Core Signature Research Articles

Abstract PR12: Identifying different functional mechanisms of invasion from mesenchymal and proneural subtypes of glioblastoma

Abstract Glioblastoma multiforme (GBM) is the most frequent, aggressive and invasive type of primary brain tumors. Clinical management of patients with GBMs is confounded by the tumor cells' tendency to migrate and invade into adjacent brain. Understanding and targeting these invasive/dispersive malignant cells would address an unmet clinical need, and have a significant impact on patient quality of life and survival. The purpose of this study is to identify expression changes associated with distinct GBM subtypes that differentiate stationary core and invasive rim tumor cells. We used GBM invasion gene expression data (GSE12689), which consists of microdissected paired core v rim sets from 19 patient samples (Hoelzinger DB, et al Neoplasia. 2005. PMID: 1490313). Gene expression data on the 19 core samples were analyzed to determine specific GBM subtypes using the mesenchymal (MES) and proneural (PN) gene expression signatures (Verhaak RG et al Cancer Cell. 2010. PMID: 2818769.) resulting in 6 instances of MES, 7 PN, 3 uncertain-MES and 3 other. After subtype analysis, differentially expressed genes (DEGs) were identified between core and rim for the Mes and PN groups. Using a p-value threshold 0.005 (or p < 0.01), 216 (468) DEGs (core vs. rim) were identified from the MES-subtype, and 497 (1,028) DEGs were identified from the PN-subtype. The larger number of DEGs in the PN subtype suggests a more substantial (greater) adaptation of transcriptional change is necessary to transition to a invasive phenotype. The smaller number of changes in the MES samples suggests that cells of this form of GBM have an inherent migratory phenotype. Curiously, there was little gene overlap between MES and PN DEGs. A knowledge mining analysis was done to characterize biological concepts represented by the DEGs in each molecular subtype. The PN rim signature was enriched for genes involved in neuronal processes associated with synapse development and function. The PN core signature displayed genes associated with cell cycle progression. The MES core and rim signature represented a less homogeneous functional grouping of genes. There was a grouping of genes associated with vesicle movement, such as ARF1 and ARF3, associated with the MES rim. One gene of importance noted in the MES rim was autotaxin, which we have previously shown to be important in GBM invasion (Hoelzinger DB, et al. J Neurooncol. 2008 PMID: 17928955). No significant enriched biological concepts were evident in the MES core, however, individual genes of interest in the core such as the insulin receptor, BDNF and sonic hedgehog (SHH) were observed. Lastly, the ligand DLL4 and notch receptor (NOTCH1) are present, suggesting Notch signaling in the MES core. In summary, we analyzed a unique clinical GBM gene expression resource to identify patterns of differentially expressed genes associated with the invasive rim of MES and PN GBM samples. Molecular subtyping of the tumor has a marked influence on the changes in gene expression coincident to invasion. Disparity in the changes in patterns of differential gene expression between MES and PN GBM argues for wide-ranging adaptations necessary to affect dispersion in the brain, including tumor:microenvironment interactions, tumor cell adoption of a invasive phenoptype, and likely proliferative-, survival- and developmental-related changes. Such wide-ranging accommodation to succeed in invasion based on molecular subtyping raises opportunities by which to assess what the points of vulnerability may be in these malignant cells while they navigate through the brain to set up distant sites of tumor growth. (Supported by NIH CA U01CA168397) This abstract is also presented as Poster B92. Citation Format: Sungwon Jung, Seungchan Kim, Jeff Kiefer, Michael E. Berens. Identifying different functional mechanisms of invasion from mesenchymal and proneural subtypes of glioblastoma. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Invasion and Metastasis; Jan 20-23, 2013; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2013;73(3 Suppl):Abstract nr PR12.

