Cytology of long-term desiccation in the desert cyanobacterium Chroococcidiopsis (Chroococcales)

Young and old cultures (up to 66 months) of two Chroococcidiopsis sp. strains isolated from the Negev desert, Israel, were examined by epifluorescence and electron microscopy. In old cultures, cell viability and autofluorescence were lower than in young cultures. An increase was seen with age in the polysaccharide content of the sheaths of nanocytes and nanocyte mother cells, and a decrease of phycobiliproteins was also seen. In the oldest cultures most of the cells were dead and in various stages of degeneration. Single living cells were scattered among the dead ones. No resting cells were formed in the oldest cultures, but many cell groups showed highly electron-dense sheaths and, in the cytoplasm, ribosomes and glycogen. These changes in cell structure may have a role in preventing water loss from the cell.     

Brain Purines in a Genetic Mouse Model of Lesch-Nyhan Disease

Abstract: Mice carrying a mutation in the gene encoding the purine salvage enzyme hypoxanthine-guanine phosphoribosyltransferase (HPRT) have recently been produced to provide an animal model for Lesch-Nyhan disease. The current-studies were conducted to characterize the consequences of the mutation on the expression of HPRT and to characterize potential changes in brain purine content in these mutants. Our results indicate that the mutant animals have no detectable HPRT-immunoreactive material on western blots and no detectable HPRT enzyme activity in brain tissue homogenates, confirming that they are completely HPRT deficient (HPRT^-). Despite the absence of HPRT-mediated purine salvage, the animals have apparently normal brain purine content. However, de novo purine synthesis, as measured by [¹⁴C]formate incorporation into brain purines, is accelerated four- to fivefold in the mutant animals. This increase in the synthesis of purines may protect the HPRT^- mice from potential depletion of brain purines despite complete impairment of HPRT-mediated purine salvage.     

Location of the major barriers to water and ion movement in young roots of Zea mays L.

The main barriers to the movement of water and ions in young roots of Zea mays were located by observing the effects of wounding various cell layers of the cortex on the roots' hydraulic conductivities and root pressures. These parameters were measured with a root pressure probe. Injury to the epidermis and cortex caused no significant change in hydraulic conductivity and either no change or a slight decline in root pressure. Injury to a small area of the endodermis did not change the hydraulic conductivity but caused an immediate and substantial drop in root pressure. When large areas of epidermis and cortex were removed (15–38% of total root mass), the endodermis was always injured and root pressure fell. The hydraulic conductance of the root increased but only by a factor of 1.2–2.7. The results indicate that the endodermis is the main barrier to the radial movement of ions but not water. The major barrier to water is the membranes and apoplast of all the living tissue. These conclusions were drawn from experiments in which hydrostatic-pressure differences were used to induce water flows across young maize roots which had an immature exodermis and an endodermis with Casparian bands but no suberin lamellae or secondary walls. The different reactions of water and ions to the endodermis can be explained by the huge difference in the permeability of membranes to these substances. A hydrophobic wall barrier such as the Casparian band should have little effect on the movement of water, which permeates membranes and, perhaps, also the Casparian bands easily. However, hydrophobic wall depositions largely prevent the movement of ions. Several hours after wounding the endodermis, root pressure recovered to some extent in most of the experiments, indicating that the wound in the endodermis had been partially healed.Abbreviations Lp_r hydraulic conductivity of root; T_1/2 = half-time of water exchange between root xylem and external medium This research was supported by a grant from EUROSILVA (project no. 39473C) to E.S., and by a Bilateral Exchange Grant jointly funded by the Deutsche Forschungsgemeinschaft and the Natural Sciences and Engineering Research Council of Canada to C.A.P. We thank Mr. Burkhard Stumpf for his excellent technicial assistance.     

Chicken oocyte growth: receptor-mediated yolk deposition

During the rapid final stage of growth, chicken oocytes take up massive amounts of plasma components and convert them to yolk. The oocyte expresses a receptor that binds both major yolk lipoprotein precursors, vitellogenin (VTG) and very low density lipoprotein (VLDL). In the present study, in vivo transport tracing methodology, isolation of coated vesicles, ligand- and immuno-blotting, and ultrastructural immunocytochemistry were used for the analysis of receptor-mediated yolk formation. The VTG/VLDL receptor was identified in coated profiles in the oocyte periphery, in isolated coated vesicles, and within vesicular compartments both outside and inside membrane-bounded yolk storage organelles (yolk spheres). VLDL particles colocalized with the receptor, as demonstrated by ultrastructural visualization of VLDL-gold following intravenous administration, as well as by immunocytochemical analysis with antibodies to VLDL. Lipoprotein particles were shown to reach the oocyte surface by passage across the basement membrane, which possibly plays an active and selective role in yolk precursor accessibility to the oocyte surface, and through gaps between the follicular granulosa cells. Following delivery of ligands from the plasma membrane into yolk spheres, proteolytic processing of VTG and VLDL by cathepsin D appears to correlate with segregation of receptors and ligands which enter disparate sub-compartments within the yolk spheres. In small, quiescent oocytes, the VTG/VLDL receptor was localized to the central portion of the cell. At onset of the rapid growth phase, it appears that this pre-existing pool of receptors redistributes to the peripheral region, thereby initiating yolk formation. Such a redistribution mechanism would obliterate the need for de novo synthesis of receptors when the oocyte's energy expenditure is to be utilized for plasma membrane synthesis, establishment and maintenance of intracellular topography and yolk formation, and preparation for ovulation.     

