Metatranscriptomics sheds light on the links between the functional traits of fungal guilds and ecological processes in forest soil ecosystems.

Soil fungi belonging to different functional guilds, such as saprotrophs, pathogens, and mycorrhizal symbionts, play key roles in forest ecosystems. To date, no study has compared the actual gene expression of these guilds in different forest soils. We used metatranscriptomics to study the competition for organic resources by these fungal groups in boreal, temperate, and Mediterranean forest soils. Using a dedicated mRNA annotation pipeline combined with the JGI MycoCosm database, we compared the transcripts of these three fungal guilds, targeting enzymes involved in C- and N mobilization from plant and microbial cell walls. Genes encoding enzymes involved in the degradation of plant cell walls were expressed at a higher level in saprotrophic fungi than in ectomycorrhizal and pathogenic fungi. However, ectomycorrhizal and saprotrophic fungi showed similarly high expression levels of genes encoding enzymes involved in fungal cell wall degradation. Transcripts for N-related transporters were more highly expressed in ectomycorrhizal fungi than in other groups. We showed that ectomycorrhizal and saprotrophic fungi compete for N in soil organic matter, suggesting that their interactions could decelerate C cycling. Metatranscriptomics provides a unique tool to test controversial ecological hypotheses and to better understand the underlying ecological processes involved in soil functioning and carbon stabilization.

ENSO and NAO affect long-term leaf litter dynamics and stoichiometry of Scots pine and European beech mixedwoods.

Litterfall dynamics (production, seasonality and nutrient composition) are key factors influencing nutrient cycling. Leaf litter characteristics are modified by species composition, site conditions and water availability. However, significant evidence on how large-scale, global circulation patterns affect ecophysiological processes at tree and ecosystem level remains scarce due to the difficulty in separating the combined influence of different factors on local climate and tree phenology. To fill this gap, we studied links between leaf litter dynamics with climate and other forest processes, such as tree-ring width (TRW) and intrinsic water-use efficiency (iWUE) in two mixtures of Scots pine (Pinus sylvestris L.) and European beech (Fagus sylvatica L.) in the south-western Pyrenees. Temporal series (18 years) of litterfall production and elemental chemical composition were decomposed following the ensemble empirical mode decomposition method and relationships with local climate, large-scale climatic indices, TRW and Scots pine's iWUE were assessed. Temporal trends in N:P ratios indicated increasing P limitation of soil microbes, thus affecting nutrient availability, as the ecological succession from a pine-dominated to a beech-dominated forest took place. A significant influence of large-scale patterns on tree-level ecophysiology was explained through the impact of the North Atlantic Oscillation (NAO) and El Niño-Southern Oscillation (ENSO) on water availability. Positive NAO and negative ENSO were related to dry conditions and, consequently, to early needle shedding and increased N:P ratio of both species. Autumn storm activity appears to be related to premature leaf abscission of European beech. Significant cascading effects from large-scale patterns on local weather influenced pine TRW and iWUE. These variables also responded to leaf stoichiometry fallen 3 years prior to tree-ring formation. Our results provide evidence of the cascading effect that variability in global climate circulation patterns can have on ecophysiological processes and stand dynamics in mixed forests.

Natural killer cells and malignant haemopathies: a model for the interaction of cancer with innate immunity.

