ABSTRACT
Insects successfully occupy most environmental niches and this success depends on surviving a broad range of environmental stressors including temperature, desiccation, xenobiotic, osmotic and infection stress. Epithelial tissues play key roles as barriers between the external and internal environments and therefore maintain homeostasis and organismal tolerance to multiple stressors. As such, the crucial role of epithelia in organismal stress tolerance cannot be underestimated. At a molecular level, multiple cell-specific signalling pathways including cyclic cAMP, cyclic cGMP and calcium modulate tissue, and hence, organismal responses to stress. Thus, epithelial cell-specific signal transduction can be usefully studied to determine the molecular mechanisms of organismal stress tolerance in vivo. This review will explore cell signalling modulation of stress tolerance in insects by focusing on cell signalling in a fluid transporting epithelium--the Malpighian tubule. Manipulation of specific genes and signalling pathways in only defined tubule cell types can influence the survival outcome in response to multiple environmental stressors including desiccation, immune, salt (ionic) and oxidative stress, suggesting that studies in the genetic model Drosophila melanogaster may reveal novel pathways required for stress tolerance.
Subject(s)
Dehydration , Drosophila melanogaster/metabolism , Malpighian Tubules/physiology , Osmotic Pressure/physiology , Oxidative Stress/physiology , Animals , Calcium/metabolism , Cyclic AMP/metabolism , Cyclic GMP/metabolism , Environment , Homeostasis , Malpighian Tubules/cytology , Mucous Membrane/physiology , Signal TransductionABSTRACT
N-Linked glycans have been shown to have an important role in the cell biology of a variety of cell surface glycoproteins, including PrP protein. It has been suggested that glycosylation of PrP can influence the susceptibility to transmissible spongiform encephalopathy and determine the characteristics of the many different strains observed in this particular type of disease. To understand the role of carbohydrates in influencing the PrP maturation, stability, and cell biology, we have produced and analyzed gene-targeted murine models expressing differentially glycosylated PrP. Transgenic mice carrying the PrP substitution threonine for asparagine 180 (G1) or threonine for asparagine 196 (G2) or both mutations combined (G3), which eliminate the first, second, and both glycosylation sites, respectively, have been generated by double replacement gene targeting. An in vivo analysis of altered PrP has been carried out in transgenic mouse brains, and our data show that the lack of glycans does not influence PrP maturation and stability. The presence of one chain of sugar is sufficient for the trafficking to the cell membrane, whereas the unglycosylated PrP localization is mainly intracellular. However, this altered cellular localization of PrP does not lead to any overt phenotype in the G3 transgenic mice. Most importantly, we found that, in vivo, unglycosylated PrP does not acquire the characteristics of the aberrant pathogenic form (PrPSc), as was previously reported using in vitro models.
Subject(s)
Neurons/metabolism , Prions/chemistry , Scrapie/metabolism , Aging , Alleles , Animals , Antibodies, Monoclonal/chemistry , Asparagine/chemistry , Blotting, Northern , Blotting, Southern , Blotting, Western , Brain/metabolism , Carbohydrates/chemistry , Cell Membrane/metabolism , Cells, Cultured , DNA/metabolism , Detergents/pharmacology , Disease Models, Animal , Embryo, Mammalian/cytology , Endopeptidase K/metabolism , Endoplasmic Reticulum/metabolism , Female , Genetic Vectors , Genotype , Glycoproteins/chemistry , Glycosylation , Golgi Apparatus/metabolism , Homozygote , Immunohistochemistry , Male , Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase/chemistry , Mice , Mice, Transgenic , Microscopy, Confocal , Models, Genetic , Mutation , Neurons/cytology , Phenotype , Polymerase Chain Reaction , Polysaccharides/chemistry , RNA/metabolism , RNA, Messenger/metabolism , Recombination, Genetic , Solubility , Stem Cells/cytology , Threonine/chemistry , Time Factors , Type C Phospholipases/metabolismABSTRACT
Expression of the PrP glycoprotein is essential for the development of the transmissible spongiform encephalopathy (TSE) or prion diseases. Although PrP is widely expressed in the mouse, the precise relevance of different PrP-expressing cell types to disease remains unclear. To address this, we generated two lines of floxed PrP gene-targeted transgenic mice using the Cre recombinase-loxP system. These floxed mice allow a functional PrP allele to be either switched "on" or "off." We demonstrate control of PrP expression for both alleles following Cre-mediated recombination, as determined by PrP mRNA and protein expression in the brain. Moreover, we show that Cre-mediated alteration of PrP expression in these mice has a major influence on the development of TSE disease. These floxed PrP mice will allow the involvement of PrP expression in specific cell types following TSE infection to be defined, which may identify potential sites for therapeutic intervention.