Ca2+ Induces Spontaneous Dephosphorylation of a Novel P5A-type ATPase

P5 ATPases constitute the least studied group of P-type ATPases, an essential family of ion pumps in all kingdoms of life. Although P5 ATPases are present in every eukaryotic genome analyzed so far, they have remained orphan pumps, and their biochemical function is obscure. We show that a P5A ATPase from barley, HvP5A1, locates to the endoplasmic reticulum and is able to rescue knock-out mutants of P5A genes in both Arabidopsis thaliana and Saccharomyces cerevisiae. HvP5A1 spontaneously forms a phosphorylated reaction cycle intermediate at the catalytic residue Asp-488, whereas, among all plant nutrients tested, only Ca(2+) triggers dephosphorylation. Remarkably, Ca(2+)-induced dephosphorylation occurs at high apparent [Ca(2+)] (K(i) = 0.25 mM) and is independent of the phosphatase motif of the pump and the putative binding site for transported ligands located in M4. Taken together, our results rule out that Ca(2+) is a transported substrate but indicate the presence of a cytosolic low affinity Ca(2+)-binding site, which is conserved among P-type pumps and could be involved in pump regulation. Our work constitutes the first characterization of a P5 ATPase phosphoenzyme and points to Ca(2+) as a modifier of its function.

Sequence-Specific Binding of Recombinant Zbed4 to DNA: Insights into Zbed4 Participation in Gene Transcription and Its Association with Other Proteins

Zbed4, a member of the BED subclass of Zinc-finger proteins, is expressed in cone photoreceptors and glial Müller cells of human retina whereas it is only present in Müller cells of mouse retina. To characterize structural and functional properties of Zbed4, enough amounts of purified protein were needed. Thus, recombinant Zbed4 was expressed in E. coli and its refolding conditions optimized for the production of homogenous and functionally active protein. Zbed4’s secondary structure, determined by circular dichroism spectroscopy, showed that this protein contains 32% α-helices, 18% β-sheets, 20% turns and 30% unordered structures. CASTing was used to identify the target sites of Zbed4 in DNA. The majority of the DNA fragments obtained contained poly-Gs and some of them had, in addition, the core signature of GC boxes; a few clones had only GC-boxes. With electrophoretic mobility shift assays we demonstrated that Zbed4 binds both not only to DNA and but also to RNA oligonucleotides with very high affinity, interacting with poly-G tracts that have a minimum of 5 Gs; its binding to and GC-box consensus sequences. However, the latter binding depends on the GC-box flanking nucleotides. We also found that Zbed4 interacts in Y79 retinoblastoma cells with nuclear and cytoplasmic proteins Scaffold Attachment Factor B1 (SAFB1), estrogen receptor alpha (ERα), and cellular myosin 9 (MYH9), as shown with immunoprecipitation and mass spectrometry studies as well as gel overlay assays. In addition, immunostaining corroborated the co-localization of Zbed4 with these proteins. Most importantly, in vitro experiments using constructs containing promoters of genes directing expression of the luciferase gene, showed that Zbed4 transactivates the transcription of those promoters with poly-G tracts.

In silico identification of an epithelial core signature in human tumors.

10628 Background: Gene expression analysis is performed on grossly selected specimens often without any microscopic analysis of tumor content. In studies where histological analyses have been performed, cases having 80% or more tumor content are used for microarray analysis. The variability in amount of epithelial and stromal cells may generate to misleading differential expression analysis and selection for wrong targets for therapeutics. It is also often unclear, whether the genes identified are stromal or epithelial in origin. The goal of this study was to identify genes that define core epithelial phenotype; these genes could provide means of normalization of expression data. Methods: The CABIG GSK microarray (HG-U133_plus_2) data consisting of 950 cell lines from carcinoma (n=562), non-carcinoma (n=385) and normal tissue (n=3) was analyzed to identify epithelial specific genes. 10 carcinomas each from 11 sites (n=110) and an equal number of non-carcinomas were randomly selected. In silico analyses were performed by 1) identifying genes differentially expressed between carcinoma and non-carcinoma samples using a one way ANOVA; 2) identifying gene signature associated with carcinoma using Predictive Analysis of Microarrays (PAM) and 3) a weighted gene coexpression network analysis (WGCNA) was performed to identify co-expression modules. A similar analysis was also performed on tissue samples (E-GEOD-12360) from carcinomas and non-carcinomas. Venn-diagram was generated to identify intersecting set. Results: Comparison of the carcinoma and non-carcinoma samples using ANOVA identified 1455 differential expressed gene probes in cell lines and 540 gene probes in tissues (FDR=1E-10). The cell lines analysis identified 5 modules and a 65-gene signature (43 core and 22 accessory set) that was specific for epithelial cells. In the tissue analysis a 188-gene signature was similarly identified. Cross-comparison identified a smaller 31 gene intersecting set; this was not associated with loss of discriminatory power. Conclusions: A 31 geneset which can be used to determine the epithelial content of heterogeneous tumors, was identified. This study has the potential to significantly impact the use of microarray based gene expression data.

