RAPD analysis of induced mutants of groundnut (Arachis hypogaea L.)

Radiation-induced mutants of groundnut cv. Spanish Improved showing distinct morphological differences and the parent were screened for RAPD variability. The analysis revealed characteristic band differences among the 12 mutants and the parent. The polymorphic RAPD bands were dominant in the F₁ and segregated in a Mendelian fashion in the F₂. The RAPD technique brought out greater genome variability than RFLP.     

Genetic analysis of the 5-azacytidine sensitivity of Escherichia coli K-12.

DNA containing 5-azacytidine (5-azaC) has been shown to form stable detergent-resistant complexes with cytosine methylases. We reasoned that if 5-azaC treatment causes protein-DNA cross-links in vivo, then mutations in DNA repair and recombination genes may increase the sensitivity of a cell to 5-azaC. We found that although recA (defective) and lexA (induction-negative) mutants of Escherichia coli were very sensitive to the drug, mutations in uvrA and ung genes had little effect on cell lethality. The sensitivity of recA strains to 5-azaC was dose dependent and was enhanced by the overproduction of a DNA cytosine methylase in the cell. Unexpectedly, a strain of E. coli carrying a recA mutation and a deletion of the DNA cytosine methylase gene (dcm) was found to be significantly sensitive to 5-azaC. Study of mutations in the pyrimidine salvage pathway of E. coli suggests that direct phosphorylation of 5-azaC, rather than phosphorylation of its degradation products, is largely responsible for the lethal effects of the drug. The addition of uracil to the growth medium has little effect on cell lethality of recA mutants, but it partially reversed the inhibition of cell growth caused by 5-azaC. This reversal of the bacteriostatic effects of the drug could not be achieved by adding cytosine or orotic acid to the growth medium and required the presence of functional UMP-pyrophosphorylase (gene upp) in the cell.     

Immune Modulation with Sulfasalazine Attenuates Immunopathogenesis but Enhances Macrophage-Mediated Fungal Clearance during Pneumocystis Pneumonia

Although T cells are critical for host defense against respiratory fungal infections, they also contribute to the immunopathogenesis of Pneumocystis pneumonia (PcP). However, the precise downstream effector mechanisms by which T cells mediate these diverse processes are undefined. In the current study the effects of immune modulation with sulfasalazine were evaluated in a mouse model of PcP-related Immune Reconstitution Inflammatory Syndrome (PcP-IRIS). Recovery of T cell-mediated immunity in Pneumocystis-infected immunodeficient mice restored host defense, but also initiated the marked pulmonary inflammation and severe pulmonary function deficits characteristic of IRIS. Sulfasalazine produced a profound attenuation of IRIS, with the unexpected consequence of accelerated fungal clearance. To determine whether macrophage phagocytosis is an effector mechanism of T cell-mediated Pneumocystis clearance and whether sulfasalazine enhances clearance by altering alveolar macrophage phagocytic activity, a novel multispectral imaging flow cytometer-based method was developed to quantify the phagocytosis of Pneumocystis in vivo. Following immune reconstitution, alveolar macrophages from PcP-IRIS mice exhibited a dramatic increase in their ability to actively phagocytose Pneumocystis. Increased phagocytosis correlated temporally with fungal clearance, and required the presence of CD4⁺ T cells. Sulfasalazine accelerated the onset of the CD4⁺ T cell-dependent alveolar macrophage phagocytic response in PcP-IRIS mice, resulting in enhanced fungal clearance. Furthermore, sulfasalazine promoted a TH2-polarized cytokine environment in the lung, and sulfasalazine-enhanced phagocytosis of Pneumocystis was associated with an alternatively activated alveolar macrophage phenotype. These results provide evidence that macrophage phagocytosis is an important in vivo effector mechanism for T cell-mediated Pneumocystis clearance, and that macrophage phenotype can be altered to enhance phagocytosis without exacerbating inflammation. Immune modulation can diminish pulmonary inflammation while preserving host defense, and has therapeutic potential for the treatment of PcP-related immunopathogenesis.     

Direct rhizogenesis and establishment of fast growing normal root organ culture of Withania somnifera Dunal

Direct rooting from leaf explants of Withania somnifera was achieved on half strength Murashige and Skoog’s medium supplemented with 15 g l⁻¹ sucrose, and different concentrations of growth regulators. Basal medium supplemented with 2.85 μM indoleacetic acid and 9.85 μM indolebutyric acid achieved maximum number of roots with 100% response. The roots were cultured on MS liquid medium for the establishment of root-organ culture with the same plant growth regulators and incubated on an orbital shaker at 80 rpm at 25 ± 2 °C. A root biomass of 6.15 ± 0.17 g was obtained after 5 weeks. When 1 g roots were inoculated to 2.5 l bubble column reactor, 47 g roots were obtained after 6 weeks. The concentration of alkaloids was increased as compared to field grown roots. The maximum concentration of withanolides (10 mg g⁻¹ dry weight) was obtained in the bioreactor.     

