Similarities in Restriction Fragment Patterns of Mitochondrial DNAs of PhytophthoraSpecies Strongly Depend on the Restriction Enzyme Used Due to Heterogeneous Base Distribution and Sequence Conservation

Mitochondrial DNAs of six morphologically different Phytophthora species were digested with 15 restriction enzymes. The numbers of restriction fragments obtained differed considerably from those theoretically expected for random base distribution. Enzymes with relatively many G and C in their recognition sequences produced significantly larger numbers of fragments. Moreover, fragments generated by most of these enzymes were more often shared by two or more species than those from enzymes with more A and T in their recognition sequence. It is concluded that base distribution in mitochondrial DNA of Phytophthora is heterogeneous,AT-rich stretches occurring scattered over the mitochondrial genome and GC-rich regions present in conserved sequences, presumably genes. A practical consequence for taxonomic RFLP studies is that optimal enzymes can be selected, depending on the desired level of resolution.     

Tetrapod phylogeny inferred from 18S and 28S ribosomal RNA sequences and a review of the evidence for amniote relationships [published erratum appears in Mol Biol Evol 1991 May;8(3):398]

The 18S ribosomal RNAs of 21 tetrapods were sequenced and aligned with five published tetrapod sequences. When the coelacanth was used as an outgroup, Lissamphibia (living amphibians) and Amniota (amniotes) were found to be statistically significant monophyletic groups. Although little resolution was obtained among the lissamphibian taxa, the amniote sequences support a sister-group relationship between birds and mammals. Portions of the 28S ribosomal RNA (rRNA) molecule in 11 tetrapods also were sequenced, although the phylogenetic results were inconclusive. In contrast to previous studies, deletion or down- weighting of base-paired sites were found to have little effect on phylogenetic relationships. Molecular evidence for amniote relationships is reviewed, showing that three genes (beta-hemoglobin, myoglobin, and 18S rRNA) unambiguously support a bird-mammal relationship, compared with one gene (histone H2B) that favors a bird- crocodilian clade. Separate analyses of four other genes (alpha- crystallin A, alpha-hemoglobin, insulin, and 28S rRNA) and a combined analysis of all sequence data are inconclusive, in that different groups are defined in different analyses and none are strongly supported. It is suggested that until sequences become available from a broader array of taxa, the molecular evidence is best evaluated at the level of individual genes, with emphasis placed on those studies with the greatest number of taxa and sites. When this is done, a bird-mammal relationship is most strongly supported. When regarded in combination with the morphological evidence for this association, it must be considered at least as plausible as a bird-crocodilian relationship.      

Constitutive expression of LIGHT on T cells leads to lymphocyte activation, inflammation, and tissue destruction.

LIGHT, a member of the TNF family of cytokines (homologous to lymphotoxin, exhibits inducible expression and competes with HSV glycoprotein D for herpesvirus entry mediator, a receptor expressed on T cells), is induced on activated T cells and mediates costimulatory and antitumor activity in vitro. Relatively little information is available on the in vivo effects of LIGHT expression, particularly within the T cell compartment. In this work, we describe transgenic mice that express human LIGHT under the control of the CD2 promoter, resulting in constitutive transgene expression in cells of the T lymphocyte lineage. LIGHT-transgenic animals exhibit abnormalities in both lymphoid tissue architecture and the distribution of lymphocyte subsets. They also show signs of inflammation that are most severe in the intestine, along with tissue destruction of the reproductive organs. These LIGHT-mediated effects were recapitulated when immune-deficient mice were reconstituted with bone marrow from LIGHT-transgenic donor mice. T cells in the LIGHT-transgenic mice have an activated phenotype and mucosal T cells exhibit enhanced Th1 cytokine activity. The results indicate that LIGHT may function as an important regulator of T cell activation, and implicate LIGHT signaling pathways in inflammation focused on mucosal tissues.     