Despite recent progress in the therapeutic approach of malignant haemopathies, their prognoses remain frequently poor. Immunotherapy offers an alternative of great interest in this context but defect or abnormal expression of human leukocyte antigens (HLA), frequently observed in cancer cells, limits its efficiency. Natural killer (NK) cells, which are able to kill target cells in a HLA-independent way, represent a novel tool in the treatment of haematological malignancies. Abnormal NK cytolytic function is observed in all the haematological malignancies studied, such as acute leukaemia, myelodysplastic syndromes or chronic myeloid/lymphoid leukaemia. Several mechanisms are involved in the alterations of NK cytotoxicity: decreased expression of activating receptors, increased expression of inhibitory receptors or defective expression of NK ligands on target cells. Further studies are needed to identify how each type of haematological malignancy escapes from the innate immune response. Attempts to increase the expression of activating receptors, to counteract inhibitory receptors expression, or to increase NK cell cytotoxic capacities could overcome tumour escape from innate immunity. These therapies are based on monoclonal antibodies or culture of NK cells in presence of cytokines or dendritic cells. Moreover, many novel drugs used in haematological malignancies [tyrosine kinase inhibitors, IMIDs(®), proteasome inhibitors, demethylating agents, histone deacetylase inhibitors (HDACis), histamine dihydrochloride] display interesting immunomodulatory properties that affect NK cells. These data suggest that combined modalities associating cytotoxic drugs with innate immunity modulators may represent a major breakthrough in tumour eradication.

Thinning affects nutrient resorption and nutrient-use efficiency in two Pinus sylvestris stands in the Pyrenees.

Needle chemical composition was measured, and nutrient resorption, nutrient-use efficiency (NUE), and other indexes were estimated for 24 months in two contrasting natural Pinus sylvestris L. forests in the western Pyrenees in Spain. For each location (Aspurz, 650 m elevation, 7% slope; Garde, 1335 m elevation, 40% slope), there were three reference plots (P0), three plots with 20% of the basal area removed (P20), and three with 30% of the basal area removed (P30). Needle P, Ca, and Mg concentrations were higher in Garde, but N concentration was higher for Aspurz, without differences for K. Nutrient-resorption efficiency of P was higher in Aspurz, of Mg higher in Garde, and there were no differences between sites in N and K. Nutrient-resorption proficiency was significantly higher in the site with lower soil nutrient availability, i.e., for P, Ca, and Mg in Aspurz, but N in Garde (no differences in K); this may be an indicator of nutrient conservation strategy. Annual nutrient productivity (A) was higher for all nutrients in Aspurz, whereas the mean residence time (MRT) was higher in Garde in all nutrients but P. NUE was significantly higher in Garde for all nutrients but P, which was more efficiently used in Aspurz. In both sites, N, P, and K concentrations were higher in the 2002 cohort, Ca in the 2000 cohort, and maximum Mg was found in the 2001 cohort. Thinning caused a reduction of Mg concentration in the 2001 cohort in Aspurz, an increase of Ca resorption proficiency in Aspurz and Mg resorption at both sites, and reduction of P, K, and Mg nutrient response efficiency (NRE) in Garde. Thinning may have caused an increase of the C:Mg ratio through facilitating the development of more biosynthesis apparatus in a more illuminated canopy, but it seemed not to affect resorption in a significant way. Changes in NRE in Garde after thinning show that forest management can affect how trees use nutrients. Our results indicate that the strategy to optimize NUE is different in each stand. In Aspurz (a Mediterranean ecosystem), pine trees carried out resorption more efficiently, while in Garde (a continental forest), trees used nutrients for longer periods of time and reduced their efficiency in using the available soil nutrients after reduced competition by thinning.

Transcription factor MBF-I interacts with metal regulatory elements of higher eucaryotic metallothionein genes.

Metallothionein (MT) gene promoters in higher eucaryotes contain multiple metal regulatory elements (MREs) that are responsible for the metal induction of MT gene transcription. We identified and purified to near homogeneity a 74-kilodalton mouse nuclear protein that specifically binds to certain MRE sequences. This protein, MBF-I, was purified employing as an affinity reagent a trout MRE that is shown to be functional in mouse cells but which lacks the G+C-rich and SP1-like sequences found in many mammalian MT gene promoters. Using point-mutated MREs, we showed that there is a strong correlation between DNA binding in vitro and MT gene regulation in vivo, suggesting a direct role of MBF-I in MT gene transcription. We also showed that MBF-I can induce MT gene transcription in vitro in a mouse extract and that this stimulation requires zinc.

Novel CD28-responsive enhancer activated by CREB/ATF and AP-1 families in the human interleukin-2 receptor alpha-chain locus.