The human granulocyte-macrophage colony-stimulating factor receptor is capable of initiating signal transduction in NIH3T3 cells.

The ability of the receptor for the hematopoietic cytokine granulocyte-macrophage colony-stimulating factor (GM-CSF) to function in non-hematopoietic cells is unknown. NIH3T3 fibroblasts were transfected with cDNAs encoding the alpha and beta subunit of the human GM-CSF receptor and a series of stable transformants were isolated that bound GM-CSF with either low (KD = 860 - > 1000 pM) or high affinity (KD = 20-80 pM). Low affinity receptors were not functional. However, the reconstituted high affinity receptors were found to be capable of activating a number of signal transduction pathways, including tyrosine kinase activity, phosphorylation of Raf-1, and the transient induction of c-fos and c-myc mRNAs. The activation of protein tyrosine phosphorylation by GM-CSF in NIH3T3 cells was rapid (< 1 min) and transient (peaking at 5-20 min) and resulted in the phosphorylation of proteins of estimated molecular weights of 42, 44, 52/53 and 58-60 kDa. Some of these proteins co-migrated with proteins from myeloid cells that were phosphorylated on tyrosine residues in response to GM-CSF. In particular, p42 and p44 were identified as mitogen-activated protein kinases (MAP kinases), and the phosphorylation on tyrosine residues of p42 and p44 MAP kinases occurred at the same time as the phosphorylation of Raf-1. However, despite evidence for activation of many mitogenic signal transduction molecules, GM-CSF did not induce significant proliferation of transfected NIH3T3 cells. These results suggest that murine fibroblasts contain signal transducing molecules that can effectively interact with the human GM-CSF receptor, and that are sufficient to activate at least some of the same signal transduction pathways this receptor activates in myeloid cells, including activation of one or more tyrosine kinase(s). However, the level of activation of signal transduction is either below a threshold of necessary activity or at least one mitogenic signal necessary for proliferation is missing.     

Drug resistance in rat colon cancer cell lines is associated with minor changes in susceptibility to cytotoxic cells

The development of resistance to anticancer drugs urges the search for different treatment modalities. Several investigators have reported the concomitant development of drug resistance and resistance to natural killer (NK), lymphokine-activated killer (LAK) or monocyte/macrophage cell lysis, while others described unchanged or even increased susceptibility. We investigated this subject in the rat colon carcinoma cell line, CC531-PAR, which is intrinsically multidrug-resistant (MDR), and in three sublines derived from this parental cell line: a cell line with an increased MDR phenotype (CC531-COL), a revertant line from CC531-COL (CC531-REV), which demonstrates enhanced sensitivity to anticancer drugs of the MDR phenotype, and an independently developed cisplatin-resistant line (CC531-CIS). In a 4-h⁵¹Cr-release assay we found no difference in susceptibility to NK cell lysis. No significant differences in lysability by adherent LAK (aLAK) cells were observed in a 4-h assay. In a prolonged 20-h⁵¹Cr-release assay an enhanced sensitivity to aLAK-cell-mediated lysis was observed in the revertant, P-glycoprotein-negative cell line and in the cisplatin-resistant cell line (CC531-CIS). None of the cell lines was completely resistant to lysis by aLAK cells. Therefore, a role for immunotherapy in the treatment of drug-resistant tumors remains a realistic option.     

BVDV and innate immunity.

Infection with bovine viral diarrhea virus (BVDV) is prevalent in the cattle population worldwide. The virus exists in two biotypes, cytopathic and non-cytopathic, depending on the effect of the viruses on cultured cells. BVDV may cause transient and persistent infections which differ fundamentally in the host's antiviral immune response. Transient infection may be due to both cytopathic and non-cytopathic biotypes of BVDV and leads to a specific immune response. In contrast, only non-cytopathic BVD viruses can establish persistent infection as a result of infection of the embryo early in its development. Persistent infection is characterized by immunotolerance specific for the infecting viral strain. In this paper we discuss the role of innate immune responses in the two types of infection. In general, both transient and persistent infections are associated with an increased frequency of secondary infections. Associated with the increased risk of such infections are, among others, impaired bacteria killing and decreased chemotaxis. Interestingly, non-cytopathic BVDV fails to induce interferon type I in cultured bovine macrophages whereas cytopathic biotypes readily trigger this response. Cells infected with non-cytopathic BVDV are also resistant to induction of interferon by double stranded RNA, a potent interferon inducer signalling the presence of viral replication in the cell. Thus, non-cytopathic BVDV may dispose of a mechanism suppressing a key element of the antiviral defence of the innate immune system. Since interferon is also important in the activation of the adaptive immune response, suppression of this signal may be essential for the establishment of persistent infection and immunotolerance.