An initiation site for meiotic gene conversion in the yeast Saccharomyces cerevisiae

An initiation site for meiotic gene conversion has been identified in the promoter region of the ARG4 gene of Saccharomyces cerevisiae. The chromosome on which initiation occurs is the recipient of genetic information during gene conversion.     

Clonal anergy induced in mature V beta 6+ T lymphocytes on immunizing Mls-1b mice with Mls-1a expressing cells

Tolerance to self-antigens has been shown to develop during ontogeny as a result of the clonal deletion of self-reactive T cells. Tolerance, or better 'nonresponsiveness', to specific antigens can also be induced in adult animals but the mechanism(s) involved are not well understood. Most murine T-helper cells that express the V beta 6 T-cell receptor gene segment are specific for Mls-1a antigens. We have therefore been able to use an anti-V beta 6 monoclonal antibody to follow the fate of Mls-1a specific T cells in adult Mls-1b mice made specifically unresponsive to Mls-1a. We show that the induced unresponsiveness is not due to clonal deletion, but rather to clonal anergy. The anergic V beta 6 T-helper cells express IL-2 receptors and undergo limited blastogenesis in vitro upon stimulation, but do not produce IL-2, in marked contrast to V beta 6 cells from naive mice. Our data appear to represent an in vivo correlate for the induction of anergy that has been observed in T-cell lines in vitro.     

Intercellular adhesion molecule-1 is an endothelial cell adhesion receptor for Plasmodium falciparum

The primary event in the pathogenesis of severe malaria in Plasmodium falciparum infection is thought to be adherence of trophozoite- and schizont-infected erythrocytes to capillary endothelium, a process called sequestration. Identifying the endothelial molecules used as receptors is an essential step in understanding this disease process. Recent work implicates the membrane glycoprotein CD36 (platelet glycoprotein IV; refs 2-5) and the multi-functional glycoprotein thrombospondin as receptors. Although CD36 has a widespread distribution on microvascular endothelium, it may not be expressed on all capillary beds where sequestration occurs, especially in the brain. The role of thrombospondin in cell adhesion, in vitro or in vivo, is less certain. We have noticed that some parasites bind to human umbilical-vein endothelial cells independently of CD36 or thrombospondin. To screen for alternative receptors, we have developed a novel cell-adhesion assay using transfected COS cells, which confirms that CD36 is a cell-adhesion receptor. In addition, we find that an endothelial-binding line of P. falciparum binds to COS cells transfected with a complementary DNA encoding intercellular adhesion molecule-1. As this molecule is widely distributed on capillaries and is inducible, this finding may be relevant to the pathogenesis of severe malaria.     

T-cell receptor delta-chain can substitute for alpha to form a beta delta heterodimer

Specific monoclonal antibodies have made possible the identification of two T-cell antigen receptor (TCR) heterodimers, alpha beta TCR and gamma delta TCR. Formation of these receptors is largely separated by the preferential pairing of alpha-TCR with beta and gamma-TCR with delta, the sequential rearrangement and expression of the TCR loci during thymic development and the deletion of the delta-loci either prior to or concomitant with alpha-rearrangement in alpha beta TCR cells. Here we show that delta-TCR can substitute for alpha in pairing with beta to form a beta delta heterodimer. This receptor is expressed on the cell surface of the T-leukaemia cell line DND41 as analysed with beta- and delta-specific monoclonal antibodies. We suggest that a variety of factors including, for example, the deletion of the delta-TCR loci, can now be understood as exclusion mechanisms operating to prevent not only the formation of gamma delta receptors, but also of beta delta T-cell receptors, thereby promoting the numerically dominant alpha beta TCR lineage. Nevertheless, some developing T-cells that do not rearrange the alpha-loci may express the beta delta TCR as described here.     

Extrusion of calcium from rod outer segments is driven by both sodium and potassium gradients

Calcium is transported across the surface membrane of both nerve and muscle by a Na+-dependent mechanism, usually termed the Na:Ca exchange. It is well established from experiments on rod outer segments that one net positive charge enters the cell for every Ca2+ ion extruded by the exchange, which is generally interpreted to imply an exchange stoichiometry of 3 Na+:1 Ca2+. We have measured the currents associated with the operation of the exchange in both forward and reversed modes in isolated rod outer segments and we find that the reversed mode, in which Ca2+ enters the cell in exchange for Na+, depends strongly on the presence of external K+. The ability of changes in external K+ concentration ([K+]o) to perturb the equilibrium level of [Ca2+]i indicates that K+ is co-transported with calcium. From an examination of the relative changes of [Ca2+]o, [Na+]o, [K+]o and membrane potential required to maintain the exchange at equilibrium, we conclude that the exchange stoichiometry is 4 Na+:1 Ca2+, 1 K+ and we propose that the exchange should be renamed the Na:Ca, K exchange. Harnessing the outward K+ gradient should allow the exchange to maintain a Ca2+ efflux down to levels of internal [Ca2+] that are considerably lower than would be possible with a 3 Na+:1 Ca2+ exchange.     