Cell-Type Independent MYC Target Genes Reveal a Primordial Signature Involved in Biomass Accumulation

The functions of key oncogenic transcription factors independent of context have not been fully delineated despite our richer understanding of the genetic alterations in human cancers. The MYC oncogene, which produces the Myc transcription factor, is frequently altered in human cancer and is a major regulatory hub for many cancers. In this regard, we sought to unravel the primordial signature of Myc function by using high-throughput genomic approaches to identify the cell-type independent core Myc target gene signature. Using a model of human B lymphoma cells bearing inducible MYC, we identified a stringent set of direct Myc target genes via chromatin immunoprecipitation (ChIP), global nuclear run-on assay, and changes in mRNA levels. We also identified direct Myc targets in human embryonic stem cells (ESCs). We further document that a Myc core signature (MCS) set of target genes is shared in mouse and human ESCs as well as in four other human cancer cell types. Remarkably, the expression of the MCS correlates with MYC expression in a cell-type independent manner across 8,129 microarray samples, which include 312 cell and tissue types. Furthermore, the expression of the MCS is elevated in vivo in Eμ-Myc transgenic murine lymphoma cells as compared with premalignant or normal B lymphocytes. Expression of the MCS in human B cell lymphomas, acute leukemia, lung cancers or Ewing sarcomas has the highest correlation with MYC expression. Annotation of this gene signature reveals Myc's primordial function in RNA processing, ribosome biogenesis and biomass accumulation as its key roles in cancer and stem cells.

Concomitant detection of IFNα signature and activated monocyte/dendritic cell precursors in the peripheral blood of IFNα-treated subjects at early times after repeated local cytokine treatments

BackgroundInterferons alpha (IFNα) are the cytokines most widely used in clinical medicine for the treatment of cancer and viral infections. Among the immunomodulatory activities possibly involved in their therapeutic efficacy, the importance of IFNα effects on dendritic cells (DC) differentiation and activation has been considered. Despite several studies exploiting microarray technology to characterize IFNα mechanisms of action, there is currently no consensus on the core signature of these cytokines in the peripheral blood of IFNα-treated individuals, as well as on the existence of blood genomic and proteomic markers of low-dose IFNα administered as a vaccine adjuvant.MethodsGene profiling analysis with microarray was performed on PBMC isolated from melanoma patients and healthy individuals 24 hours after each repeated injection of low-dose IFNα, administered as vaccine adjuvant in two separate clinical trials. At the same time points, cytofluorimetric analysis was performed on CD14+ monocytes, to detect the phenotypic modifications exerted by IFNα on antigen presenting cells precursors.ResultsAn IFNα signature was consistently observed in both clinical settings 24 hours after each repeated administration of the cytokine. The observed modulation was transient, and did not reach a steady state level refractory to further stimulations. The molecular signature observed ex vivo largely matched the one detected in CD14+ monocytes exposed in vitro to IFNα, including the induction of CXCL10 at the transcriptional and protein level. Interestingly, IFNα ex vivo signature was paralleled by an increase in the percentage and expression of costimulatory molecules by circulating CD14+/CD16+ monocytes, indicated as natural precursors of DC in response to danger signals.ConclusionsOur results provide new insights into the identification of a well defined molecular signature as biomarker of IFNα administered as immune adjuvants, and for the characterization of new molecular and cellular players, such as CXCL10 and CD14+/CD16+ cells, mediating and possibly predicting patient response to these cytokines.

