HIV-specific CD8+ T cell function in children with vertically acquired HIV-1 infection is critically influenced by age and the state of the CD4+ T cell compartment

The immunology of vertical HIV transmission differs from that of adult infection in that the immune system of the infant is not fully matured, and the factors that influence the functionality of CD8(+) T cell responses against HIV in children remain largely undefined. We have investigated CD8(+) T cell responses in 65 pediatric subjects with vertically acquired HIV-1 infection. Vigorous, broad, and Ag dose-driven CD8(+) T cell responses against HIV Ags were frequently observed in children who were older than 3 years of age and maintained CD4(+) T cell counts >400 cells/ micro l. In contrast, younger age or a CD4(+) T cell count <400 cells/ micro l was associated with poor CD8(+) T cell responses and high HIV loads. Furthermore, subjects with a severely depleted and phenotypically altered CD4(+) T cell compartment had circulating Gag-specific CD8(+) T cells with impaired IFN-gamma production. When viral load was not suppressed by antiviral treatment, subjects that fell below the putative age and CD4(+) T cell count thresholds had significantly reduced CD8(+) T cell responses and significantly higher viral loads. Thus, the data suggest that fully effective HIV-specific CD8(+) T cell responses take years to develop despite an abundance of Ag in early life, and responses are further severely impaired, independent of age, in children who have a depleted or skewed CD4(+) T cell compartment. The results are discussed in relation to differences between the neonatal and adult immune systems in the ability to respond to HIV infection.     

Identification of new aromatic cytokinins in Arabidopsis thaliana and Populus x canadensis leaves by LC-(+)ESI-MS and capillary liquid chromatography/frit-fast atom bombardment mass spectrometry

A search for naturally occurring aromatic cytokinins (ARCKs) in Arabidopsis thaliana plants and Populus x canadensis leaves led to the discovery of four new plant hormone substances: 6-(2-methoxybenzylamino)purine (ortho-methoxytopolin, MeoT), 6-(3-methoxybenzylamino)purine (meta-methoxytopolin, MemT) (Fig. 1) and their 9-beta-D-ribofuranosyl derivatives. These substances were identified by liquid chromatography electrospray ionization mass spectrometry [LC (+)ESI-MS] and capillary-liquid chromatography/frit-fast atom bombardment-mass spectrometry [CapLC/frit-FAB-MS] after pre-column derivatization. The chemical structures were subsequently confirmed by chemical synthesis. Because of lack of heavy labelled internal standards, the endogenous levels of methoxytopolins in A. thaliana plants, Populus x canadensis leaves and samples derived from cultures of Agrobacterium tumefaciens strain GV3101 were determined by enzyme-linked immunosorbent assay (ELISA) of HPLC-fractionated extracts. While the levels of MeoT, MemT and their ribosides in A. thaliana shoots and Populus x canadensis leaves were relatively low (approximately 0.25-10 pmol g-1 FW for MeoT and MemT, respectively), the A. tumefaciens strain produced up to 600 times more of the newly identified substances. Cytokinin activity of methoxytopolines was demonstrated in three bioassays testing their ability to stimulate tobacco callus growth, to delay chlorophyll degradation in excised wheat leaves, and to induce betacyanin synthesis in Amaranthus caudatus var. atropurpurea cotyledons. Notably, their anti-senescing activity in the wheat leaf assay exceeded that of BAP and Z by almost 200%. Methoxytopolins are proposed to be new members of the biologically active aromatic cytokinin family, which might have specific physiological functions.     

The role of auxin-binding protein 1 in the expansion of tobacco leaf cells

Tobacco leaf was used to investigate the mechanism of action of auxin-binding protein 1 (ABP1). The distributions of free auxin, ABP1, percentage of leaf nuclei in G2 and the amount of auxin-inducible growth were each determined in control tobacco leaves and leaves over-expressing Arabidopsis ABP1. These parameters were compared with growth of tobacco leaves, measured both spatially and temporally throughout the entire expansion phase. Within a defined window of leaf development, juvenile leaf cells that inducibly expressed Arabidopsis ABP1 prematurely advanced nuclei to the G2 phase. The ABP1-induced increase in cell expansion occured before the advance to the G2 phase, indicating that the ABP1-induced G2 phase advance is an indirect effect of cell expansion. The level of ABP1 was highest at the position of maximum cell expansion, maximum auxin-inducible growth and where the free auxin level was the lowest. In contrast, the position of maximum cell division correlated with higher auxin levels and lower ABP1 levels. Consistent with the correlations observed in leaves, tobacco cells (BY-2) in culture displayed two dose-dependent responses to auxin. At a low auxin concentration, cells expanded, while at a relatively higher concentration, cells divided and incorporated [3H]-thymidine. Antisense suppression of ABP1 in these cells dramatically reduced cell expansion with negligible effect on cell division. Taken together, the data suggest that ABP1 acts at a relatively low level of auxin to mediate cell expansion, whereas high auxin levels stimulate cell division via an unidentified receptor.     

