Multidrug resistance (MDR) is a major clinical obstacle in treatment of gastric cancer. Previously, using 2D electrophoresis-mass spectrometry, we identified RhoGDI2 as a contributor to 5-FU resistance in colon cancer cells, and also confer gastric cancer cells resistance to 5-FU. Here, we reported RhoGDI2 also induced MDR in gastric cancer cell line (MKN-45). To explore the underlining mechanism, we detected the mRNA, protein expression, activity of P-glycoprotein (P-gp) in MKN-45 stably transfected with RhoGDI2 expressing or control vector. All the mRNA, protein level, activity were increased by 130%, 230%, 35% respectively after ectopic expression of RhoGDI2. RhoGDI2 was correlated with P-gp expression in gastric cancer tissues as detected by immunohistochemistry. To further study how RhoGDI2 up-regulates P-gp expression, we tested the activity of Rac1 in MKN-45/RhoGDI2 and MKN-45/GFP. Ectopic expression of RhoGDI2 increased Rac1 activity (P < 0.05). For more important, silencing of Rac1 expression by siRNA decreased P-gp expression to undetectable level. Overall, these findings suggest that RhoGDI2 up-regulates P-gp expression via Rac1 to induce MDR.

Genetic and transcriptional profiling, as well as surface marker identification of single circulating tumor cells (CTCs) have been demonstrated. However, quantitatively profiling of functional proteins at single CTC resolution has not yet been achieved, owing to the limited purity of the isolated CTC populations and a lack of single-cell proteomic approaches to handle and analyze rare CTCs. Here, we develop an integrated microfluidic system specifically designed for streamlining isolation, purification and single-cell secretomic profiling of CTCs from whole blood. Key to this platform is the use of photocleavable ssDNA-encoded antibody conjugates to enable a highly purified CTC population with <75 'contaminated' blood cells. An enhanced poly-L-lysine barcode pattern is created on the single-cell barcode chip for efficient capture rare CTC cells in microchambers for subsequent secreted protein profiling. This system was extensively evaluated and optimized with EpCAM-positive HCT116 cells seeded into whole blood. Patient blood samples were employed to assess the utility of the system for isolation, purification and single-cell secretion profiling of CTCs. The CTCs present in patient blood samples exhibit highly heterogeneous secretion profile of IL-8 and VEGF. The numbers of secreting CTCs are found not in accordance with CTC enumeration based on immunostaining in the parallel experiments.

Overexpressed CEACAM6 in tumor tissues plays important roles in invasion, metastasis and anoikis resistance in a variety of human cancers. We recently reported that CEACAM6 expression is upregulated in Gastric cancer (GC) tissues and promoted GC metastasis. Here, we report that CEACAM6 promotes peritoneal metastases in vivo and is negatively correlated with E-cadherin expression in GC tissues. Overexpressed CEACAM6 induced epithelial-mesenchymal transition (EMT) in GC, as measured by increases in the EMT markers N-cadherin, Vimentin and Slug while E-cadherin expression was decreased in CEACAM6-overexpressing GC cells; opposing results were observed in CEACAM6-silenced cells. Furthermore, E-cadherin expression was negatively correlated with depth of tumor invasion, lymph node metastasis and TNM stage in GC tissues. Additionally, CEACAM6 elevated matrix metalloproteinase-9 (MMP-9) activity in GC, and anti-MMP-9 antibody could reverse the increasing invasion and migration induced by CEACAM6. CEACAM6 also increased the levels of phosphorylated AKT, which is involved in the progression of a variety of human tumors. We further observed that LY294002, a PI3K inhibitor, could reverse CEACAM6-induced EMT via mesenchymal-epithelial transition. These findings suggest that CEACAM6 enhances invasion and metastasis in GC by promoting EMT via the PI3K/AKT signaling pathway.

Androgen receptor (AR) plays an important role in many kinds of cancers. However, the molecular mechanisms of AR in gastric cancer (GC) are poorly characterized. Here, we investigated the role of AR in GC cell migration, invasion and metastatic potential. Our data showed that AR expression was positively correlated with lymph node metastasis and late TNM stages. These findings were accompanied by activation of AKT and upregulation of matrix metalloproteinase 9 (MMP9). AR overexpression induced increases in GC cell migration, invasion and proliferation in vitro and in vivo. These effects were attenuated by inhibition of AKT, AR and MMP9. AR overexpression upregulated MMP9 protein levels, whereas this effect was counteracted by AR siRNA. Inhibition of AKT by siRNA or an inhibitor (MK-2206 2HC) decreased AR protein expression in both stably transfected and parental SGC-7901 cells. Luciferase reporter and chromatin immunoprecipitation assays demonstrated that AR bound to the AR-binding sites of the MMP9 promoter. In summary, AR overexpression induced by AKT phosphorylation upregulated MMP9 by binding to its promoter region to promote gastric carcinogenesis. The AKT/AR/MMP9 pathway plays an important role in GC metastasis and may be a novel therapeutic target for GC treatment.

Elevated MELK expression is featured in multiple tumors and correlated with tumorigenesis and tumor development. This study is aimed to investigate the mechanisms of MELK-mediated development of gastric cancer.

