Genome origins of Triticum cylindricum,Triticum triunciale,and Triticum ventricosum (Poaceae) inferred from variation in restriction patterns of repeated nucleotide sequences: a methodological study

Three methods of phylogenetic inferences on polyploid plants employing variation in restriction sites in repeated nucleotide sequences were compared. Allotetraploid Triticum species of well-established origin were used as a model. Methods based on determination of the proportion of restriction fragments shared between a polyploid and its diploid relatives generated biased results because of uneven numbers of restriction fragments among diploid species and presence of common bands in phylogenetically related diploid species. A method employing restriction fragments unique to a diploid species (marker bands) was not affected by either factor and generated results consistent with cytogenetic inferences. It is shown that the latter method can be used to investigate the origin of a polyploid species even when one of its progenitors is extinct or when the polyploid and its diploid progenitors have diverged.     

Virus-induced immunosuppression: kinetic analysis of the selection of a mutation associated with viral persistence.

Infection of neonatal mice with lymphocytic choriomeningitis virus (LCMV) strain Armstrong (ARM) results in a lifelong persistent infection. Viral variants (cytotoxic T lymphocyte [CTL] negative, persistence positive [CTL- P+]) can be isolated from the lymphoid tissues of such mice. Adult mice inoculated with these CTL- P+ viruses fail to generate sufficient cytotoxic T lymphocytes to clear the acute infection and become persistently infected. By contrast, inoculation of a similar dose of the parental ARM virus (CTL+ P-) into adult mice leads to the generation of a vigorous virus-specific CTL response that clears the infection. Sequence analysis revealed a phenylalanine (Phe)-to-Leucine (Leu) change at amino acid 260 of the viral glycoprotein (GP) as a marker for variant viruses with the CTL- P+ phenotype. An RNA PCR assay that detects the variant GP sequence and thus allows kinetic studies of the selection of the Leu at position 260 was developed. We found that although CTL- P+ viruses are known to be lymphotropic, mature T and B cells were not required for the generation and selection of the Leu at GP amino acid 260. Kinetically, in mice infected at birth with LCMV ARM, as early as 3 weeks postinfection the Phe-to-Leu change was found in virus in the serum. By 5 weeks, viral nucleic acid obtained from peritoneal macrophages, spleen, lymph nodes, and liver showed the Phe-to-Leu change. At 2 months postinfection, the Leu change was detected in virus from the thymus, heart, lung, and kidney. By contrast, virus replicating in the central nervous system showed only minimal levels of the Leu change by 4 months and as long as 1 year postinfection. In vitro studies showed that the parental LCMV ARM CTL+ P- virus replicates more efficiently and outcompetes CTL- P+ virus in a cultured neuronal cell line, indicating that differential growth properties in neurons are likely the basis for the selection of the parental virus over the CTL- P+ variant in the brain.     

Phytochrome-mediated effects on extracellular peroxidase activity, lignin content and bending resistance in etiolated Vicia faba epicotyls

Etiolated Vicia faba seedlings were exposed to continuous red light to investigate whether changes in extracellular peroxidase activity were correlated in time and localization with changes in extension growth and/or lignin content in the subapical region of the epicotyl. Continuous red light: (a) increased extracellular peroxidase activity after a lag of ca 0.5 h, followed by a maximum peak after 2.5 h due to slightly acidic isoforms (pI = 6–6.5, according to isoelectrofocusing gels), a minimum after 4 h and a second maximum after 8 h due to acidic isoforms (pI=4–5), (b) increased lignin content and epicotyl resistance to bending after a lag of ca 4 h, i.e. simultaneously with changes in acidic extracellular peroxidase activity, and (c) reduced extension growth to a stable rate after a lag of ca 1 h, not coinciding with the kinetics of any of the extracellular peroxidase isoforms. These effects of continuous red light were at least partially mediated by phytochrome. Tissue printing and anatomical studies revealed red light effects on extracellular peroxidase activity and lignin content mainly in the outer cortical parenchyma. The results are consistent with the involvement of phyto-chrome-mediated effects on extracellular peroxidases (acidic isoforms) in the transduction chain leading to lignin responses to red light.     