The interaction of interleukin-2 (IL-2) with its receptor (IL-2R) critically regulates the T-cell immune response, and the alpha chain CD25/IL-2Ralpha is required for the formation of the high-affinity receptor. Tissue-specific, inducible expression of the IL-2Ralpha gene is regulated by at least three positive regulatory regions (PRRI, PRRII, and PRRIII), but none responded to CD28 engagement in gene reporter assays although CD28 costimulation strongly amplifies IL-2Ralpha gene transcription. By DNase I hypersensitivity analysis, we have identified a novel TCR-CD3- and CD28-responsive enhancer (CD28rE) located 8.5 kb 5' of the IL-2Ralpha gene. PRRIV/CD28rE contains a functional CRE/TRE element required for CD28 signaling. The T-cell-specific, CD28-responsive expression of the IL-2Ralpha gene appears controlled through PRRIV/CD28rE by cooperation of CREB/ATF and AP-1 family transcription factors.

Elf-1 and Stat5 bind to a critical element in a new enhancer of the human interleukin-2 receptor alpha gene.

The interleukin 2 receptor alpha-chain (IL-2R alpha) gene is a key regulator of lymphocyte proliferation. IL-2R alpha is rapidly and potently induced in T cells in response to mitogenic stimuli. Interleukin 2 (IL-2) stimulates IL-2R alpha. transcription, thereby amplifying expression of its own high-affinity receptor. IL-2R alpha transcription is at least in part controlled by two positive regulatory regions, PRRI and PRRII. PRRI is an inducible proximal enhancer, located between nucleotides -276 and -244, which contains NF-kappaB and SRE/CArG motifs. PRRII is a T-cell-specific enhancer, located between nucleotides -137 and -64, which binds the T-cell-specific Ets protein Elf-1 and HMG-I(Y) proteins. However, none of these proximal regions account for the induction of IL-2R alpha transcription by IL-2. To find new regulatory regions of the IL-2R alpha gene, 8.5 kb of the 5' end noncoding sequence of the IL-2R alpha gene have been sequenced. We identified an 86-nucleotide fragment that is 90% identical to the recently characterized murine IL-2-responsive element (mIL-2rE). This putative human IL-2rE, designated PRRIII, confers IL-2 responsiveness on a heterologous promoter. PRRIII contains a Stat protein binding site that overlaps with an EBS motif (GASd/EBSd). These are essential for IL-2 inducibility of PRRIII/CAT reporter constructs. IL-2 induced the binding of Stat5a and b proteins to the human GASd element. To confirm the physiological relevance of these findings, we carried out in vivo footprinting experiments which showed that stimulation of IL-2R alpha expression correlated with occupancy of the GASd element. Our data demonstrate a major role of the GASd/EBSd element in IL-2R alpha regulation and suggest that the T-cell-specific Elf-1 factor can serve as a transcriptional repressor.

Vanin-1-/- mice exhibit a glutathione-mediated tissue resistance to oxidative stress.

Vanin-1 is an epithelial ectoenzyme with pantetheinase activity and generating the amino-thiol cysteamine through the metabolism of pantothenic acid (vitamin B(5)). Here we show that Vanin-1(-/-) mice, which lack cysteamine in tissues, exhibit resistance to oxidative injury induced by whole-body gamma-irradiation or paraquat. This protection is correlated with reduced apoptosis and inflammation and is reversed by treating mutant animals with cystamine. The better tolerance of the Vanin-1(-/-) mice is associated with an enhanced gamma-glutamylcysteine synthetase activity in liver, probably due to the absence of cysteamine and leading to elevated stores of glutathione (GSH), the most potent cellular antioxidant. Consequently, Vanin-1(-/-) mice maintain a more reducing environment in tissue after exposure to irradiation. In normal mice, we found a stress-induced biphasic expression of Vanin-1 regulated via antioxidant response elements in its promoter region. This process should finely tune the redox environment and thus change an early inflammatory process into a late tissue repair process. We propose Vanin-1 as a key molecule to regulate the GSH-dependent response to oxidative injury in tissue at the epithelial level. Therefore, Vanin/pantetheinase inhibitors could be useful for treatment of damage due to irradiation and pro-oxidant inducers.

