SummaryThe plant genome is a highly redundant and dynamic genome. Here, we show that double antisense plants lacking the two major hydrogen peroxide-detoxifying enzymes, ascorbate peroxidase (APX) and catalase (CAT), activate an alternative/redundant defense mechanism that compensates for the lack of APX and CAT. A similar mechanism was not activated in single antisense plants that lacked APX or CAT, paradoxically rendering these plants more sensitive to oxidative stress compared to double antisense plants. The reduced susceptibility of double antisense plants to oxidative stress correlated with suppressed photosynthetic activity, the induction of metabolic genes belonging to the pentose phosphate pathway, the induction of monodehydroascorbate reductase, and the induction of IMMUTANS, a chloroplastic homologue of mitochondrial alternative oxidase. Our results suggest that a co-ordinated induction of metabolic and defense genes, coupled with the suppression of photosynthetic activity, can compensate for the lack of APX and CAT. In addition, our findings demonstrate that the plant genome has a high degree of plasticity and will respond differently to different stressful conditions, namely, lack of APX, lack of CAT, or lack of both APX and CAT.

Glioblastoma multiforme (GBM) is a lethal cancer that responds poorly to radiotherapy and chemotherapy. Glioma cancer-initiating cells have been shown to recapitulate the characteristic features of GBM and mediate chemotherapy and radiation resistance. However, it is unknown whether the cancer-initiating cells contribute to the profound immune suppression in GBM patients. Recent studies have found that the activated form of signal transducer and activator of transcription 3 (STAT3) is a key mediator in GBM immunosuppression. We isolated and generated CD133+ cancer-initiating single colonies from GBM patients and investigated their immunesuppressive properties. We found that the cancer-initiating cells inhibited T-cell proliferation and activation, induced regulatory Tcells, and triggered T-cell apoptosis. The STAT3 pathway is constitutively active in these clones and the immunosuppressive properties were markedly diminished when the STAT3 pathway was blocked in the cancer-initiating cells. These findings indicate that cancer-initiating cells contribute to the immune evasion of GBM and that blockade of the STAT3 pathway has therapeutic potential. Mol Cancer Ther; 9(1); 67-78. ©2010 AACR.

Purpose: Glioblastoma multiforme is a lethal cancer that responds poorly to therapy. Glioblastoma multiforme cancer-initiating cells have been shown to mediate resistance to both chemotherapy and radiation; however, it is unknown to what extent these cells contribute to the profound immunosuppression in glioblastoma multiforme patients and if strategies that alter their differentiation state can reduce this immunosuppression.Experimental Design: We isolated a subpopulation of cells from glioblastoma multiforme that possessed the capacity for self-renewal, formed neurospheres in vitro, were capable of pluripotent differentiation, and could initiate tumors in vivo. The immune phenotype of these cells was characterized including the elaboration of immunosuppressive cytokines and chemokines by ELISA. Functional immunosuppressive properties were characterized based on the inhibition of T-cell proliferation and effector responses, triggering of T-cell apoptosis, and induction of FoxP3 + regulatory T cells. On altering their differentiation state, the immunosuppressive phenotype and functional assays were reevaluated. Results: We found that the cancer-initiating cells markedly inhibited T-cell proliferation and activation, induced regulatory T cells, and triggered T-cell apoptosis that was mediated by B7-H1 and soluble Galectin-3. These immunosuppressive properties were diminished on altering the differentiation of the cancer-initiating cells.

The activation of signal transducer and activator of transcription 3 (STAT3) has been identified as a key mediator that drives the fundamental components of melanoma malignancy, including immune suppression in melanoma patients. Increasing evidence also suggests that regulatory T cells (Tregs) are important in suppressing anti-tumor immunity and play a dominant role in negating efficacious immunotherapy approaches. We hypothesized that WP1066, a novel inhibitor of STAT3 signaling, reverses immune suppression through the inhibition of Tregs and that this contributes to the antitumor activity of these agents against melanoma brain metastases. We found that the mean percentage of peripheral blood mononuclear cells expressing phosphorylated STAT3 (p-STAT3) was significantly elevated in samples from patients with melanoma brain metastases compared to healthy donors, 16.13% ± 2.48% vs 4.17% ± 1.79%. The p-STAT3 inhibitor WP1066 enhanced CD3+ (which contained Tregs) but not CD8+ T cell cytotoxicity against human A375 melanoma cells, indicating that this p-STAT3 blockade agent did not directly activate CD8+ T cells. Furthermore, the p-STAT3 inhibitor did not enhance the cytotoxicity of CD3+CD25-T cells (from which Tregs were excluded), indicating that the enhanced cytotoxicity of WP1066 is secondary to its inhibition of Tregs. This was confirmed by demonstrating that WP1066 inhibited FoxP3+ Treg induction in a dose-dependent manner. Moreover, CD3+ T cells exhibited markedly enhanced levels of phosphorylated ZAP-70, a critical proximal signal in T cell activation, after exposure to WP1066. Similar effects were not observed in Treg-depleted CD3+CD25-T cell populations, confirming that the T cell activation by WP compounds is secondary to their inhibition of the Tregs. These results suggest that WP1066 enhances T cell cytotoxicity against melanoma through inhibition of Tregs.

Eph receptor tyrosine kinases and ephrins are required for axon patterning and plasticity in the developing nervous system. Typically, Eph-ephrin interactions promote inhibitory events; for example, prohibiting the entry of neural cells into certain embryonic territories. Here, we show that distinct subsets of motor neurons that express EphA4 respond differently to ephrin-A5.

The tomato ghost and Arabidopsis immutans variegation mutants define orthologous genes for a chloroplast quinol oxidase that bears similarity to the mitochondrial alternative oxidase. In this study tomato fruit ripening was used as a model to explore the function of this oxidase. It was found that a lack of GH impacts the biogenesis of chloroplasts and chromoplasts, as well as pericarp tissue morphogenesis, during the ripening process. Most ripening-related genes are expressed normally during gh fruit development, but late in the process there is a specific repression of mRNA accumulation from carotenoid biosynthetic genes, perhaps due to retrograde (plastid-to-nucleus) signalling pathways mediated by lycopene. Metabolic profiling revealed that wild-type red-ripe fruit accumulate multiple isomers of phytoene, phytofluene, and z z z z -carotene, but that the composition of these isomers is different in ripe gh fruit. Profiling also revealed that there is an enhanced ratio of b b b b -carotene to lycopene in the ripe gh fruit. This explains the orange colour of these fruit, and is consistent with the proposal that the b b b b -cyclization step plays a pivotal role in regulating the partitioning of pathway intermediates. The total carotenoid content of the mutant fruit is significantly higher than normal due to an accumulation of phytoene. Phytoene accumulation, together with an up-regulation of GH transcripts during ripening, supports the notion that GH plays a central role in carotenogenesis in chromoplasts. However, GH mRNAs are also expressed in tissues and organs other than fruit. This is consistent with the possibility that GH mediates quinol oxidation from a variety of pathways that converge on plastoquinol in plastid membranes.

The Arabidopsis immutans mutation affects plastid differentiation and the morphogenesis of white and green sectors in variegated plants.

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