The roles of calcium and manganese ions in the in vitro conversion of 1-aminocyclopropane-1-carboxylic acid to ethylene by lentil root membranes

Ca²⁺ and Mn²⁺ activate the conversion of 1-aminocyclopropane-1-carboxylic acid (ACC) by root microsomes of Vicia lens as they do in other similar systems. The preparation of microsomes in the presence of Mn²⁺ greatly increases their ability to convert ACC into ethylene, without addition of Mn²⁺ in the reaction mixture. Ca²⁺ does not have this property. The effect could not be attributed to Mn²⁺ entrapping into membrane vesicles (sonication followed by repelleting had no effect) but, possibly, in part to Mn²⁺-mediated binding to microsomes of a soluble factor favouring the conversion of ACC to C₂H₄. Although no direct correlation could be established in vitro between ethylene-forming-enzyme (EFE) and peroxidase activities, some soluble peroxidases might be this soluble factor. Mn²⁺ favoured attachment to membranes of some peroxidase activity from the soluble fraction and from commercial HRP and lipoxygenase. This binding effect of Mn₂₊ cannot be readily distinguished from its role in the generation of a chain of free radicals and in redox mechanisms.     

Serotonin Levels in the Rabbit Retina Are Elevated Following Intraocular Injection of Forskolin

Analysis of the mammalian retina for serotonin immunoreactivity suggests an absence of the amine. However, following an intraocular injection of forskolin (1 microM) into a rabbit eye 1 h before analysis of the retina, serotonin immunoreactivity is associated with a subpopulation of amacrine cells. These cells correspond in size and position to the "indoleamine-accumulating cells" of the retina. Biochemical experiments show that forskolin treatment produces an increase in levels of endogenous serotonin and 5-hydroxytryptophan but has no effect on the uptake of serotonin or tryptophan or the metabolism of 5-hydroxytryptophan. These results suggest that the "indoleamine-accumulating cells" in the retina are "serotonergic cells" and that the level of amine is elevated sufficiently for localisation following forskolin treatment. It would appear that forskolin either directly or indirectly activates tryptophan hydroxylase.     

Heterogeneity of the glutathione transferase genes encoding enzymes responsible for insecticide degradation in the housefly

One of the four glutathione-S-transferases (GST) that is overproduced in the insecticide-resistant Cornell-R strain of the housefly (Musca domestica) produces an activity that degrades the insecticide dimethyl parathion and conjugates glutathione to lindane. In earlier work, it was shown that the resistant Cornell-R carries an amplification, probably a duplication, of one or more of its GST loci and that this amplification is directly related to resistance. Using polymerase chain reaction (PCR) amplification with genomic DNA, multiple copies of the gene encoding the parathion-degrading activity (called MdGst-3) were subcloned from both the ancestral, insecticide-susceptible strain BPM and from the insecticide-resistant Cornell-R. In BPM, three different MdGst-3 genes were identified while in Cornell-R, 12 different MdGst-3 sequences were found that, though closely related to ancestral genes, had diverged by a few nucleotides. This diversity in MdGst-3 genomic sequences in Cornell-R is reflected in the expressed sequences, as sampled through a cDNA bank. Population heterozygosity cannot account for these multiple GST genes. We suggest that selection for resistance to insecticides has resulted in not only amplification of the MdGst-3 genes but also in the divergence of sequence between the amplified copies. Received: 22 November 1995 / Accepted: 23 February 1996     

Mutation-Selection Balance at a Modifier-of-Imprinting Locus

We propose a pair of population genetic models for a modifier-of-imprinting locus for which different genotypes imprint different proportions of an imprintable target locus in their gametes. The two models examine the situations in which imprinting is advantageous or disadvantageous, and we discuss three cases for which the modifier is respectively partially dominant, dominant, or recessive. The models predict the stable equilibrium frequencies of the mutant modifier and functionally diploid individuals in a large population in terms of up to four parameters: the mutation rate at the modifier locus, V; the selection coefficient against the disadvantageous phenotype, s; the proportion of unimprinted eggs produced by homozygotes for the mutant modifier, θ, and, in the partially dominant models, the dominance parameter, k. The equilibrium frequency of the mutant phenotypes is shown to be approximately twice that of standard Mendelian models: 2V/s or 4V/s when the modifier is recessive or dominant, respectively. Mathematical equivalences between these and nonimprinting models are noted.     

The scl gene product is required for the generation of all hematopoietic lineages in the adult mouse.