Bus, a bushy Arabidopsis CYP79F1 knockout mutant with abolished synthesis of short-chain aliphatic glucosinolates

A new mutant of Arabidopsis designated bus1-1 (for bushy), which exhibited a bushy phenotype with crinkled leaves and retarded vascularization, was characterized. The phenotype was caused by an En-1 insertion in the gene CYP79F1. The deduced protein belongs to the cytochrome P450 superfamily. Because members of the CYP79 subfamily are believed to catalyze the oxidation of amino acids to aldoximes, the initial step in glucosinolate biosynthesis, we analyzed the level of glucosinolates in a CYP79F1 null mutant (bus1-1f) and in an overexpressing plant. Short-chain glucosinolates derived from methionine were completely lacking in the null mutant and showed increased levels in the overexpressing plant, indicating that CYP79F1 uses short-chain methionine derivatives as substrates. In addition, the concentrations of indole-3-ylmethyl-glucosinolate and the content of the auxin indole-3-acetic acid and its precursor indole-3-acetonitrile were increased in the bus1-1f mutant. Our results demonstrate for the first time that the formation of glucosinolates derived from methionine is mediated by CYP79F1 and that knocking out this cytochrome P450 has profound effects on plant growth and development.     

Selective loss of innate CD4(+) V alpha 24 natural killer T cells in human immunodeficiency virus infection

V alpha 24 natural killer T (NKT) cells are innate immune cells involved in regulation of immune tolerance, autoimmunity, and tumor immunity. However, the effect of human immunodeficiency virus type 1 (HIV-1) infection on these cells is unknown. Here, we report that the V alpha 24 NKT cells can be subdivided into CD4(+) or CD4(-) subsets that differ in their expression of the homing receptors CD62L and CD11a. Furthermore, both CD4(+) and CD4(-) NKT cells frequently express both CXCR4 and CCR5 HIV coreceptors. We find that the numbers of NKT cells are reduced in HIV-infected subjects with uncontrolled viremia and marked CD4(+) T-cell depletion. The number of CD4(+) NKT cells is inversely correlated with HIV load, indicating depletion of this subset. In contrast, CD4(-) NKT-cell numbers are unaffected in subjects with high viral loads. HIV infection experiments in vitro show preferential depletion of CD4(+) NKT cells relative to regular CD4(+) T cells, in particular with virus that uses the CCR5 coreceptor. Thus, HIV infection causes a selective loss of CD4(+) lymph node homing (CD62L(+)) NKT cells, with consequent skewing of the NKT-cell compartment to a predominantly CD4(-) CD62L(-) phenotype. These data indicate that the key immunoregulatory NKT-cell compartment is compromised in HIV-1-infected patients.     

Environmental and auxin regulation of wood formation involves members of the Aux/IAA gene family in hybrid aspen

Indole acetic acid (IAA/auxin) profoundly affects wood formation but the molecular mechanism of auxin action in this process remains poorly understood. We have cloned cDNAs for eight members of the Aux/IAA gene family from hybrid aspen (Populus tremula L. x Populus tremuloides Michx.) that encode potential mediators of the auxin signal transduction pathway. These genes designated as PttIAA1-PttIAA8 are auxin inducible but differ in their requirement of de novo protein synthesis for auxin induction. The auxin induction of the PttIAA genes is also developmentally controlled as evidenced by the loss of their auxin inducibility during leaf maturation. The PttIAA genes are differentially expressed in the cell types of a developmental gradient comprising the wood-forming tissues. Interestingly, the expression of the PttIAA genes is downregulated during transition of the active cambium into dormancy, a process in which meristematic cells of the cambium lose their sensitivity to auxin. Auxin-regulated developmental reprogramming of wood formation during the induction of tension wood is accompanied by changes in the expression of PttIAA genes. The distinct tissue-specific expression patterns of the auxin inducible PttIAA genes in the cambial region together with the change in expression during dormancy transition and tension wood formation suggest a role for these genes in mediating cambial responses to auxin and xylem development.     