[Imaging of the postoperative knee in athletes: articular cartilage, menisci, and ligaments]. / Imagerie du genou du sportif opéré: cartilage, ménisques et ligaments.

This article focuses on the common surgical procedures and reviews the clinical and imaging features of the postoperative knee following prior surgical treatment of meniscal, articular cartilage, and ligamentous injuries.

[Onychomycosis by yeast not common in diabetics of a health center]. / Onicomicosis por levaduras no comunes en diabéticos de un centro de salud.

BACKGROUND: Mexican diabetic population frequently presents mycosis under foot hyperkeratosis; however, in another type of onychomycosis as the ones that is assumed Candida albicans is the causal agent, it is unknown the frequency, the prevalence and if another Candida species or other yeasts are found. OBJECTIVE: Evaluate the frequency of yeasts causing onychomycosis in diabetic patients looked after in public institutions of health of the State of Hidalgo, Mexico, and its association with clinical epidemiological variables. MATERIALS AND METHODS: An observational, descriptive and transversal study was made on 261 patients, from which one nail sample of each one was obtained, used to isolate and identify dermatophytes and yeasts; the results were statistically correlated with 24 epidemiological parameters. The clinical study was done through interrogation and by medical exploration in order to evaluate Tinea pedis and onychomycosis. RESULTS: Onychomycosis were caused by Candida guilliermondii, Candida parapsilosis, Candida glabrata, Candida krusei, Candida spp., Kodamaea ohmeri, Prototheca wickerhamii and unidentified yeasts. The prevalence for general onychomycosis, by dermatophytes, mixed onychomycosis and by yeasts were: 24.1, 19.5, 2.3 and 14.6%, respectively. Patients with significant probability to be diagnosed as having onychomycosis by yeasts are those wearing open shoes (2.59%); technicians and professionals (10.49%) and alcohol drinkers (3.72%). CONCLUSION: The fact that Candida albicans is not present in this study as causal agent of onychomycosis, and emerging and non-common yeasts were indeed isolated, creates new challenges. It is remarked the clinical criterion that when onychomycosis is suspected in diabetics, the diagnosis for culturing dermatophytes and yeasts should be included.

IL-2 and long-term T cell activation induce physical and functional interaction between STAT5 and ETS transcription factors in human T cells.

Activation of Stat5 by many cytokines implies that it cannot alone insure the specificity of the regulation of its target genes. We have evidenced a physical and functional interaction between members of two unrelated transcription factor families, Ets-1, Ets-2 and Stat5, which could contribute to the proliferative response to interleukin 2. Competition with GAS- and EBS-specific oligonucleotides and immunoassays with a set of anti-Stat and anti-Ets families revealed that the IL-2-induced Stat5-Ets complex recognizes several GAS motifs identified as target sites for activated Stat5 dimers. Coimmunoprecipitation experiments evidenced that a Stat5/Ets-1/2 complex is formed in vivo in absence of DNA. GST-pull down experiments demonstrated that the C-terminal domain of Ets-1 is sufficient for this interaction in vitro. Cotransfection experiments in Kit225 T cells resulted in cooperative transcriptional activity between both transcription factors in response to a combination of IL-2, PMA and ionomycin. A Stat5-Ets protein complex was the major inducible DNA-binding complex bound to the human IL-2rE GASd/EBSd motif in long-term proliferating normal human T cells activated by CD2 and CD28. These results suggest that the inducible Stat5-Ets protein interaction plays a role in the regulation of gene expression in response to IL-2 in human T lymphocytes.

Relief of cyclin A gene transcriptional inhibition during activation of human primary T lymphocytes via CD2 and CD28 adhesion molecules.