Signal transduction through the CD4 receptor involves the activation of the internal membrane tyrosine-protein kinase p56lck

The CD4 T-cell surface antigen is an integral membrane glycoprotein of relative molecular mass 55,000 which binds class II major histocompatibility complex (MHC) molecules expressed on antigen presenting cells (APCs). It is thought to stabilize physical interactions between T cells and APCs (for a review, see ref. 1). Evidence is accumulating that suggests that CD4 can transduce an independent signal during T-cell activation. It has recently been shown that CD4 expressed on human and murine T cells is physically associated with the Src-related tyrosine protein kinase p56lck (refs 7, 8). These results indicate that CD4 can function as a signal transducer and suggest that tyrosine phosphorylation events may be important in CD4-mediated signalling. Here, we present evidence that cross-linking of the CD4 receptor induces a rapid increase in the tyrosine-specific protein kinase activity of p56lck and is associated with the rapid phosphorylation of one of the subunits (zeta) of the T-cell receptor complex on tyrosine residues. These data provide direct evidence for a specific CD4 signal transduction pathway that is mediated through p56lck and suggest that some of the tyrosine phosphorylation events detected during antigen-mediated T-cell activation may result from signalling through this surface molecule.     

Vesicular removal by oligodendrocytes of membrane attack complexes formed by activated complement

Oligodendrocytes synthesize myelin in the central nervous system and maintain it in lamellar sheaths around axons. Techniques for studying oligodendrocyte development in vitro can be used, indirectly, to investigate the myelin injury that occurs in human and experimental demyelinating disease. Cell-mediated immune mechanisms are necessary but not sufficient to induce myelin damage in vivo; more recently complement has also been implicated in the pathogenesis both of multiple sclerosis and experimental allergic encephalomyelitis. Previously we have demonstrated that antibody-independent complement activation occurs in vitro at the oligodendrocyte surface. Here we show that the ensuing oligodendrocyte injury is reversible, and that recovery involves the release of membrane-attack complex-enriched vesicles from the surface of viable cells. The demonstration of morphologically and immunochemically identical vesicles in the cerebrospinal fluid of patients with multiple sclerosis suggests that reversible complement-mediated injury contributes to myelin damage in vivo.     

A recombinant immunotoxin consisting of two antibody variable domains fused to Pseudomonas exotoxin

Antibodies and growth factors have been chemically coupled to different toxins to produce cytotoxic molecules that selectively kill cells bearing appropriate antigens or receptors. Antibody-toxin conjugates (immunotoxins) produced using conventional chemical coupling techniques have several undesirable characteristics. The smallest binding unit of an antibody is an Fv fragment which consists of a light and heavy chain variable domain. Recently, active single chain Fv fragments of antibodies have been produced in Escherichia coli by attaching the light and heavy chain variable domains together with a peptide linker. Here we describe the construction and expression in E. coli of a single chain antibody toxin fusion protein, anti-Tac(Fv)-PE40, in which the variable regions of anti-Tac, a monoclonal antibody to the p55 subunit of the human interleukin-2 receptor, are joined in peptide linkage to PE40, a modified form of Pseudomonas exotoxin lacking its binding domain. Anti-Tac(Fv)-PE40 was very cytotoxic to two interleukin-2 receptor-bearing human cell lines but was not cytotoxic to receptor-negative cells.     

Vesicle fusion following receptor-mediated endocytosis requires a protein active in Golgi transport

In reconstitution studies N-ethylmaleimide, a sulphydryl alkylating reagent, inhibits both fusion of endocytic vesicles and vesicular transport in the Golgi apparatus. We show here that the same N-ethylmaleimide-sensitive factor that catalyses the vesicle-mediated transport within Golgi stacks is also required for endocytic vesicle fusion. Thus, it is likely that a common mechanism for vesicle fusion exists for both the secretory and endocytic pathways of eukaryotic cells.     

Sequence of an unusually large protein implicated in regulation of myosin activity in C. elegans

The Caenorhabditis elegans gene unc-22 encodes a very large muscle protein, called twitchin, which consists of a protein kinase domain and several copies of two short motifs. The sequence of twitchin has unexpected similarities to the sequences of proteins of the immunoglobulin superfamily, cell adhesion molecules and vertebrate muscle proteins, including myosin light-chain kinase. These homologies, together with results from earlier genetic and molecular analyses, indicate that twitchin is involved in a novel mechanism of myosin regulation.