Genomic characterization of LIGHT reveals linkage to an immune response locus on chromosome 19p13.3 and distinct isoforms generated by alternate splicing or proteolysis.

LIGHT is a member of the TNF cytokine superfamily that signals through the lymphotoxin (LT)beta receptor and the herpesvirus entry mediator. LIGHT may function as a costimulatory factor for the activation of lymphoid cells and as a deterrent to infection by herpesvirus, which may provide significant selective pressure shaping the evolution of LIGHT. Here, we define the molecular genetics of the human LIGHT locus, revealing its close linkage to the TNF superfamily members CD27 ligand and 4-1BB ligand, and the third complement protein (C3), which positions LIGHT within the MHC paralog on chromosome 19p13.3. An alternately spliced isoform of LIGHT mRNA that encodes a transmembrane-deleted form is detected in activated T cells and gives rise to a nonglycosylated protein that resides in the cytosol. Furthermore, membrane LIGHT is shed from the cell surface of human 293 T cells. These studies reveal new mechanisms involved in regulating the physical forms and cellular compartmentalization of LIGHT that may contribute to the regulation and biological function of this cytokine.     

Cutting edge: a novel viral TNF receptor superfamily member in virulent strains of human cytomegalovirus.

The UL144 open reading frame found in clinical isolates of human CMV (HCMV) encodes a structural homologue of the herpesvirus entry mediator, a member of the TNFR superfamily. UL144 is a type I transmembrane glycoprotein that is expressed early after infection of fibroblasts; however, it is retained intracellularly. A YXXZ motif in the highly conserved cytoplasmic tail contributes to UL144 subcellular distribution. The finding that no known ligand of the TNF family binds UL144 suggests that its mechanism of action is distinct from other known viral immune evasion genes. Specific Abs to UL144 can be detected in the serum of a subset of HCMV seropositive individuals infected with HIV. This work establishes a novel molecular link between the TNF superfamily and herpesvirus that may contribute to the ability of HCMV to escape immune clearance.     

The lymphotoxin-beta receptor is necessary and sufficient for LIGHT-mediated apoptosis of tumor cells

LIGHT is a tumor necrosis factor (TNF) ligand superfamily member, which binds two known cellular receptors, lymphotoxin-beta receptor (LTbetaR) and the herpesvirus entry mediator (HveA). LIGHT is a homotrimer that activates proapoptotic and integrin-inducing pathways. Receptor binding residues via LIGHT were identified by introducing point mutations in the A' --> A" and D --> E loops of LIGHT, which altered binding to LTbetaR and HveA. One mutant of LIGHT exhibits selective binding to HveA and is inactive triggering cell death in HT29.14s cells or induction of ICAM-1 in fibroblasts. Studies with HveA- or LTbetaR-specific antibodies further indicated that HveA does not contribute, either cooperatively or by direct signaling, to the death pathway activated by LIGHT. LTbetaR, not HveA, recruits TNF receptor-associated factor-3 (TRAF3), and LIGHT-induced death is blocked by a dominant negative TRAF3 mutant. Together, these results indicate that TRAF3 recruitment propagates death signals initiated by LIGHT-LTbetaR interaction and implicates a distinct biological role for LIGHT-HveA system.     

Evaluation of the <Emphasis Type="Italic">OPTC</Emphasis> gene in primary open angle glaucoma: functional significance of a silent change

Background  

We investigated the molecular basis of primary open-angle glaucoma (POAG) using Opticin (OPTC) as a candidate gene on the basis of its expression in the trabecular meshwork cells involved in the disease pathogenesis. Two hundred POAG patients and 100 controls were enrolled in this study. The coding sequence of OPTC was amplified by PCR from genomic DNA of POAG patients, followed by SSCP, DHPLC and DNA sequencing. Subsequent bioinformatic analysis, site-directed mutagenesis, quantitative RT-PCR and western blot experiments were performed to address the functional significance of a 'silent' change in the OPTC coding region while screening for mutations in POAG patients.