Cyclin A transcription is cell cycle regulated and induced by cell proliferative signals. To understand the mechanisms underlined in this regulation in normal human cells, we have analysed in vivo protein-DNA interactions at the Cyclin A locus in primary T lymphocytes. Stimulation of purified T lymphocytes by a combination of monoclonal antibodies directed at CD2 and CD28 adhesion molecules gives rise to a long lasting proliferation in the absence of accessory cells. Cyclin A was observed after 4 days of costimulation with anti CD2 + CD28 whereas stimulation by anti CD2 or anti CD28 alone was not effective. In vivo genomic DMS footprinting revealed upstream of the major transcription initiation sites, the presence of at least three protein binding sites, two of which were constitutively occupied. They bind in vitro respectively ATF-1 and NF-Y proteins. The third site was occupied in quiescent cells or in cells stimulated by anti CD2 or anti CD28 alone. The mitogenic combination of anti CD2 + anti CD28 released the footprint as cells were committed to proliferation. Consistent with theses results, nuclear extracts prepared from quiescent cells formed a specific complex with this element, whereas extracts prepared from cells treated with anti CD2 + anti CD28 failed to do so after cells entered a proliferative state.

Physical interaction of the bHLH LYL1 protein and NF-kappaB1 p105.

The LYL1 gene was first identified upon the molecular characterization of the t(7;9)(q35;p13) translocation associated with some human T-cell acute leukemias (T-ALLs). In adult tissues, LYL1 expression is restricted to hematopoietic cells with the notable exclusion of the T cell lineage. LYL1 encodes a basic helix-loop-helix (bHLH) protein highly related to TAL-1, whose activation is also associated with a high proportion of human T-ALLs. A yeast two-hybrid system was used to identify proteins that specifically interact with LYL1 and might mediate its activities. We found that p105, the precursor of NF-kappaB1 p50, was the major LYL1-interacting protein in this system. The association between LYL1 and p105 was confirmed both in vitro and in vivo in mammalian cells. Biochemical studies indicated that the interaction was mediated by the bHLH motif of LYL1 and the ankyrin-like motifs of p105. Ectopic expression of LYL1 in a human T cell line caused a significant decrease in NF-kappaB-dependent transcription, associated with a reduced level of NF-kappaB1 proteins.

Detection of CBFbeta/MYH11 fusion transcripts in acute myeloid leukemia: heterogeneity of cytological and molecular characteristics.

Pericentric inversion of chromosome 16, translocation (16;16) and del(16q), resulting in a chimerical fusion of CBFbeta and MYH11 genes, are typically seen in the M4Eo French-American-British (FAB) classification subset of acute myelogenous leukemia (AML). In this study, we analyzed 70 cases of acute non-lymphoblastic leukemia, mainly of the M4 or M5 type. We report the very unusual presence of the t(16;16) and CBFbeta/MYH11 fusion transcript in an M7 patient. Ten M4Eo and four non-M4Eo patients presented an inv(16), t(16;16) or CBFbeta/MYH11 fusion transcript. In most cases, the common 'A-type' CBFbeta/MYH11 fusion transcript was detected. In addition to the eight different breakpoints and the three alternative splicing variants already described, evidence of a new CBFbeta/MYH11 fusion transcript was found which involves a 785-bp deletion of MYH11. Moreover, two patients had an unusual transcript, to our knowledge only observed once. Only one patient had abnormal eosinophilic differentiation without chromosome 16 cytogenetic abnormalities or detectable CBFbeta/MYH11 fusion. Conversely, only one patient presented CBFbeta/MYH11 fusion without abnormal eosinophilic differentiation. Altogether, our data suggest a correlation between the CBFbeta/MYH11 fusion transcript and characteristic abnormal eosinophilic differentiation, whatever the FAB subtype or the percentage of abnormal eosinophils

Hematological reconstitution and gene therapy: retroviral transfer of the bacterial beta-galactosidase activity into human hematopoietic CD34+ cell populations and into T lymphocytes derived from the peripheral blood.