Abstract 5168: Unraveling the molecular signature that drives breast cancer-induced brain metastasis in a mouse xenograft model

Abstract Brain metastases are most feared complication in breast cancer (BCa). Nearly 20% of patients with advanced BCa are eventually diagnosed with brain lesions, making breast tumors the main source of metastatic brain disease in women. Presently available treatment regimens are not capable of significantly treating the BCa-induced brain metastases due to their inability to penetrate the blood brain barrier. Rodent model systems have been reported for brain metastasis in BCa but the current models for brain metastasis have limitations. As such, there is a need of an efficient model system that can significantly contribute towards our understanding of different factors from both host and tumor leading to brain metastasis. Previously we isolated and characterized B6TC cells, derived from the stable spontaneous fusion of MDA-MB-231/GFP/Neo and ZR-75-1/GFP/puro in mouse bone marrow microenvironment. The low ER-expressing B6TC has propensity to metastasize to brain in addition to lung and bone, to express CD44 and ALDH, and to form mammospheres indicating its stem cell-like features. In this study using B6TC, we have developed an efficient and novel mouse model for studying BCa-induced brain metastasis. We performed RNA microarray to identify genes whose RNA levels are significantly altered in B6TC cells by greater than 2-fold at the false discovery rate of 0.001, when compared to the parental metastatic MDA-MB-231 or non-aggressive ZR-75-1 cells. Interestingly we identified genes like MMP1, HB-EGF, ST3GAL1, PTGS2, ITGA3, and CXCR4 implicated in metastasis regulation. Then we derived a Luc-GFP transfected B6TC line and enriched it by 4 successive in vivo passages to obtain a highly aggressive B6TC/Luc-GFP cell line with 100% brain metastatic frequency. A second round of RNA microarray was performed with highly brain metastatic B6TC/Luc-GFP and parental B6TC cells to derive a core signature of up- and down-regulated genes to select unique potential brain metastatic genes by comparing two sets of microarray data. Apart from potent brain metastatic genes detected earlier, we identified ZEB2, an EMT activator, a crucial promoter for metastasis, and a modulator of TGF beta signaling pathway, which shows up-regulation over successive in-vivo passages. Further analyses are underway to establish a refined signature of BCa-induced brain metastasis in our novel model system. Our study is significant in that they suggest potential brain metastasis genes for BCa. This will not only provide valuable insight into molecular mechanism of BCa-induced brain metastasis but also lay the foundation to identify novel prognostic and therapeutic markers of brain metastases, leading ultimately to the discovery of novel molecularly targeted drugs to prevent and eradicate BCa metastasis initiation, progression and recurrence. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 5168. doi:10.1158/1538-7445.AM2011-5168

Core epithelial-to-mesenchymal transition interactome gene-expression signature is associated with claudin-low and metaplastic breast cancer subtypes

The epithelial-to-mesenchymal transition (EMT) produces cancer cells that are invasive, migratory, and exhibit stem cell characteristics, hallmarks of cells that have the potential to generate metastases. Inducers of the EMT include several transcription factors (TFs), such as Goosecoid, Snail, and Twist, as well as the secreted TGF-beta1. Each of these factors is capable, on its own, of inducing an EMT in the human mammary epithelial (HMLE) cell line. However, the interactions between these regulators are poorly understood. Overexpression of each of the above EMT inducers up-regulates a subset of other EMT-inducing TFs, with Twist, Zeb1, Zeb2, TGF-beta1, and FOXC2 being commonly induced. Up-regulation of Slug and FOXC2 by either Snail or Twist does not depend on TGF-beta1 signaling. Gene expression signatures (GESs) derived by overexpressing EMT-inducing TFs reveal that the Twist GES and Snail GES are the most similar, although the Goosecoid GES is the least similar to the others. An EMT core signature was derived from the changes in gene expression shared by up-regulation of Gsc, Snail, Twist, and TGF-beta1 and by down-regulation of E-cadherin, loss of which can also trigger an EMT in certain cell types. The EMT core signature associates closely with the claudin-low and metaplastic breast cancer subtypes and correlates negatively with pathological complete response. Additionally, the expression level of FOXC1, another EMT inducer, correlates strongly with poor survival of breast cancer patients.

Repetitive Antigen Stimulation Induces Stepwise Transcriptome Diversification but Preserves a Core Signature of Memory CD8+ T Cell Differentiation

Repetitive antigen stimulation by prime-boost vaccination or pathogen reencounter increases memory CD8(+) T cell numbers, but the impact on memory CD8(+) T cell differentiation is unknown. Here we showed that repetitive antigen stimulations induced accumulation of memory CD8(+) T cells with uniform effector memory characteristics. However, genome-wide microarray analyses revealed that each additional antigen challenge resulted in the differential regulation of several hundred new genes in the ensuing memory CD8(+) T cell populations and, therefore, in stepwise diversification of CD8(+) T cell transcriptomes. Thus, primary and repeatedly stimulated (secondary, tertiary, and quaternary) memory CD8(+) T cells differed substantially in their molecular signature while sharing expression of a small group of genes and biological pathways, which may constitute a core signature of memory differentiation. These results reveal the complex regulation of memory CD8(+) T cell differentiation and identify potential new molecular targets to dissect the function of memory cells generated by repeated antigen stimulation.