Shoot-derived auxin is essential for early lateral root emergence in Arabidopsis seedlings

Lateral root formation is profoundly affected by auxins. Here we present data which indicate that light influences the formation of indole-3-acetic acid (IAA) in germinating Arabidopsis seedlings. IAA transported from the developing leaves to the root system is detectable as a short-lived pulse in the roots and is required for the emergence of the lateral root primordia (LRP) during early seedling development. LRP emergence is inhibited by the removal of apical tissues prior to detection of the IAA pulse in the root, but this treatment has minimal effects on LRP initiation. Our results identify the first developing true leaves as the most likely source for the IAA required for the first emergence of the LRP, as removal of cotyledons has only a minor effect on LRP emergence in contrast to removal of the leaves. A basipetal IAA concentration gradient with high levels of IAA in the root tip appears to control LRP initiation, in contrast to their emergence. A significant increase in the ability of the root system to synthesize IAA is observed 10 days after germination, and this in turn is reflected in the reduced dependence of the lateral root emergence on aerial tissue-derived auxin at this stage. We propose a model for lateral root formation during early seedling development that can be divided into two phases: (i) an LRP initiation phase dependent on a root tip-localized IAA source, and (ii) an LRP emergence phase dependent on leaf-derived IAA up to 10 days after germination.     

Transmembrane topology of the NuoL,M and N subunits of NADH:quinone oxidoreductase and their homologues among membrane-bound hydrogenases and bona fide antiporters

Nicotinamide adenine dinucleotide-reduced form (NADH):quinone oxidoreductase (respiratory Complex I), F420H2 oxidoreductase and complex, membrane-bound NiFe-hydrogenase contain protein subunits homologous to a certain type of bona fide antiporters. In Complex I, these polypeptides (NuoL/ND5, NuoM/ND4, NuoN/ND2) are most likely core components of the proton pumping mechanism, and it is thus important to learn more about their structure and function. In this work, we have determined the transmembrane topology of one such polypeptide, and built a 2D structural model of the protein valid for all the homologous polypeptides. The experimentally determined transmembrane topology was different from that predicted by majority vote hydrophobicity analyses of members of the superfamily. A detailed phylogenetic analysis of a large set of primary sequences shed light on the functional relatedness of these polypeptides.     

High number of diatom species in first-year ice from the Chukchi Sea

Our study describes the species composition of microalgae, primarily diatoms, in two ice cores collected from the Chukchi Sea in early June 1998. At least 251 species were present in 2 cores collected 10 m apart in first-year ice. This is a greater number of algal species in ice from one locality than has been recorded from any other area of the Arctic. Microalgae were distributed throughout the 173-cm-long core, but abundance and species composition varied among different sections of the core, with maximum species richness (108 and 103 species in the 94- to 103- and 103- to 113-cm sections, respectively) occurring in the middle sections. More than 237 species were recorded from this core. Only the bottom 20 cm of the shorter (110 cm) core was analysed and it contained 135 algal species, still an extraordinarily high number of species. Marine species dominated both cores, but typical brackish and freshwater species were also present. None of these species, however, had more than 1% relative abundance. It should be noted, though, that there were several distinct, but unidentified, species of unknown origin. Characteristic ice algal species (e.g. Nitzschia frigida, Navicula pelagica, solitary Navicula spp., in addition to Cylindrotheca closterium) were the numerical dominants in most sections of the long core, but phytoplankton and benthic species were quite abundant in some sections. One section was dominated by a blue-green bacterium, presumably of the genus Anabaena. The species composition is consistent with several different mechanisms for algal incorporation into ice (i.e. seawater filtration ice, seeding from the sea floor, freshwater input). Over time, ice dynamics and sources of ice in the Chukchi Sea appear to result in high numbers of algal species in the ice. It is also likely that season of collection contributed to the high number of species observed. Determining the geographical area of origin for the different species is however difficult, due to the large-scale pattern of ice circulation.