We report the possibility to transfer marker genes coding for beta-galactosidase activity using retroviral vectors into human peripheral blood CD34+ cells, peripheral blood T-lymphocytes and into the growth factor-dependent human hematopoietic cell line TF-1. Using the MFG-nisLacZ and the FLac vector and various packaging cell lines, we demonstrated retroviral transfer and high expression of a bacterial beta-galactosidase activity induced by the nisLacZ gene or the Sh-ble/LacZ gene. Kinetics of expression of the transgenes were analyzed both in primary cells and cell lines. Absence of cytotoxicity related to the expression of the bacterial beta-galactosidase was assessed in both cell types. These results open interesting prospectives for the use of the beta-galactosidase activity to mark and follow the fate of genetically modified cells isolated from patients prior to reimplantation.

The neuroergonomic evaluation of human machine interface design in air traffic control using behavioral and EGG/ERP measures.

The Air Traffic Control (ATC) environment is complex and safety-critical. Whilst exchanging information with pilots, controllers must also be alert to visual notifications displayed on the radar screen (e.g., warning which indicates a loss of minimum separation between aircraft). Under the assumption that attentional resources are shared between vision and hearing, the visual interface design may also impact the ability to process these auditory stimuli. Using a simulated ATC task, we compared the behavioral and neural responses to two different visual notification designs--the operational alarm that involves blinking colored "ALRT" displayed around the label of the notified plane ("Color-Blink"), and the more salient alarm involving the same blinking text plus four moving yellow chevrons ("Box-Animation"). Participants performed a concurrent auditory task with the requirement to react to rare pitch tones. P300 from the occurrence of the tones was taken as an indicator of remaining attentional resources. Participants who were presented with the more salient visual design showed better accuracy than the group with the suboptimal operational design. On a physiological level, auditory P300 amplitude in the former group was greater than that observed in the latter group. One potential explanation is that the enhanced visual design freed up attentional resources which, in turn, improved the cerebral processing of the auditory stimuli. These results suggest that P300 amplitude can be used as a valid estimation of the efficiency of interface designs, and of cognitive load more generally.

Centrofacial malignant granulomas. Clinicopathologic study of 40 cases and review of the literature.

An important problem in the treatment of centrofacial ulcerations is to establish a precise diagnosis, since similar clinical and microscopic findings can result from many different causes (as in the centrofacial malignant granuloma syndrome [CFMG]). A comprehensive surgical biopsy protocol (known as SNFMI/GMCF), involving microbiology, parasitology, immunology and pathology laboratories, allowed us to evaluate and to treat 40 cases of CFMG, who form the basis of this report. In 13 of them, specific diagnoses were found and curative treatments could be given. In the remaining 27, the optical microscopy pattern met the criteria for CFMG without identifiable origin or the presence of so-called lethal midline granulomas; however, a more precise evaluation with the help of immunofluorescence studies led to the recognition of malignant lymphoma (ulcerative lymphoma of the midface [ULM]). Most of these lymphomas belonged to the T cell lineage; the others were of B lymphoid origin, or, more rarely, of histiocytic origin. Patients with ULM received radiotherapy and chemotherapy with a response rate of 70.3%; however, the toxicity was significant, with frequent occurrence of chemotherapy-induced neutropenia followed by severe infectious facial cellulitis. Six patients were enrolled in a preliminary open trial of treatment with recombinant alpha-2b interferon with little success. Three patients were treated with radiation therapy only, and survived. Thus, CFMG is a syndrome with specific causes and treatments, requiring multiple extensive biopsies to make the correct diagnosis. The recognition of ULM as the cause of the previously called "lethal midline granulomas" leads logically to the use of chemotherapy with growth factors in order to ameliorate its bad prognosis.

Highly efficient retroviral gene transfer into human primary T lymphocytes derived from peripheral blood.