DNA microarray profiling of genes differentially regulated by the histone deacetylase inhibitors vorinostat and LBH589 in colon cancer cell lines

BackgroundDespite the significant progress made in colon cancer chemotherapy, advanced disease remains largely incurable and novel efficacious chemotherapies are urgently needed. Histone deacetylase inhibitors (HDACi) represent a novel class of agents which have demonstrated promising preclinical activity and are undergoing clinical evaluation in colon cancer. The goal of this study was to identify genes in colon cancer cells that are differentially regulated by two clinically advanced hydroxamic acid HDACi, vorinostat and LBH589 to provide rationale for novel drug combination partners and identify a core set of HDACi-regulated genes.MethodsHCT116 and HT29 colon cancer cells were treated with LBH589 or vorinostat and growth inhibition, acetylation status and apoptosis were analyzed in response to treatment using MTS, Western blotting and flow cytometric analyses. In addition, gene expression was analyzed using the Illumina Human-6 V2 BeadChip array and Ingenuity® Pathway Analysis.ResultsTreatment with either vorinostat or LBH589 rapidly induced histone acetylation, cell cycle arrest and inhibited the growth of both HCT116 and HT29 cells. Bioinformatic analysis of the microarray profiling revealed significant similarity in the genes altered in expression following treatment with the two HDACi tested within each cell line. However, analysis of genes that were altered in expression in the HCT116 and HT29 cells revealed cell-line-specific responses to HDACi treatment. In addition a core cassette of 11 genes modulated by both vorinostat and LBH589 were identified in both colon cancer cell lines analyzed.ConclusionThis study identified HDACi-induced alterations in critical genes involved in nucleotide metabolism, angiogenesis, mitosis and cell survival which may represent potential intervention points for novel therapeutic combinations in colon cancer. This information will assist in the identification of novel pathways and targets that are modulated by HDACi, providing much-needed information on HDACi mechanism of action and providing rationale for novel drug combination partners. We identified a core signature of 11 genes which were modulated by both vorinostat and LBH589 in a similar manner in both cell lines. These core genes will assist in the development and validation of a common gene set which may represent a molecular signature of HDAC inhibition in colon cancer.

Tungsten isotopes as tracers of core–mantle interactions: The influence of subducted sediments

The postulated difference in W isotopic composition of the Earth’s core of ∼2 ε W units, compared to the bulk silicate earth (BSE) has previously been used to search for evidence of core–mantle interaction (CMI) in ocean island basalts (OIB). The absence of W isotope anomalies has thus been taken as evidence that CMI does not occur. However, the addition of subducted sediment with high W to the sources of OIB could obscure a core signature. This possibility brings into question the utility of W isotopes as tracers for CMI. To accurately consider the effects of sediment addition to mantle sources of OIB with respect to W requires improved constraints on the abundances of W in subducting sediment. Here, we present high-precision W abundance data (and other HFSE) for a suite of sediments from the Banda subduction regime in East Indonesia. Subducting East Indonesian sediments have trace element concentrations that resemble those of average upper continental crust (UCC), making these sediments valuable to consider as typical of subducted sediments. Average W abundances of 2.1 ppm, corrected for carbon content coupled with current models of 0.5% core addition and 1% sediment addition to EM1 or HIMU plume, suggest that a model hybrid source should exhibit values of ε W = −0.24 with ∼25 ppb W. Prior studies have not reported such low W isotopic compositions or high estimated W concentrations present in the sources of either Hawaiian or French Polynesian lavas, so such large additions of core material to these plume sources seems unlikely. Given these constraints, core contributions to these source, if present, can be no more than ∼0.1%.