Homozygosity for a null mutation in the scl gene causes mid-gestational embryonic lethality in the mouse due to failure of development of primitive hematopoiesis. Whilst this observation established the role of the scl gene product in primitive hematopoiesis, the death of the scl null embryos precluded analysis of the role of scl in later hematopoietic development. To address this question, we created embryonic stem cell lines with a homozygous null mutation of the scl gene (scl-/-) and used these lines to derive chimeric mice. Analysis of the chimeric mice demonstrates that the scl-/- embryonic stem cells make a substantial contribution to all non-hematopoietic tissues but do not contribute to any hematopoietic lineage. These observations reveal a crucial role for the scl gene product in definitive hematopoiesis. In addition, in vitro differentiation assays with scl-/- embryonic stem cells showed that the scl gene product was also required for formation of hematopoietic cells in this system.     

The development of phloem anastomoses between vascular bundles and their role in xylem regeneration after wounding in Cucurbita and Dahlia

The differentiation of phloem anastomoses linking the longitudinal vascular bundles has been studied in stem internodes of Cucurbita maxima Duchesne, C. pepo L. and Dahlia pinnata Cav. These anastomoses comprise naturally occurring regenerative sieve tubes which redifferentiate from interfascicular parenchyma cells in the young internodes. In all three species, severing a vascular bundle in a young internode resulted in regeneration of xylem to form a curved by-pass immediately around the wound. The numerous phloem anastomoses in these young internodes were not involved in this process, the regenerated vessels originating from interfascicular parenchyma alone. Conversely, in mature internodes of Dahlia, the regenerated vessels originated from initials of the interfascicular cambia, and their phloem anastomoses did not influence the pattern of xylogenesis. On the other hand, in old internodes of Cucurbita, in which an interfascicular cambium was not yet developed, the parenchyma cells between the bundles had lost the ability to redifferentiate into vessel elements, and instead, regenerated vessels were produced in the phloem anastomoses. Thus, the wounded region of the vascular bundle was not bypassed via the shortest, curved pathway, but by more circuitous routes further away from the wound. Some of the regenerated vessels produced in the phloem anastomoses were extremely wide, and presumably efficient conductors of water. It is proposed that the dense network of phloem anastomoses developed during evolution as a mechanism of adaptation to possible damage in mature internodes by providing flexible alternative pathways for efficient xylem regeneration in plants with limited or no interfascicular cambium.This paper is dedicated to the memory of the late Isaac Blachmann (deceased 19 November 1995), father-in-law of the senior author, for encouragement and advice throughout the yearsThis research was supported by an International Scientific Exchange Award to R.A. from the Israel Academy of Sciences and The Royal Society.     

Gender specialization and distyly in hoary puccoon,LITHOSPERMUM CROCEUM (Boraginaceae)

This 2-yr study of gender and its relation to the sizes of pin and thrum plants in Lithospermum croceum was performed in central Michigan. The population was composed of 51-52% thrums, 42-43% pins, and ≈6% homostyles in 1988 and 1989. Pins produced more flowers and fruits than thrums, and a greater proportion of their ovaries produced more than one nutlet. However, the number of fruits produced by pins and thrums in 1988 did not differ when plant size is a covariate in ANCOVA. The number of seeds produced by plants in 1989 is also unaffected by floral morph when size is a covariate. Instead, plant size is the most important predictor of fecundity in both years. The average percent fruit sets (fruits/ovaries) of the morphs did not differ in either year. Herbivory was more common in 1988 than in 1989, but there was no difference in the frequency with which pins and thrums were attacked. Furthermore, the effect of herbivory on fecundity did not differ for the morphs. The fecundity of neither morph was limited by pollen in 1989. Apparent gender specialization was a consequence of subtle differences in the vegetative sizes of the morphs.     

Multidrug resistance evaluation by confocal microscopy in primary urothelial cancer explant colonies

Assessing functional multidrug resistance (MDR) status in clinical biopsy material using drug autofluorescence has potential applications to clinical management. The small size of many cystoscopy specimens has led us to develop, as an alternative to flow cytometry, a protocol for studying epirubicin accumulation in adherent colonies of primary bladder cancer cells viewed live andin situ by confocal microscopy. The limitations to quantitation inherent in this technique are compensated for by preservation of cellular organisation and the elimination of non-malignant cells. Biopsy material is disaggregated and explanted into culture-grade petri dishes. After incubation for three to seven days plaques of epithelial cells have developed. Classical patterns of sensitive and resistant drug distribution are observed. Cells of the rolled edges of the colony accumulate more drug than those of the inner epithelial monolayer. Some central areas of larger colonies give the appearance of drug arrested at the intercellular junctions to give a fenestrated pattern. These observations contribute to the understanding of mechanisms in MDR as well as forming the basis for a clinical urological MDR evaluation protocol.