T lymphocytes are a promising cell vehicle for gene therapy purposes. By cocultivating retroviral vector producing cells and target cells, highly efficient gene transfer was achieved with activated human T lymphocytes derived from peripheral blood with vectors carrying different forms of the bacterial beta-galactosidase gene including the regular LacZ gene, the Sh-ble::LacZ gene and the nlsLacZ gene. Infection kinetics of T cells activated by a combination of monoclonal antibodies directed against CD2 and CD28 indicated that the highest efficiencies of transduction were obtained when the cocultivation began 4 days after stimulation. In fact, with the FLac vector, a new retroviral vector which expresses the Sh-ble::LacZ gene, we observed up to 78% transduction efficiency assessed by X-gal staining performed 2 days after the end of the cocultivation. Expression of the transduced genes was observed throughout the period of culture. Neither the cocultivation step nor the expression of the transduced Sh-ble::LacZ gene altered cell culture proliferation or the expression of selected cell surface antigens. In addition, we showed that CD4+ and CD8+ T cells were equally transduced.

Activation of the HIV type 1 long terminal repeat by X-irradiation involves two main Re1/NF-kappa B DNA-binding complexes.

Transcription of human immunodeficiency virus type 1 (HIV-1) is regulated by multiple cis-acting regulatory elements located in the viral long terminal repeats (LTRs). HIV-1 LTR enhancer is activated by a variety of heterologous viral, chemical, and physical agents. Studies have demonstrated that irradiation by X-rays induces transcription under the control of the HIV-1 LTR and that ionizing radiations activate DNA binding of the nuclear transcription factor NF-kappa B. Using various constructs expressing a reporter gene under the control of complete or deleted LTRs of HIV-1, we evidenced that a sequence located in the U3 region was involved in X-ray activation of the HIV-1 LTR in the human colonic carcinoma cell line HT29. The cis-acting element conferring X-ray responsiveness is indistinguishable from HIV NF-kappa B tandem repeat binding sites (HIV-1, kappa B). The present work has examined the effects of X-irradiation on the NF-kappa B transcription factor. Furthermore, we characterized the subunit composition of the two major nuclear NF-kappa B complexes that bind HIV-1 kappa B after X-ray irradiation.

Secretion of extracellular factor(s) induced by X-irradiation activates the HIV type 1 long terminal repeat through its kappaB motif.

X-irradiation has been used in the treatment of several human diseases, including AIDS-related-malignancies. X-irradiation might induce the transcription and the replication of human immunodeficiency virus type 1 (HIV-1) and enhance nuclear factor kappa B (NF-kappaB). In the present article we show that the activation of the HIV-1 long terminal repeat (LTR) by direct X-irradiation can be mimicked by coculture of transfected cells with X-irradiated nontransfected (HIV-1-negative) cells. In the human colonic carcinoma cell line HT29, the activation seems to depend on an extracellular factor(s) released by a cell line treated with X-rays. The HIV-1 LTR cis-acting element conferring X-indirect responsiveness was identified as the kappaB tandem motif. The two main nuclear HIV-1 kappaB-binding complexes activated by X-direct and -indirect irradiation were the NF-kappaB p50/p65 and c-Rel/p65 heterodimers. Nuclear NF-kappaB activation was dependent on protein neosynthesis. It was partially inhibited by 100 microM pyrrolidine dithiocarbamate, a potent antioxidant drug, but was not correlated with a significant decrease in cellular IkappaBalpha. Furthermore, X-irradiation induces the expression of several cytokine genes generally associated with stress response and antibodies against interleukin 6 and TNF-alpha partially inhibited the X-indirect activation of the HIV-1 LTR. The use of protein kinase C (PKC)-specific inhibitor and of forskolin, an adenylate cyclase activator, suggests that a PKC-dependent pathway and the cAMP intracellular concentration could play a role in the X-indirect enhancement of HIV-1 LTR transcription in the HT29 cell line. In addition, supernatants of an X-irradiated HT29 cell culture activated the HIV-1 stimulation in infected peripheral blood monocytes.