Two independent gene signatures in pediatric t(4;11) acute lymphoblastic leukemia patients

Gene expression profiles become increasingly more important for diagnostic procedures, allowing clinical predictions including treatment response and outcome. However, the establishment of specific and robust gene signatures from microarray data sets requires the analysis of large numbers of patients and the application of complex biostatistical algorithms. Especially in case of rare diseases and due to these constrains, diagnostic centers with limited access to patients or bioinformatic resources are excluded from implementing these new technologies. In our study we sought to overcome these limitations and for proof of principle, we analyzed the rare t(4;11) leukemia disease entity. First, gene expression data of each t(4;11) leukemia patient were normalized by pairwise subtraction against normal bone marrow (n = 3) to identify significantly deregulated gene sets for each patient. A 'core signature' of 186 commonly deregulated genes present in each investigated t(4;11) leukemia patient was defined. Linking the obtained gene sets to four biological discriminators (HOXA gene expression, age at diagnosis, fusion gene transcripts and chromosomal breakpoints) divided patients into two distinct subgroups: the first one comprised infant patients with low HOXA genes expression and the MLL breakpoints within introns 11/12. The second one comprised non-infant patients with high HOXA expression and MLL breakpoints within introns 9/10. A yet homogeneous leukemia entity was further subdivided, based on distinct genetic properties. This approach provided a simplified way to obtain robust and disease-specific gene signatures even in smaller cohorts.

A Core MYC Gene Expression Signature Is Prominent in Basal-Like Breast Cancer but Only Partially Overlaps the Core Serum Response

BackgroundThe MYC oncogene contributes to induction and growth of many cancers but the full spectrum of the MYC transcriptional response remains unclear.Methodology/Principal FindingsUsing microarrays, we conducted a detailed kinetic study of genes that respond to MYCN or MYCNΔMBII induction in primary human fibroblasts. In parallel, we determined the response to steady state overexpression of MYCN and MYCNΔMBII in the same cell type. An overlapping set of 398 genes from the two protocols was designated a ‘Core MYC Signature’ and used for further analysis. Comparison of the Core MYC Signature to a published study of the genes induced by serum stimulation revealed that only 7.4% of the Core MYC Signature genes are in the Core Serum Response and display similar expression changes to both MYC and serum. Furthermore, more than 50% of the Core MYC Signature genes were not influenced by serum stimulation. In contrast, comparison to a panel of breast cancers revealed a strong concordance in gene expression between the Core MYC Signature and the basal-like breast tumor subtype, which is a subtype with poor prognosis. This concordance was supported by the higher average level of MYC expression in the same tumor samples.Conclusions/SignificanceThe Core MYC Signature has clinical relevance as this profile can be used to deduce an underlying genetic program that is likely to contribute to a clinical phenotype. Therefore, the presence of the Core MYC Signature may predict clinical responsiveness to therapeutics that are designed to disrupt MYC-mediated phenotypes.

Rethinking School Leadership Programs: The USC Ed.D. Program in Perspective

The Ed.D. program at the University of Southern California focuses on leadership in urban education. The article describes the rationale and guiding principles for the program and its design in terms of core knowledge, inquiry, signature pedagogy, concentrations, and capstone experiences. The article concludes with an assessment of our contributions to date and a set of challenges that will direct our efforts in the near future.

Identification of Cellular Pathways Mediating Progression of Monoclonal Gammopathy (MGUS) to Multiple Myeloma (MM) Using Gene Expression Profiling (GEP).

