Evolution of bacterial pathogenesis

The evolution of bacteria is associated with continuous generation of novel genetic variants. The major driving forces in this process are point mutations, genetic rearrangements, and horizontal gene transfer. A large number of human and animal bacterial pathogens have evolved the capacity to produce virulence factors that are directly involved in infection and disease. Additionally, many bacteria express resistance traits against antibiotics. Both virulence factors and resistance determinants are subject to intrastrain genetic and phenotypic variation. They are often encoded on unstable DNA regions. Thus, they can be readily transferred to bacteria of the same species or even to non-related prokaryotes. This review article focuses on the main mechanisms of bacterial microevolution responsible for the rapid emergence of variants with novel virulence and resistance properties. In addition, processes of macroevolution are described with special emphasis on gene transfer and fixation of adaptive mutations in the genome of pathogens.     

Permeability of rat heart myocytes to cytochrome c

Rat heart myocytes undergoing progressive damage demonstrate morphological changes of shortening and swelling followed by the formation of intracellular vacuoles and plasma membrane blebbing. The damaged myocytes displayed impaired N,N'-tetramethyl-p-phenyldiamine (TMPD) ascorbate-stimulated respiratory activity which was restored by the addition of reduced cytochrome c to the cell culture medium. To clarify the role played by cytochrome c in the impairment of cell respiration, polarographic, spectrophotometric and fluorescence as well as electron microscopy imaging experiments were performed. TMPD/ascorbate-stimulated respiratory activity returned to control levels, at approximately 20 microM cytochrome c, establishing the threshold below which the turnover rate by cytochrome c oxidase in the cell depends on cytochrome concentration. Mildly damaged cardiac myocytes, as indicated by cell shortening, retention of visible striations and free-fluorescein exclusion, together with the absence of lactate dehydrogenase leakage and exclusion of trypan blue, were able to oxidize exogenous cytochrome c and were permeable to fluorescein-conjugated cytochrome c. The results, while consistent with an early cytochrome c release observed at the beginning of cell death, elucidate the role played by cytochrome c in the kinetic control of mitochondrial electron transfer under pathological conditions, particularly those involving the terminal part of the respiratory chain. These data are the first to demonstrate that the sarcolemma of cardiac myocytes, damaged but still viable, is permeable to cytochrome c.     

Beyond blood pressure: new roles for angiotensin II

Since the discovery 100 years ago by Tigerstedt and Bergman of renin, an acid protease generating angiotensin peptide, numerous discoveries have advanced our understanding of the renin-angiotensin system (RAS). The recent cloning of angiotensin receptors and the availability of specific receptor ligands have allowed characterization of angiotensin-receptor-mediated actions, and an increasing number of studies using biochemical, pharmacological and molecular biological methods has focused on the many different physiological actions of the RAS in various tissues. Angiotensin II, the main effector peptide of the RAS, exerts most of its known actions in blood pressure control and body fluid homeostasis via the AT, receptor. AT, receptors not only play a role in growth control and cell differentiation but have been implicated in apoptosis and tissue regeneration. This review focuses on the extrarenal functions of angiotensin, especially in neuronal cells and the nervous system, and on recent advances in angiotensin receptor research.     

Notch signalling pathway mediates hair cell development in mammalian cochlea

The mammalian cochlea contains an invariant mosaic of sensory hair cells and non-sensory supporting cells reminiscent of invertebrate structures such as the compound eye in Drosophila melanogaster. The sensory epithelium in the mammalian cochlea (the organ of Corti) contains four rows of mechanosensory hair cells: a single row of inner hair cells and three rows of outer hair cells. Each hair cell is separated from the next by an interceding supporting cell, forming an invariant and alternating mosaic that extends the length of the cochlear duct. Previous results suggest that determination of cell fates in the cochlear mosaic occurs via inhibitory interactions between adjacent progenitor cells (lateral inhibition). Cells populating the cochlear epithelium appear to constitute a developmental equivalence group in which developing hair cells suppress differentiation in their immediate neighbours through lateral inhibition. These interactions may be mediated through the Notch signalling pathway, a molecular mechanism that is involved in the determination of a variety of cell fates. Here we show that genes encoding the receptor protein Notch1 and its ligand, Jagged 2, are expressed in alternating cell types in the developing sensory epithelium. In addition, genetic deletion of Jag2 results in a significant increase in sensory hair cells, presumably as a result of a decrease in Notch activation. These results provide direct evidence for Notch-mediated lateral inhibition in a mammalian system and support a role for Notch in the development of the cochlear mosaic.     

