Pollen tubes extend through pistil tissues and are guided to ovules where they release sperm for fertilization. Although pollen tubes can germinate and elongate in a synthetic medium, their trajectory is random and their growth rates are slower compared to growth in pistil tissues. Furthermore, interaction with the pistil renders pollen tubes competent to respond to guidance cues secreted by specialized cells within the ovule. The molecular basis for this potentiation of the pollen tube by the pistil remains uncharacterized. Using microarray analysis in Arabidopsis, we show that pollen tubes that have grown through stigma and style tissues of a pistil have a distinct gene expression profile and express a substantially larger fraction of the Arabidopsis genome than pollen grains or pollen tubes grown in vitro. Genes involved in signal transduction, transcription, and pollen tube growth are overrepresented in the subset of the Arabidopsis genome that is enriched in pistil-interacted pollen tubes, suggesting the possibility of a regulatory network that orchestrates gene expression as pollen tubes migrate through the pistil. Reverse genetic analysis of genes induced during pollen tube growth identified seven that had not previously been implicated in pollen tube growth. Two genes are required for pollen tube navigation through the pistil, and five genes are required for optimal pollen tube elongation in vitro. Our studies form the foundation for functional genomic analysis of the interactions between the pollen tube and the pistil, which is an excellent system for elucidation of novel modes of cell–cell interaction.

Purpose Aurora A and B are oncogenic serine/threonine kinases that regulate mitosis. Over-expression of Auroras promotes resistance to microtubule targeted agents. We investigated mechanistic synergy by inhibiting the mitotic spindle apparatus in the presence of MLN8237 [M], an Aurora A inhibitor with either vincristine [MV] or docetaxel [MD] in aggressive B-NHL. The addition of rituximab [R] to MV or MD was evaluated for synthetic lethality. Experimental Design Aggressive B-NHL cell subtypes were evaluated in vitro and in vivo for target modulation and anti-NHL activity with single agents, doublets and triplets by analyzing cell proliferation, apoptosis, tumor growth, survival and mechanisms of response/relapse by gene expression profiling with protein validation. Results MV is synergistic while MD is additive for cell proliferation inhibition in B-NHL cell culture models. Addition of R to MV is superior to MD but both significantly induce apoptosis compared to doublet therapy. Mouse xenograft models of mantle cell lymphoma showed modest single agent activity for M, R, D and V with tumor growth inhibition (TGI) of ~10–15%. Of the doublets, MV caused tumor regression, while TGI was observed with MD (~55–60%) and MR (~25–50%) respectively. Although MV caused tumor regression, mice relapsed 20 days after stopping therapy. In contrast, MVR was curative, while MDR led to TGI of ~85%. PCNA, Aurora B, cyclin B1, cyclin D1 and Bcl-2 proteins of harvested tumors confirmed response and resistance to therapy. Conclusions Addition of R to MV is a novel therapeutic strategy for aggressive B-NHL and warrants clinical trial evaluation.

The detrimental effects of spaceflight and simulated microgravity on the immune system have been extensively documented. We report here microarray gene expression analysis, in concert with quantitative RT-PCR, in young adult C57BL/6NTac mice at 8 weeks of age after exposure to spaceflight aboard the space shuttle (STS-118) for a period of 13 days. Upon conclusion of the mission, thymus lobes were extracted from space flown mice (FLT) as well as age- and sex-matched ground control mice similarly housed in animal enclosure modules (AEM). mRNA was extracted and an automated array analysis for gene expression was performed. Examination of the microarray data revealed 970 individual probes that had a 1.5-fold or greater change. When these data were averaged (n = 4), we identified 12 genes that were significantly up- or down-regulated by at least 1.5-fold after spaceflight (P < or = 0.05). The genes that significantly differed from the AEM controls and that were also confirmed via QRT-PCR were as follows: Rbm3 (up-regulated) and Hsph110, Hsp90aa1, Cxcl10, Stip1, Fkbp4 (down-regulated). QRT-PCR confirmed the microarray results and demonstrated additional gene expression alteration in other T cell related genes, including: Ctla-4, IFN-alpha2a (up-regulated) and CD44 (down-regulated). Together, these data demonstrate that spaceflight induces significant changes in the thymic mRNA expression of genes that regulate stress, glucocorticoid receptor metabolism, and T cell signaling activity. These data explain, in part, the reported systemic compromise of the immune system after exposure to the microgravity of space.

Pearson correlation coefficient for expression analysis of the Lymphoma/Leukemia Molecular Profiling Project (LLMPP) demonstrated Aurora A and B are highly correlated with MYC in DLBCL and mantle cell lymphoma (MCL), while both Auroras correlate with BCL2 only in DLBCL. Auroras are up-regulated by MYC dysregulation with associated aneuploidy and resistance to microtubule targeted agents such as vincristine. Myc and Bcl2 are differentially expressed in U-2932, TMD-8, OCI-Ly10 and Granta-519, but only U-2932 cells over-express mutated p53. Alisertib [MLN8237 or M], a highly selective small molecule inhibitor of Aurora A kinase, was synergistic with vincristine [VCR] and rituximab [R] for inhibition of cell proliferation, abrogation of cell cycle checkpoints and enhanced apoptosis versus single agent or doublet therapy. A DLBCL (U-2932) mouse model showed tumor growth inhibition (TGI) of ∼10–20% (p = 0.001) for M, VCR and M-VCR respectively, while R alone showed ∼50% TGI (p = 0.001). M-R and VCR-R led to tumor regression [TR], but relapsed 10 days after discontinuing therapy. In contrast, M-VCR-R demonstrated TR with no relapse >40 days after stopping therapy with a Kaplan-Meier survival of 100%. Genes that are modulated by M-VCR-R (CENP-C, Auroras) play a role in centromere-kinetochore function in an attempt to maintain mitosis in the presence of synthetic lethality. Together, our data suggest that the interaction between alisertib plus VCR plus rituximab is synergistic and synthetic lethal in Myc and Bcl-2 co-expressing DLBCL. Alisertib plus vincristine plus rituximab [M-VCR-R] may represent a new strategy for DLBCL therapy.