Events mediating transformation from pre-malignant MGUS to MM is currently not well defined. Recurrent genetic abnormalities such as t(11;14), t(4;14), hyperdiploidy and chromosome 13 deletion are already present in MGUS at relatively similar frequency to MM. The unified deregulation of D-type cyclins is also already present in MGUS. Previous GEP studies have revealed little differences between MGUS and MM. A more recent study using a large patient cohort and new generation Affymetrix genechip identifies an MGUS signature but the functional and biological significance underlying this signature is unknown. In the current study, we analyze a cohort of 22 MGUS and 101 MM from the Mayo Clinic with GEP performed on Affymetrix U133A genechip using gene set enrichment analysis, a method that analyze differentially expressed gene between 2 phenotypes of interest in the context of published or curated genesets that represent specific biological, chemical, or molecular perturbation to cells. This method increases the sensitivity of identifying low but significant changes in gene expression. We made further modification to the original method that allows assessment in individual samples rather than the average across a phenotype further increasing the specificity of the output. In this analysis, 313 genesets were significant enriched for genes over-expressed in MM compared to MGUS, representing potential activated pathways that mediate transformation. When MM samples and genesets were clustered using the enrichment score for each genesets and samples, 4 cluster of genesets emerged, one including a number of MYC genesets, one including a number of cell cycle related genesets, one including genesets related to metabolic activity and another including a number of IFN related genesets. Further dissection of these correlated genesets to identify common enriched genes (leading edge genes) led to identification of a MYC core signature, tRNA core signature, Proteosome core signature and metabolic core signature which are highly correlated. From known literature and biology, it is likely that MYC activation leads to downstream activation of protein synthesis (tRNA signature), degradation (proteasome signature), and metabolic pathway (metabolic signature). This is verified by GEP results from in vitro modulation of MYC activation in cell lines. In addition, a cell cycle core signature and IFN core signature was identified. Activation of IFN and MYC core signatures accounts for almost 90% of MM patients. The remaining patients have a metabolic signature without MYC or IFN activation. The activation of the different core signatures is significantly correlated with certain TC classes when assessed by Chi-Square test. IFN activation is significantly correlated with D1 subtypes and negatively correlated with D2 subtypes. MYC activation is negatively correlated with t(11;14). Similar patterns were observed in a validation dataset of 351 MM patients from UAMS (GSE2677) with GEP performed on the U133plus2.0 chip. These results are validated at the protein level using IHC on TMA of the Mayo cohort. The activation of the IFN and MYC pathway may represent predominant mechanism in MGUS to MM progression.

RXR heterodimerization allosterically activates LXR binding to the second NR box of activating signal co-integrator-2

ASC-2 (activating signal co-integrator-2) is a transcriptional co-activator that mediates the transactivation of NRs (nuclear receptors) via direct interactions with these receptors. ASC-2 contains two separate NR-interaction domains harbouring a core signature motif, LXXLL (where X is any amino acid), named the NR box. Although the first NR box (NR box-1) of ASC-2 interacts with many different NRs, the second NR box (NR box-2) specifically interacts with only LXR (liver X receptor), whose transactivation in vivo requires heterodimerization with RXR (retinoid X receptor). Interestingly, RXR has been shown to enhance the LXR transactivation, even in the absence of LXR ligand via a unique mechanism of allosteric regulation. In the present study we demonstrate that LXR binding to an ASC-2 fragment containing NR box-2 (Co4aN) is enhanced by RXR and even further by liganded RXR. We also identified specific residues in Co4aN involved in its interaction with LXR that were also required for the ASC-2-mediated transactivation of LXR in mammalian cells. Using these mutants, we found that the Co4aN-LXR interaction surface is not altered by the presence of RXR and RXR ligand and that the Ser(1490) residue is the critical determinant for the LXR-specific interaction of Co4aN. Notably the NR box-2, but not the NR box-1, is essential for ASC-2-mediated transactivation of LXR in vivo and for the interaction between LXR-RXR and ASC-2 in vitro. These results indicate that RXR does not interact directly with NR box-1 of ASC-2, but functions as an allosteric activator of LXR binding to NR box-2 of ASC-2.

Functional Network Analysis of the Transcriptomes of Mesenchymal Stem Cells Derived from Amniotic Fluid, Amniotic Membrane, Cord Blood, and Bone Marrow

Using high-density oligonucleotide microarrays and functional network analyses, we examined whether MSCs derived from four different origins exhibited unique gene expression profiles individually and then compared the gene expression profiles of all MSCs with those of fetal organs. Our results indicated that within each group of MSCs from the same origin, the variability of the gene expression levels was smaller than that between groups of different origins. Functional genomic studies revealed the specific roles of MSCs from different origins. Our results suggest that amniotic fluid MSCs may initiate interactions with the uterus by upregulating oxytocin and thrombin receptors. Amniotic membrane MSCs may play a role in maintaining homeostasis of fluid and electrolytes by regulating the networks of endothelin, neprilysin, bradykinin receptors, and atrial natriuretic peptide. Cord blood MSCs may be involved in innate immune systems as the neonatal defense system against the earliest encountered pathogens. Adult bone marrow MSCs may be an important source not only of all blood lineages but also of bone formation. However, in spite of the different gene expression profiles seen in MSCs derived from different origins, a set of core gene expression profiles was preserved in these four kinds of MSCs. The core signature transcriptomes of all MSCs, when contrasted against those of fetal organs, included genes involved in the regulation of extracellular matrix and adhesion, transforming growth factor-beta receptor signaling, and the Wnt signaling pathways. Disclosure of potential conflicts of interest is found at the end of this article.

