Arabidopsis thaliana ENHANCED DISEASE SUSCEPTIBILITY1 (EDS1) controls defense activation and programmed cell death conditioned by intracellular Toll-related immune receptors that recognize specific pathogen effectors. EDS1 is also needed for basal resistance to invasive pathogens by restricting the progression of disease. In both responses, EDS1, assisted by its interacting partner, PHYTOALEXIN-DEFICIENT4 (PAD4), regulates accumulation of the phenolic defense molecule salicylic acid (SA) and other as yet unidentified signal intermediates. An Arabidopsis whole genome microarray experiment was designed to identify genes whose expression depends on EDS1 and PAD4, irrespective of local SA accumulation, and potential candidates of an SA-independent branch of EDS1 defense were found. We define two new immune regulators through analysis of corresponding Arabidopsis loss-of-function insertion mutants. FLAVIN-DEPENDENT MONOOXYGENASE1 (FMO1) positively regulates the EDS1 pathway, and one member (NUDT7) of a family of cytosolic Nudix hydrolases exerts negative control of EDS1 signaling. Analysis of fmo1 and nudt7 mutants alone or in combination with sid2-1, a mutation that severely depletes pathogen-induced SA production, points to SA-independent functions of FMO1 and NUDT7 in EDS1-conditioned disease resistance and cell death. We find instead that SA antagonizes initiation of cell death and stunting of growth in nudt7 mutants.

Globally suppressed T-cell function has been described in many patients with cancer to be a major hurdle for the development of clinically efficient cancer immunotherapy. Inhibition of antitumor immune responses has been mainly linked to inhibitory factors present in cancer patients. More recently, increased frequencies of CD4 ؉ CD25 hi regulatory T cells (T reg cells) have been described as an additional mechanism reducing immunity. We assessed 73 patients with B-cell chronic lymphocytic leukemia ( IntroductionHuman and murine CD4 ϩ CD25 ϩ T cells contain cells that suppress antigen-specific T-cell immune responses. [1][2][3][4][5] These naturally occurring regulatory CD4 ϩ CD25 ϩ T cells originate from the thymus and play a central role in the maintenance of peripheral tolerance by suppression of autoreactive T-cell populations. In murine models, regulatory T cells (T reg cells) prevent autoimmune and inflammatory diseases 1,6,7 and inhibit antitumor immune responses. [8][9][10][11][12] Although a truly unique marker for T reg cells is still not available, several molecules have been associated with these cells including cytotoxic T lymphocyte-associated protein 4 (CTLA4), [13][14][15][16] glucocorticoid-induced tumor necrosis factor receptor-related protein (GITR, TNFRSF18), 17,18 Forkhead box P3 (FOXP3), [19][20][21] Lselectin (CD62L, SELL), 22,23 and OX40 antigen (CD134, TNFRSF4). 23,24 In humans, T reg cells are enriched within the CD4 ϩ CD25 hi population, whereas CD4 ϩ CD25 lo T cells represent mainly previously activated T helper cells. 25 These CD4 ϩ CD25 hi T reg cells inhibit proliferation and cytokine release by conventional CD4 ϩ CD25 Ϫ T cells. 26 Decrease of these cells was found in patients with autoimmune diseases, 27-31 whereas an increase of T reg cells in patients after allogeneic bone marrow transplantation was associated with a reduced graft-versus-host disease. [32][33][34][35] In patients with malignant melanoma, 36 Hodgkin lymphoma, 37 or ovarian, 38,39 gastric, 40,41 lung, 39,42 breast, 43,44 and pancreatic cancer 43 inhibitory CD4 ϩ CD25 ϩ T cells are also increased. In an elegant study, Curiel et al 38 demonstrated that functional T reg cells were enriched in ascites from women with ovarian cancer, migrated toward CCL22 expressed by tumor cells and tumor-associated macrophages, and specifically inhibited antitumor immunity. Moreover, within this setting, the increase of T reg cells predicted poor survival. 38 Only recently, studies assessing a potential influence of chemotherapy on T reg cells have been initiated. In mice, low-dose cyclophosphamide decreased the number of T reg cells. 45 Based on these observations we were interested in understanding whether CD4 ϩ CD25 hi T cells are also increased and possess inhibitory capacities in B-cell chronic lymphocytic leukemia (CLL) and, if so, to assess the frequency and function in the context of stage of disease and prior therapy. CLL, the most common type of leukemia in the Western hemisphere, 46 is characterized by clonal proliferat...

Immune tolerance is a central mechanism counteracting tumor-specific immunity and preventing effective anticancer immunotherapy. Induction of tolerance requires a specific environment in which tolerogenic dendritic cells (DCs) play an essential role deviating the immune response away from effective immunity. It was recently shown that maturation of DCs in the presence of PGE2 results in upregulation of indoleamine 2,3-dioxygenase (IDO) providing a potential mechanism for the development of DC-mediated Tcell tolerance. Here, we extend these findings, demonstrating a concomitant induction of IDO and secretion of soluble CD25 after DC maturation in the presence of PGE2. While maturation of DCs induced IDO expression on transcriptional level, only integration of PGE2 signaling led to up-regulation of functional IDO protein as well as significant expression of cell-surface and soluble CD25 protein. As a consequence, T-cell proliferation and cytokine production were significantly inhibited, which was mediated mainly by IDO-induced tryptophan depletion. Of importance, we demonstrate that different carcinoma entities associated with elevated levels of PGE2 coexpress CD25 and IDO in peritumoral dendritic cells, suggesting that PGE2 might influence IDO expression in human DCs in the tumor environment. We therefore suggest PGE2 to be a mediator of early events during induction of immune tolerance in cancer.

Owing to its clinical accessibility, peripheral blood is probably the best source for the assessment of differences or changes in gene expression associated with disease or drug response and therapy. Gene expression patterns in peripheral blood cells greatly depend on temporal and interindividual variations. However, technical aspects of blood sampling, isolation of cellular components, RNA isolation techniques and clinical aspects such as time to analysis and temperature during processing have been suggested to affect gene expression patterns. We therefore assessed gene expression patterns in peripheral blood from 29 healthy individuals by using Affymetrix microarrays. When RNA isolation was delayed for 20-24 h-a typical situation in clinical studies-gene signatures related to hypoxia were observed, and downregulation of genes associated with metabolism, cell cycle or apoptosis became dominant preventing the assessment of gene signatures of interindividual variation. Similarly, gene expression patterns were strongly dependent on choice of cell and RNA isolation and preparation techniques. We conclude that for large clinical studies, it is crucial to reduce maximally the time to RNA isolation. Furthermore, prior to study initiation, the cell type of interest should already be defined. Our data therefore will help to optimize clinical studies applying gene expression analysis of peripheral blood to exploit drug responses and to better understand changes associated with disease.