Mutations in ATP2A2, encoding a Ca2+ pump, cause Darier disease

Darier disease (DD) is an autosomal-dominant skin disorder characterized by loss of adhesion between epidermal cells (acantholysis) and abnormal keratinization. Recently we constructed a 2.4-Mb, P1-derived artificial chromosome contig spanning the DD candidate region on chromosome 12q23-24.1. After screening several genes that mapped to this region, we identified mutations in the ATP2A2 gene, which encodes the sarco/endoplasmic reticulum Ca2(+)-ATPase type 2 isoform (SERCA2) and is highly expressed in keratinocytes. Thirteen mutations were identified, including frameshift deletions, in-frame deletions or insertions, splice-site mutations and non-conservative missense mutations in functional domains. Our results demonstrate that mutations in ATP2A2 cause DD and disclose a role for this pump in a Ca(2+)-signalling pathway regulating cell-to-cell adhesion and differentiation of the epidermis.     

The Pendred syndrome gene encodes a chloride-iodide transport protein

Pendred syndrome is the most common form of syndromic deafness and characterized by congenital sensorineural hearing loss and goitre. This disorder was mapped to chromosome 7 and the gene causing Pendred syndrome (PDS) was subsequently identified by positional cloning. PDS encodes a putative transmembrane protein designated pendrin. Pendrin is closely related to a family of sulfate transport proteins that includes the rat sulfate-anion transporter (encoded by Sat-1; 29% amino acid sequence identity), the human diastrophic dysplasia sulfate transporter (encoded by DTD; 32%) and the human sulfate transporter 'downregulated in adenoma' (encoded by DRA; 45%). On the basis of this homology and the presence of a slightly modified sulfate-transporter signature sequence comprising its putative second transmembrane domain, pendrin has been proposed to function as a sulfate transporter. We were unable to detect evidence of sulfate transport following the expression of pendrin in Xenopus laevis oocytes by microinjection of PDS cRNA or in Sf9 cells following infection with PDS-recombinant baculovirus. The rates of transport for iodide and chloride were significantly increased following the expression of pendrin in both cell systems. Our results demonstrate that pendrin functions as a transporter of chloride and iodide, but not sulfate, and may provide insight into thyroid physiology and the pathophysiology of Pendred syndrome.     

Myosins from plants

Molecular cloning and sequence analysis of myosin genes from Arabidopsis thaliana and electron microscopic observation of a myosin from characean alga have revealed that overall structure of plant unconventional myosins is similar to that of the class V myosins. These plant unconventional myosins have two heads, a coiled-coil tail of varied length and a globular tail piece at the end. The tail piece is probably a site for membrane interaction. Characean myosin is of special interest because it can translocate actin filaments at a velocity several times faster than muscle myosin, which must have evolved to support the quick movement of animals in the struggle for their lives.     

Arachidonic acid release by ionomycin and phorbol ester is similar in C127 epithelial cells expressing wild-type or mutated (ΔF508) cystic fibrosis transmembrane conductance regulator

The Ca²⁺ ionophore ionomycin induced cytosolic [Ca²⁺]_i elevation as well as strong activation of Cl⁻ efflux in mouse mammary epithelial cell lines expressing wild-type or mutated (deletion of phenylalaline 508) cystic fibrosis transmembrane conductance regulator (CFTR) or vector. Ionomycin-induced Cl⁻ efflux was abolished by the intracellular Ca²⁺ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid, whereas both activators and inhibitors of phospholipase A₂ had no effect, indicating the involvement of Ca²⁺-dependent Cl^- channels. Stimulation of arachidonic acid release by ionomycin and phorbol ester was not significantly different between wild-type or mutated cell lines, whereas vector-transfected cells exhibited a significant higher release, which was shown to be due to larger amount of immunoreactive cytosolic phospholipase A₂. These results indicate that phospholipase A₂ activity of C127 cells was not influenced by the presence of wild-type or mutated CFTR. Received 27 April 1999; received after revision 11 June 1999; accepted 23 July 1999     

Triplet repeat disorders: discussion of molecular mechanisms

Comparison of the growing number of disorders known to be associated with triplet repeat expansions reveals both common features and a diversity of molecular pathways. Despite significant progress towards the characterization of proteins coded by the mutant genes, the complex nature of these disorders requires identification of all molecular components of the triplet repeat pathways. In this brief review we will discuss recent progress in determining the molecular mechanisms of disorders with unstable trinucleotide mutations. Received 13 January 1999; received after revision 8 March 1999; accepted 9 March 1999     

Genomic instability in Gadd45a-deficient mice.

Gadd45a-null mice generated by gene targeting exhibited several of the phenotypes characteristic of p53-deficient mice, including genomic instability, increased radiation carcinogenesis and a low frequency of exencephaly. Genomic instability was exemplified by aneuploidy, chromosome aberrations, gene amplification and centrosome amplification, and was accompanied by abnormalities in mitosis, cytokinesis and growth control. Unequal segregation of chromosomes due to multiple spindle poles during mitosis occurred in several Gadd45a -/- cell lineages and may contribute to the aneuploidy. Our results indicate that Gadd45a is one component of the p53 pathway that contributes to the maintenance of genomic stability.