Effects of recombinant human gamma interferon on intracellular survival of Bordetella pertussis in human phagocytic cells

Abstract Several studies have demonstrated that Bordetella pertussis has the ability to enter and survive intracellularly within human polymorphonuclear leukocytes (PMNL) and human monocytes/macrophages. The effects of human recombinant gamma interferon (IFN-γ) on the survival of B. pertussis in PMNL and human monocytes, and on the oxidative burst activity of PMNL and human monocytes in response to B. pertussis were assessed in this study. IFN-γ partially increased intracellular killing of phagocytosed B. pertussis in human monocytes, as determined by an orange acridine-crystal violet assay. In contrast, IFN-γ did not enhance intracellular killing of B. pertussis in PMNL. No significant increase of superoxide production was noted in human monocytes in response to B. pertussis when stimulated with various concentrations of IFN-γ. The partial increase of B. pertussis killing by IFN-γ within monocytes, together with poor production of superoxide may explain how B. pertussis can survive within human phagocytic cells, and thus cause a more prolonged course of the disease.     

Three new HLA-G alleles and their linkage disequilibria with HLA-A

Three new allelic forms of the HLA-G DNA sequence (HLA-G*II, HLA-G*III, and HLA-G*IV) have been identified. With the HLA-G*I sequence (previously designated HLA 6.0) as a reference, HLA-G*II shows a silent (G A) mutation at the third base of codon 57, HLA-G*III bears a non-synonymous (A T), but conservative, (Thr Ser) substitution at the first base of codon 31, and HLA-G*IV shows two silent substitutions: (A T) at the third base of codon 107 and (G A) at the third base of codon 57. A rapid method of singling out each allele on genomic DNA has been developed by using polymerase chain reaction amplification followed by restriction endonuclease treatment. Also, more or less strong linkage disequilibria has been found between most HLA-A alleles and either HLA-G*I or *II, both being the most prevalent alleles in the population, with a genotypic frequency of 0.55 and 0.38, respectively; HLA-G*III is very rare and HLA-G*IV has a genotypic frequency of 0.07. An evolutive classification of HLA-A alleles results according to their association with either HLA-G*I or HLA-G*II, which does not correlate with the classical serological cross-reacting groups classification. The finding of a strong and selective A/G linkage disequilibria with most HLA-A alleles, together with the existence of less frequent random A/G associations, may suggest that there exist in different haplotypes true and varied A/G genetic distances (and not a recombinational hotspot). It may be inferred from preliminary data that in primates HLA-A/G haplotypes bearing G*II may have appeared later than those bearing G*I.The nucleotide sequence data reported in this paper have been submitted to the GenBank and EMBL nucleotide sequence databases and have been assigned the following accession numbers: EMBL-X60983 (HLA-G*II), GenBank-M99048 (HLA-G*III), and GenBank-L07784 (HLA-G*IV).The contribution to this paper by P. Morales and A. Corell is equal, and the order of authorship is arbitrary. Correspondence to: A. Arnaiz-Villena.     

Biomass structure and dry matter dynamics of subtropical alluvial and exotic Ligustrum forests at the Río de la Plata,Argentina

Biomass, litterfall, litter standing crop, and decomposition was studied in a native subtropical alluvial forest locally known as Selva Marginal (SM) and an exotic Ligustrum lucidum forest (LF) at the Reserva Integral de Punta Lara, Buenos Aires Province, 34°47S and 58°1W. The alluvial forest site was at the southern limit of distribution of subtropical forests in South America. The Ligustrum forest was invading disturbed areas. Total biomass was 147.7 Mg/ha (86% aboveground and 14% belowground) in the SM, and 71.4 Mg/ha (93% and 7%, respectively) in the LF. Litterfall was 10.3 Mg/ha·yr and 13.8 Mg/ha·yr respectively. Annual leaf decomposition rate was greater for Ligustrum (k=4.07) than for SM species (k=1.48). The mean residence time of aboveground biomass was 12 yr for the SM and 5 yr for the LF. The k₁ values (litterfall/standing crop) were 1.9 and 2.0 for SM and LF respectively. The influence of coastal road and wall in the hydroperiod, native forested wetland ecosystem survival and exotic forest invasion is discussed.     