Background Previous studies show gene expression alterations in rat embryo hearts and cell lines that correspond to the cardio-teratogenic effects of trichloroethylene (TCE) in animal models. One potential mechanism of TCE teratogenicity may be through altered regulation of calcium homeostatic genes with a corresponding inhibition of cardiac function. It has been suggested that TCE may interfere with the folic acid/methylation pathway in liver and kidney and alter gene regulation by epigenetic mechanisms. According to this hypothesis, folate supplementation in the maternal diet should counteract TCE effects on gene expression in the embryonic heart. Approach To identify transcriptional targets altered in the embryonic heart after exposure to TCE, and possible protective effects of folate, we used DNA microarray technology to profile gene expression in embryonic mouse hearts with maternal TCE exposure and dietary changes in maternal folate. Results Exposure to low doses of TCE (10ppb) caused extensive alterations in transcripts encoding proteins involved in transport, ion channel, transcription, differentiation, cytoskeleton, cell cycle and apoptosis. Exogenous folate did not offset the effects of TCE exposure on normal gene expression and both high and low levels of folate produced additional significant changes in gene expression. Conclusions A mechanism where TCE induces a folate deficiency does not explain altered gene expression patterns in the embryonic mouse heart. The data further suggest that use of folate supplementation, in the presence of this toxin, may be detrimental and non-protective of the developing embryo.

BackgroundNumerous microarray-based prognostic gene expression signatures of primary neoplasms have been published but often with little concurrence between studies, thus limiting their clinical utility. We describe a methodology using logistic regression, which circumvents limitations of conventional Kaplan Meier analysis. We applied this approach to a thrice-analyzed and published squamous cell carcinoma (SQCC) of the lung data set, with the objective of identifying gene expressions predictive of early death versus long survival in early-stage disease. A similar analysis was applied to a data set of triple negative breast carcinoma cases, which present similar clinical challenges.MethodsImportant to our approach is the selection of homogenous patient groups for comparison. In the lung study, we selected two groups (including only stages I and II), equal in size, of earliest deaths and longest survivors. Genes varying at least four-fold were tested by logistic regression for accuracy of prediction (area under a ROC plot). The gene list was refined by applying two sliding-window analyses and by validations using a leave–one-out approach and model building with validation subsets. In the breast study, a similar logistic regression analysis was used after selecting appropriate cases for comparison.ResultsA total of 8594 variable genes were tested for accuracy in predicting earliest deaths versus longest survivors in SQCC. After applying the two sliding window and the leave-one-out analyses, 24 prognostic genes were identified; most of them were B-cell related. When the same data set of stage I and II cases was analyzed using a conventional Kaplan Meier (KM) approach, we identified fewer immune-related genes among the most statistically significant hits; when stage III cases were included, most of the prognostic genes were missed. Interestingly, logistic regression analysis of the breast cancer data set identified many immune-related genes predictive of clinical outcome.ConclusionsStratification of cases based on clinical data, careful selection of two groups for comparison, and the application of logistic regression analysis substantially improved predictive accuracy in comparison to conventional KM approaches. B cell-related genes dominated the list of prognostic genes in early stage SQCC of the lung and triple negative breast cancer.

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Whole genome analyses were performed to test the hypothesis that temporal cortical gene expression differs between epilepsy patients rendered seizure-free versus non-seizure-free following anterior temporal lobectomy with amygdalohippocampectomy (ATL/AH). Twenty four patients underwent ATL/AH to treat medically intractable seizures of temporal lobe origin (mean age 35.5 years, mean follow up 42.2 months), they were then dichotomized into seizure-free and non-seizure-free groups. Tissue RNA was isolated from the lateral temporal cortex and gene expression analysis was performed. Whole genome data were analyzed for prognostic value for seizure-free outcome following ATL/AH by logistic regression. Genes that could distinguish seizure outcome groups were identified based on providing an accuracy of >0.90 judging by area under the receiver operating characteristic curve, AUC, with a P value of the slope coefficient of <0.05. Four genes and seven RNA probes were with prognostic value for post-operative seizure-free outcome. Gene expression associated with seizure-free outcome included relative down-regulation of: zinc finger protein 852 (ZNF852); CUB domain containing protein 2 (CDCP2); proline-rich transmembrane protein 1 (PRRT1); hypothetical LOC440200 (FLJ41170); RNA probe 8047763; RNA probe 8126238; RNA probe 8113489; RNA probe 8092883; RNA probe 7935228; RNA probe 806293 and RNA probe 8104131. This study describes the predictive value of temporal cortical gene expression for seizure-free outcome after ATL/AH. Four genes and seven RNA probes were found to predict post-operative seizure-free outcome. Future prospective investigation of these genes and probes in human brain tissue and blood could establish new biomarkers predictive of seizure outcome following ATL/AH.