Transforming capacities of avian erythroblastosis virus mutants deleted in the erbA or erbB oncogenes

Mutants of avian erythroblastosis virus (AEV) were constructed by deleting large nucleotide segments in each of the viral oncogenes termed v-erbA and v-erbB. Mutants in erbA (erbA ^?B⁺) retained the ability to transform fibroblasts in vitro, and these cells exhibited most of the transformation characteristics that typify wild-type AEV-transformed fibroblasts. In addition, the mutants induced small erythroid colonies upon infection of bone marrow cells in culture. Chickens inoculated with erbA ^?B⁺ virus or with erbA ^?B⁺-transformed cells developed sarcomas or atypical erythroid leukemias. The erythroid cells transformed in vivo or in vitro by the erbA ^?B⁺ viruses appeared not to be as tightly blocked in differentiation as wild-type transformed cells. In contrast, fibroblasts infected with the erbA ⁺B^? mutant resembled normal cells in all transformation parameters tested, and no bone marrow cell transformation was observed with the mutant. The results indicate that the main transforming properties of AEV are encoded in erbB and that its effects are enhanced by erbA.     

Verification of the structure of the complex sex chromosome system in Lagorchestes conspicillatus Gould (Marsupialia: Mammalia)

The complex sex chromosome system of Lagorchestes conspicillatus has been reinvestigated using G-banding, Hoechst 33258 sensitivity, and Ag staining. These investigations demonstrate that, as proposed, three exchanges have been involved in the evolution of this system. An autosome was translocated to the original X and the homologue of that autosome was translocated to the original Y. An additional autosome has been translocated to the Y. There is no sex vesicle at meiosis in the male, and no association between the original X and Y elements of the compound chromosomes. The inadequacies of the present terminology for complex sex chromosomes are considered and an alternative system suggested.     

Relationship between enhanced turnover of phosphatidylinositol and lymphocyte activation by mitogens.

1. Various lectins [phaseolus vulgaris phytohaemagglutinin, Glycine max (soy-bean) agglutinin, Triticum vulgaris (wheat-germ) agglutinin and Axinella polyploides agglutinin] and antibodies to pig Ig (immunoglobulin) that are found by pig lymphocytes were assessed in terms of their capacities to stimulate lymphocyte transformation and to enhance phosphatidylinositol turnover. Transformation was measured after 45h of culture by incorporation of [6-(3)H]thymidine into DNA, whereas phosphatidylinositol metabolism was assessed after 1h of cultuis and G. max agglutinins and rabbit antibodies to pig Ig) increased phosphatidylinositol turnover, but non-transforming agents (T. vulgaris and A. polyploides agglutinins and Fab fragments of rabbit antibodies to pig Ig) failed to induce any significant enhancement. Subsequent cross-linkage of the bound, non-transforming Fab fragments with a goat antiserum to rabbit Ig stimulated transformation and phosphatidylinositol turnover. 3. Each transforming agent gave characteristic optimal dose responses that were similar for both phosphatidylinositol turnover and transformation. 4. The results indicate that activation of T- and B-lymphocytes is accompanied by enhanced phosphatidylinositol turnover and that in the case of B-cells this enhancement depends on the cross-linkage of surface receptors. They are consistent with the proposal that turnover represents an essential early step in the transformation process.     

Analysis of a deleted MC29 provirus: gag sequences are not required for fibroblast transformation.

Recovered avian myelocytomatosis virus HBI is an MC29-related virus that induces lymphoid tumors in chickens rather than the predominant neoplastic disease induced by wild-type MC29 (namely, endotheliomas). An analysis of the structure of the HBI provirus(es) in the tumors demonstrated that the provirus(es) could be either full size or deleted. One tumor was found to be clonal in that it contained a single provirus which had been partially deleted; this raised a question concerning the role of this provirus in the maintenance of tumor growth. To characterize the detailed structure of this provirus and determine its biological activity, it was molecularly cloned from tumor DNA. Sequencing confirmed that the provirus contained a deletion which effectively removed the whole gag gene. However, the provirus was shown to encode a myc-specific protein, presumably initiating from within the myc gene, and to be biologically active when it was transfected onto quail embryo fibroblasts. Our results suggest that myc alone is sufficient to transform quail embryo fibroblasts and to maintain tumor growth in vivo.     

Vitronectin—A major cell attachment-promoting protein in fetal bovine serum

Bovine serum is a constituent of most media used for the culture of animal cells. The adhesion-promoting properties of serum are generally attributed to fibronectin, yet there have been frequent reports of other adhesion-promoting molecules in bovine serum. Using a technique in which adhesive proteins are visualized after separation by SDS-PAGE, we graphically confirm the presence of a second cell attachment protein in bovine serum and present the evidence that this molecule is the bovine equivalent of vitronectin. The molecular size of this protein is in the same range as the size of the adhesive human plasma protein, vitronectin. The bovine protein also shared with human vitronectin an affinity for glass, and it could be purified by a combination of glass bead and ion exchange chromatography. The isolated bovine protein had varying proportions of an 80 and a 65 kD polypeptide. It showed immunological cross-reactivity with anti-human vitronectin and with anti-human somatomedin B. Somatomedin B is a serum peptide which has a NH2-terminal sequence identical to that of human vitronectin. The identity of the bovine protein as vitronectin was established by showing that its NH2-terminal amino acid sequence is strongly homologous with those of human vitronectin and somatomedin B. Quantitation of the adhesive activities of fibronectin and vitronectin in bovine plasma and fresh serum showed that more activity is associated with vitronectin than with fibronectin. The preponderance of vitronectin was particularly clear in fetal bovine serum intended for cell culture. In various batches, cell attachment activity attributable to vitronectin was 8-16-fold greater than that of fibronectin, making vitronectin the main adhesive protein in routine cell culture media.     

Detachment of cells from culture substrate by soluble fibronectin peptides

The synthetic cell attachment-promoting peptides from fibronectin (Pierschbacher, M. D., and E. Ruoslahti, 1984, Nature (Lond.)., 309:30-33) were found to detach cultured cells from the substratum when added to the culture in a soluble form. Peptides ranging in length from tetrapeptide to heptapeptide and containing the active L-arginyl-glycyl-L-aspartic acid (Arg-Gly-Asp) sequence had the detaching activity, whereas a series of different peptides with chemically similar structures had no detectable effect on any of the test cells. The Arg-Gly-Asp-containing peptides caused detachment of various cell lines of different species and histogenetic origin. Studies with defined substrates showed that the active peptides could inhibit the attachment of cells to vitronectin in addition to fibronectin, indicating that vitronectin is recognized by cells through a similar mechanism as fibronectin. The peptides did not inhibit the attachment of cells to collagen. However, cells cultured on collagen-coated plastic for 24-36 h, as well as cells with demonstrable type I or type VI collagen in their matrix, were susceptible to the detaching effect of the peptides. These results indicate that the recognition mechanism(s) by which cells bind to fibronectinand vitronectin plays a major role in the substratum attachment of cells and that collagens may not be directly involved in cell-substratum adhesion. Since vitronectin is abundant in serum, it is probably an important component in mediating the attachment of cultured cells. The independence of the effects of the peptide on the presence of serum and the susceptibility of many different cell types to detachment by the peptide show that the peptides perturb an attachment mechanism that is intrinsic to the cells and fundamentally significant to their adhesion.