Species specificity of iron delivery in hybridomas

Summary Studies with Human x Human (HxH), Human x Mouse (HxM), and Mouse x Mouse (MxM) hybridomas have enabled us to define specific factors that affect hybridoma growth in a species-specific manner. Three transferrins and three lipophilic iron chelates have been tested for their ability to support hybridoma proliferation and antibody production. The results of these studies demonstrate that HxH hybridomas do not respond to bovine transferrin a⁺ concentrations up to 100 μg/ml and are approximately 100-fold less responsive to mouse transferrin than to human transferrin. HxM and MxM hybridomas respond equally to human or mouse transferrin but are 100-fold less sensitive to bovine transferrin. An antibody to the human transferrin receptor inhibited the growth-promoting activity of human or mouse transferrin on HxH hybridomas but was ineffective on HxM hybridomas. This semonstrated the functionality of the human transferrin receptor in HxH hybridomas and that human, mouse, and bovine transferrin were interacting through the mouse transferrin receptor in HxM hybridomas. HxH and HxM hybridomas respond similarly to three different iron chelates exhibiting 80 to 110% of the growth response to human transferrin. MxM hybridomas fail to respond to the iron chelates at similar concentrations, suggesting that the human genome present in the other hybridoma species confers a unique ability for utilizing iron when delivered in this form.     

Developmental modulation of deoxyribonucleic acid-binding proteins of Bacillus subtilis during sporulation stages

The isolation of deoxyribonucleic acid (DNA)-binding proteins from various stages of growth and sporulation of Bacillus subtilis is described. After adsorption and elution from phosphocellulose, the proteins were fractionated according to their ability to adsorb to denatured calf thymus DNA-cellulose or native B. subtilis DNA-cellulose. The proteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and purification was monitored by a nitrocellulose filter binding assay. Approximately 1% of the proteins in the crude extract adsorbed to denatured calf thymus DNA-cellulose and 0.1% adsorbed to native B. subtilis DNA-cellulose. Each class of proteins varied qualitatively and quantitatively as sporulation proceeded. Several proteins from the exponential phase of growth that bound to denatured DNA were lost by T(0), whereas at T(5) new polypeptides appeared. Fewer changes in the profile of proteins with affinity for native DNA were observed between exponential phase and T(0); however, the dominant species in these eluates were clearly different.     

Detecting non-neutral heterogeneity across a region of DNA sequence in the ratio of polymorphism to divergence

Natural selection, in the form of balancing selection or selective sweeps, can result in a decoupling of the amounts of molecular polymorphism and divergence. Thus natural selection can cause some areas of DNA sequence to have greater silent polymorphism, relative to divergence between species, than other areas. It would be useful to have a statistical test for heterogeneity in the polymorphism to divergence ratio across a region of DNA sequence, one that could identify heterogeneity greater than that expected from the neutral processes of mutation, drift, and recombination. The only currently available test requires that a region be arbitrarily divided into sections that are compared with each other, and the subjectivity of this division could be problematic. Here a test is proposed in which runs of polymorphic and fixed sites are counted, where a "run" is a set of one or more sites of one type preceded and followed by the other type. The number of runs is smaller than otherwise expected if polymorphisms are clumped together. By simulating neutral evolution and comparing the observed number of runs to the simulations, a statistical test is possible which does not require any a priori decisions about subdivision.      

Hydrolysis, synthesis, and release of acetylcholine in the isolated heart

The occurrence of unhydrolyzed acetylcholine (ACh) in the cardiac perfusate during vagal stimulation in the absence of cholinesterase inhibition has been demonstrated by several methods. Because some ACh was found unhydrolyzed in the extracellular space for several seconds after vagal stimulation (half-time of decay 2.5 s), it appears that the prolonged time course of the cardiac responses to bursts of vagal activity is determined by a slow rate of transmitter inactivation (diffusion plus hydrolysis) in addition to slowly operating postsynaptic mechanisms mediated by activation of the muscarinic receptor. The neuronal uptake of choline in isolated heart preparations was found to be Na+ dependent, sensitive to hemicholinium 3, and activated by vagal stimulation. Activation occurred after a delay of 1 or 2 min and slowly faded within 5 min after stimulation. Resting release of ACh was insensitive to extracellular Ca2+ and to muscarinic feedback inhibition, in contrast to the evoked transmitter release. Inasmuch as atropine increased ACh release by vagal and field stimulation to the same extent, muscarinic feedback inhibition is likely to occur at postganglionic parasympathetic neurons. Adrenergic agonists and propranolol did not significantly change the release of ACh.     

Inversion polymorphism in Drosophila obscura

The inversion polymorphism of Drosophila obscura Fallen, a European species of the obscura group of the subgenus Sophophora, is described. A total of 21 inversions have been recorded; they are located in the five large chromosomes of the species (a dot chromosome is also present) and form 25 gene arrangements present in the species' natural populations. Strains from five different countries were studied. Two of these inversions were found to be pericentric, and the remaining were paracentric. The presence of "hot" points (multibreaks) was noticed. The distribution of the relative lengths of inversions conformed to the Van Valen-Levins distribution, contrary to what happens in D. subobscura. Observations also showed that there is no crossover inhibition between nonoverlapping inversions. The phylogeny of chromosome C was reconstructed; the existence of several arrangements not found was postulated; and the primitive gene arrangement, linking D. obscura to its closely related species was identified, as well as the primitive gene arrangements of the other chromosomes. Photographic maps of the chromosomes of D. obscura are provided here.     

The phylogeny of nine species of the Drosophila obscura group inferred by the banding homologies of chromosomal regions. I. Element B

The phylogenetic relationships among nine Drosophila species belonging to the obscura group were investigated by establishing the segments displaying banding homologies in their element B (equivalent to the U element of D. subobscura). The phylogenetic ordering of the species was accomplished using overlapping inversions. Two African species, D. kitumensis and D. microlabis, were investigated. These species are homosequential for their element B gene arrangement but differ from that of D. obscura by several rearrangements. Drosophila obscura seems to be most closely related to D. subsilvestris, from which the respective element B gene arrangements differ at least by six inversions. Three species, D. obscura, D. ambigua, and D. tristis, are closely related and form a cluster. Drosophila obscura displays an element B polymorphism for a pericentric inversion for which D. ambigua is fixed for one gene arrangement and D. tristis for the other. Both D. ambigua and D. tristis share a short distal inversion in the small arm of the chromosome, and differ in this respect from D. obscura. Drosophila madeirensis, D. guanche, and D. subobscura all share the same element B gene arrangement, which is acrocentric, but metacentric in all the other species mentioned. It was found that the gene arrangements of the species from the obscura cluster seem to occupy an intermediate position between those of the species of the D. subobscura cluster and those of the African one. The data reported generally are in good agreement with information provided in the literature.