Improvement of somatic embryogenesis inFeijoa sellowiana berg (Myrtaceae) by manipulation of culture media composition

Summary Somatic embryos could be induced from the cotyledons of zygotic embryos from immature fruits ofFeijoa sellowiana Berg (Feijoa) in the presence of a wide range of concentrations of fructose, glucose, maltose, and sucrose. Mannitol or sorbitol alone were ineffective. The highest frequencies of induction (99%) and the greatest number of somatic embryos per explant (134) were obtained with 0.4M fructose and 0.3M sucrose, respectively. This sucrose concentration also showed greater induction capacity than equimolar combinations of its monosaccharide constituents combined. Somatic embryo development was arrested at the globular stage at concentrations higher than 0.5M of all the sugars tested. When transferred to solid germination medium containing 2.0 mg/liter (5.77μM) gibberellic acid, 0.5 mg/liter (2.32μM) kinetin, and 0.029M sucrose, somatic embryos formed under 0.3 or 0.4M sucrose had better germination capacity than those induced under lower (0.1 and 0.2M) concentrations, as assessed by the frequency of explants presenting germinated embryos and by the number of plants obtained from those explants. On liquid media of similar composition somatic embryos did not germinate. Our data suggest that high (0.3 to 0.4M) carbohydrate levels improve somatic embryogenesis by acting both as carbon source and as osmotic regulator.     

A τ Fragment Containing a Repetitive Sequence Induces Bundling of Actin Filaments

Abstract: Much indirect evidence suggests that the interconnections of actin microfilaments with the microtubule system are mediated by microtubule-associated proteins (MAPs). In this study we provide new data to support the interaction of a specific tubulin-binding domain on τ with actin in vitro. In actin polymerization assays, the synthetic peptide VRSKIGSTENLKHQPGGG, corresponding to the first repetitive sequence of τ protein, increased turbidity at 320 nm in a dose-dependent fashion. A salient feature of the τ peptide-induced assembly process is the formation of a large amount of actin filament bundles, as revealed by electron microscopic analysis. An increase in the τ peptide concentration resulted in a proportional increase in the bundling of actin filaments. It is interesting that a gradual decrease of pH within the range 7.6–4.7 resulted in a higher effect of τ peptide in promoting bundles of actin filaments. A similar pH-dependent effect was observed for τ protein-induced bundling. An analysis of the mechanisms that operate in the peptide induction of actin filament bundles suggests the involvement of electrostatic forces, because the neutralization of ɛ-aminolysyl residues by selective carbamoylation resulted in a complete loss of the peptide induction of actin bundles. The data suggest that a τ repetitive sequence (also found in MAP-2 and MAP-4) containing a common tubulin binding motif may constitute a functional domain on τ for the dynamics of the interconnections between actin filaments and microtu-bules.     

Vaccination to prevent persistent viral infection.

Persistent virus infections are increasingly being recognized as a significant cause of human morbidity and mortality. To establish persistence, a virus must establish infection and evade eradication by the host immune response, in particular by cytotoxic T lymphocytes (CTL). We have studied a virus that establishes persistence in part by suppressing the CTL response of the infected host. The virus persists in many cell types, including lymphocytes and macrophages. We show that prior immunization with a vaccine designed to induce CTL (in the absence of antiviral antibody) confers complete protection against subsequent establishment of persistence in all tissues analyzed. The vaccine can be designed to express as few as 10 amino acids of a viral protein that comprise the CTL epitope. Further, two CTL epitopes for two discrete MHC haplotypes can be successfully used in a single vaccine that protects both strains of mice. Hence, a "string of CTL epitopes" (beads) concept for vaccination is feasible. Finally, the CTL vaccine provided protection against the establishment of persistence by an immunosuppressive virus.     

Regulation of ammonium ion assimilation enzymes inNeurospora crassa nit-2 andms-5 mutant strains

InNeurospora crassa thenit-2 andnmr-1 (ms-5) loci represent the major control genes encoding regulatory proteins that allow the coordinated expression of various systems involved with the utilization of a secondary nitrogen source. In this paper we examine the effect of thenit-2 andms-5 (nmr-1 locus) mutations on the regulation of the ammonium assimilation enzymes, glutamine synthetase and glutamate dehydrogenase, which are regulated by the products of these genes; however, glutamate synthase is not so regulated. Glutamine synthetase and glutamate dehydrogenase levels are also regulated by the amino nitrogen content. We present evidence that thems-5 andgln ^r strains, which behave very similarly in their resistance to glutamine repression, are different and map in different loci.