A scheme for lunar inner core detection

Very precise measurements of the lunar gravity field could detect a solid inner core. For synchronous rotation, the equator planes of the inner core and mantle should be tilted with respect to the ecliptic plane and precess along that plane with the same 18.6 yr period. Generally, two different tilts would cause a static inner core gravity field to appear as small periodic (27.212 d) variations in the mantle‐referenced C21 and S21 coefficients. Tidal variations also contribute to the C21 variation so a much improved k2 Love number would be required. Model computations suggest that the inner core signature is likely to be very small requiring sensitive gravity measurements. In principle, a signature analogous to the Moon's should be present for other synchronous satellites with interior liquid layers and also Mercury.

All Memory Lymphocytes Share a Common Differentiation Program.

Although memory lymphocytes in CD4, CD8 and B cell lineages are characterized by a strikingly diverse array of functions and phenotypes, they share common properties of longevity, lowered response threshold and the ability to self-renew. The development of these memory functions is essential for immunity to pathogens and tumors, but it is not known how diverse lineages acquire these same properties. We therefore studied the transcriptional profile of memory differentiation to determine whether the development of memory in disparate lineages involves a common differentiation program. To identify a core signature of memory differentiation, we used cross-species genomic analysis to identify genes differentially expressed by both human and murine memory CD8 T cells compared to their naive precursors. We identified 220 genes significantly increased in human memory-phenotype cells, and found this signature to be highly conserved in two different TCR transgenic murine models of memory differentiation (P<0.001). Our analysis thus revealed an evolutionarily conserved transcriptional signature of CD memory differentiation. Several of the genes in this signature are known to be required for CD8 memory development, while others are novel markers of memory differentiation. We validated the signature at the protein and functional level, and demonstrated that it is highly enriched in the gene expression profiles of human CD8 T cells specific for CMV, influenza and EBV, suggesting that diverse viral pathogens can induce a similar differentiation program. We next tested whether this conserved signature is necessary for memory cells to be fully functional. To address this question, we studied memory differentiation in TCR transgenic T cells responding acute or chronic LCMV infection. Adult mice resolve an acute LCMV infection and then exhibit long-term CD8 T cell memory. In contrast, chronic LCMV infection results in exhausted CD8 cells that fail to demonstrate the properties of memory cells. Hierarchical clustering of the memory signature genes showed marked differences between functional memory cells and virally-exhausted T cells. Thus the signature of memory differentiation is disrupted in CD8 cells that fail to manifest the properties of memory. Finally, we tested the conserved CD8 memory signature in gene expression profiles of human memory CD4 and memory B cells compared to their naïve precursors. We found that the CD8 memory signature was highly enriched not only in CD4 memory (P<0.001) but also in B cell memory (P<0.001). Thus a single transcriptional signature is a general feature of memory differentiation in CD8, CD4 and B cells, and disruption of this signature is associated with defective T cell memory. Analysis of the regulation of this transcriptional program may help identify the mechanisms governing memory differentiation. Our data represent a novel approach to predicting the function of antigen-specific memory cells in vivo, and suggest that drugs which target this single differentiation program could profoundly influence the formation of immunologic memory.

Interaction between iron and post‐perovskite at core‐mantle boundary and core signature in plume source region

Interaction between the lower mantle and core is essential for understanding the nature of D″ layer at the core‐mantle boundary (CMB). Here, we report the reaction between post‐perovskite (PPv) and metallic iron under the condition of the CMB, for example, 139 GPa and 3000 Kelvin. Analytical transmission electron microscope (ATEM) analysis revealed that significant amount of oxygen up to 6.3 weight percent (wt.%) and silicon up to 4.0 wt.% can be dissolved into molten iron. The dihedral angle between PPv and molten iron is 67 degrees. Thus, a small amount of core metal of about 2 volume percent (vol.%) can be trapped without separation in the PPv region at the CMB. The amount of core metal trapped by this mechanism can produce the isotopic signature of the outer core in the plume source at the base of the lower mantle.