either no effects or only slight effects on nodulation The inhibitory effect of far-red light on nodulation was partly reduced by subsequent exposure to red light, provided that the same part of the plant was exposed to both red and far-red light,viz either the root or the shoot. When different parts of the plant were exposed to red and far-red light respectively, there was no interaction between the two kinds of light on nodulation. Plants whose roots were exposed to far-red light did not subsequently show stem elongation.Nodules were found to develop on the roots of pea plants grown in the dark, provided that the plants were kept at or below 22°C. At 25°C nodulation was almost absent. Nodulation was decreased by addition of kinetin and IAA. In contrast to plants grown in the light pea plants grown in the dark, inoculated with either an effective or ineffective strain of Rhizobium, developed equal numbers of nodules. Exposure to red light slightly increased the percentage of nodulated plants but decreased the number of nodules per plant. Exposure to far-red light slightly decreased both the percentage of nodulated plants and the number of nodules per plant. The effect of far-red light was counteracted by red light andvice versa.     

Rates of synonymous substitution in plant nuclear genes

Summary The rate of synonymous nucleotide substitution in nuclear genes of higher plants has been estimated. The rate varies among genes by a factor of up to two, in a manner that is not immediately explicable in terms of base composition or codon usage bias. The average rate, in both monocots and dicots, is about four times higher than that in chloroplast genes. This leads to an estimated absolute silent substitution rate of 6 × 10^–9 substitutions per site per year that falls within the range of average rates (2–8 × 10^–9) seen in different mammalian nuclear genomes.     

Quenching of Hoechst 33258 fluorescence in erythroid precursors.

Quenching of the fluorescence of DNA-bound Hoechst 33258 in erythroid precursors was studied by flow cytometry and cytochemistry. This quenching artifact may affect the measurement of ploidy in specific cases. The bone marrow cells of two patients with hemolytic disease and active erythropoiesis contained subpopulations of cells with an apparent hypodiploid DNA content as measured by flow cytometry of paraformaldehyde-fixed cells stained with Hoechst 33258. No aneuploidy was detected in either of the two cases when cells were stained with mithramycin or 7-aminoactinomycin D. Cells exhibiting reduced Hoechst 33258 fluorescence expressed glycophorin A and low amounts of CD36, and were therefore erythroid precursors. In one case studied, the number of cells with reduced Hoechst 33258 fluorescence and glycophorin A expressed agreed well with the number of cells containing nuclear hemoglobin. In the other case, hemoglobin was present in a significant proportion of nucleated cells. Calculated values for the efficiency of resonance energy transfer from Hoechst 33258 to hemoglobin were in accordance with the observed levels of quenching (approximately 10%). However, the results could also be explained by hemoglobin reabsorption of Hoechst 33258 fluorescence. Nuclei stained with Hoechst 33258 showed uniform fluorescence, probably due to extraction of hemoglobin during the isolation procedure.