Two closely linked tomato HKT coding genes are positional candidates for the major tomato QTL involved in Na+/K+ homeostasis

The location of major quantitative trait loci (QTL) contributing to stem and leaf [Na⁺] and [K⁺] was previously reported in chromosome 7 using two connected populations of recombinant inbred lines (RILs) of tomato. HKT1;1 and HKT1;2, two tomato Na⁺‐selective class I‐HKT transporters, were found to be closely linked, where the maximum logarithm of odds (LOD) score for these QTLs located. When a chromosome 7 linkage map based on 278 single‐nucleotide polymorphisms (SNPs) was used, the maximum LOD score position was only 35 kb from HKT1;1 and HKT1;2. Their expression patterns and phenotypic effects were further investigated in two near‐isogenic lines (NILs): 157‐14 (double homozygote for the cheesmaniae alleles) and 157‐17 (double homozygote for the lycopersicum alleles). The expression pattern for the HKT1;1 and HKT1;2 alleles was complex, possibly because of differences in their promoter sequences. High salinity had very little effect on root dry and fresh weight and consequently on the plant dry weight of NIL 157‐14 in comparison with 157‐17. A significant difference between NILs was also found for [K⁺] and the [Na⁺]/[K⁺] ratio in leaf and stem but not for [Na⁺] arising a disagreement with the corresponding RIL population. Their association with leaf [Na⁺] and salt tolerance in tomato is also discussed.     

On single and multiple models of protein families for the detection of remote sequence relationships

Background  

The detection of relationships between a protein sequence of unknown function and a sequence whose function has been characterised enables the transfer of functional annotation. However in many cases these relationships can not be identified easily from direct comparison of the two sequences. Methods which compare sequence profiles have been shown to improve the detection of these remote sequence relationships. However, the best method for building a profile of a known set of sequences has not been established. Here we examine how the type of profile built affects its performance, both in detecting remote homologs and in the resulting alignment accuracy. In particular, we consider whether it is better to model a protein superfamily using a single structure-based alignment that is representative of all known cases of the superfamily, or to use multiple sequence-based profiles each representing an individual member of the superfamily.     

A novel surface protein of Trichomonas vaginalis is regulated independently by low iron and contact with vaginal epithelial cells

Background  

Trichomonosis caused by Trichomonas vaginalis is the number one, non-viral sexually transmitted disease (STD) that affects more than 250 million people worldwide. Immunoglobulin A (IgA) has been implicated in resistance to mucosal infections by pathogens. No reports are available of IgA-reactive proteins and the role, if any, of this class of antibody in the control of this STD. The availability of an IgA monoclonal antibody (mAb) immunoreactive to trichomonads by whole cell (WC)-ELISA prompted us to characterize the IgA-reactive protein of T. vaginalis.     

Sex and rank in competitive brood hierarchies influence stress levels in nestlings of a sexually dimorphic bird

Studies of sibling competition within brood hierarchies have rarely assessed simultaneously the effects of sex and rank in the brood hierarchy on traits other than offspring mortality and differential growth. We studied the expression of heat-shock proteins (Hsps) to assess the physiological stress response to different combinations of sex and position within competitive brood hierarchies in the black kite Milvus migrans (Bodd.), a sexually dimorphic raptor showing facultative siblicide. Senior males showed higher stress levels than did senior females and younger siblings of each sex as revealed by Hsp60 values. The analysis of Hsp70 levels indicated that nestlings from broods in which the senior chick was a male showed higher stress levels than did nestlings from broods in which the senior chick was a female. In addition, levels of Hsp60 were related negatively to nutritional condition expressed as levels of plasmatic albumin. This suggests that the sex of senior chicks may be key in determining their stress level and that of their siblings, which is probably associated with sibling competition by fighting within brood hierarchies. The comparatively higher stress levels of senior males (and their siblings) may be a consequence of their ability to exploit their potential advantage from being the head start while avoiding a possible competitive disadvantage from being the smaller sex, independent of environmental conditions determining the probability of brood reduction. Differential stress levels depending on sex and rank in the brood hierarchy may be a consequence of parental control of offspring behaviour through differential resource allocation (e.g. yolk androgens) or it may reflect adaptations of particular chicks (senior males) to enhance their competitive ability within brood hierarchies.  © 2006 The Linnean Society of London, Biological Journal of the Linnean Society , 2006, 88 , 383–390.