The data presented in this article is related to the research article entitled "Biochemical Fractionation of Time-Resolved Drosophila Embryos Reveals Similar Transcriptomic Alterations in Replication Checkpoint and Histone mRNA Processing Mutants" (Lefebvre et al., 2017) [1]. This article provides a spatiotemporal transcriptomic analysis of early embryogenesis and shows that mutations in the checkpoint factor grapes/Chk1 and the histone mRNA processing factor SLBP selectively impair zygotic gene expression. Here, lists of transcripts enriched in early syncytial embryos, late blastoderm embryos, cytoplasmic and nuclear extracts of blastoderm embryos are made public, along with transcription factor motif occurrence for genes enriched in each context. In addition, extensive lists of genes down-regulated upon Chk1 and SLBP protein depletion in embryos are released to enable further analyses.

Gastric cancer is the second most common cause of global cancer-related mortality. Although dedifferentiation predicts poor prognosis in gastric cancer, the molecular mechanism underlying dedifferentiation, which could provide fundamental insights into tumor development and progression, has yet to be elucidated. Furthermore, the molecular mechanism underlying the effects of hexamethylene bisacetamide (HMBA), a recently discovered differentiation inducer, requires investigation and there are no reported studies concerning the effect of HMBA on gastric cancer.

MicroRNA has been recently recognized as playing a prominent role in tumorigenesis and metastasis. Here, we report that miR-338-3p was epigenetically silenced in gastric cancer, and its down-regulation was significantly correlated with gastric cancer clinicopathological features. Strikingly, restoring miR-338-3p expression in SGC-7901 gastric cancer cells inhibited proliferation, migration, invasion and tumorigenicity in vitro and in vivo, at least partly through inducing apoptosis. Furthermore, we demonstrate the oncogene SSX2IP is a target of miR-338-3p. We propose that miR-338-3p functions as a tumor suppressor in gastric cancer, and the methylation status of its CpG island could serve as a potential diagnostic marker for gastric cancer.

MTA2 gene belongs to metastasis associated family, and is highly expressed in some solid tumors, including gastric cancer. Its biological function in gastric cancer is currently undefined.

Vertebrates require tremendous molecular diversity to defend against numerous small hydrophobic chemicals. UDP-glucuronosyltransferases (UGTs) are a large family of detoxification enzymes that glucuronidate xenobiotics and endobiotics, facilitating their excretion from the body. The UGT1 gene cluster contains a tandem array of variable first exons, each preceded by a specific promoter, and a common set of downstream constant exons, similar to the genomic organization of the protocadherin (Pcdh), immunoglobulin, and T-cell receptor gene clusters. To assist pharmacogenomics studies in Chinese, we sequenced nine first exons, promoter and intronic regions, and five common exons of the UGT1 gene cluster in a population sample of 253 unrelated Chinese individuals. We identified 101 polymorphisms and found 15 novel SNPs. We then computed allele frequencies for each polymorphism and reconstructed their linkage disequilibrium (LD) map. The UGT1 cluster can be divided into five linkage blocks: Block 9 (UGT1A9), Block 9/7/6 (UGT1A9, UGT1A7, and UGT1A6), Block 5 (UGT1A5), Block 4/3 (UGT1A4 and UGT1A3), and Block 3' UTR. Furthermore, we inferred haplotypes and selected their tagSNPs. Finally, comparing our data with those of three other populations of the HapMap project revealed ethnic specificity of the UGT1 genetic diversity in Chinese. These findings have important implications for future molecular genetic studies of the UGT1 gene cluster as well as for personalized medical therapies in Chinese.

Although subcellular mRNA trafficking has been demonstrated as a mechanism to control protein distribution, it is generally believed that most protein localization occurs subsequent to translation. To address this point, we developed and employed a high-resolution fluorescent in situ hybridization procedure to comprehensively evaluate mRNA localization dynamics during early Drosophila embryogenesis. Surprisingly, of the 3370 genes analyzed, 71% of those expressed encode subcellularly localized mRNAs. Dozens of new and striking localization patterns were observed, implying an equivalent variety of localization mechanisms. Tight correlations between mRNA distribution and subsequent protein localization and function, indicate major roles for mRNA localization in nucleating localized cellular machineries. A searchable web resource documenting mRNA expression and localization dynamics has been established and will serve as an invaluable tool for dissecting localization mechanisms and for predicting gene functions and interactions.

Stress granules (SGs) are transient ribonucleoprotein (RNP) aggregates that form during cellular stress and are increasingly implicated in human neurodegeneration. To study the proteome and compositional diversity of SGs in different cell types and in the context of neurodegeneration-linked mutations, we used ascorbate peroxidase (APEX) proximity labeling, mass spectrometry, and immunofluorescence to identify ∼150 previously unknown human SG components. A highly integrated, pre-existing SG protein interaction network in unstressed cells facilitates rapid coalescence into larger SGs. Approximately 20% of SG diversity is stress or cell-type dependent, with neuronal SGs displaying a particularly complex repertoire of proteins enriched in chaperones and autophagy factors. Strengthening the link between SGs and neurodegeneration, we demonstrate aberrant dynamics, composition, and subcellular distribution of SGs in cells from amyotrophic lateral sclerosis (ALS) patients. Using three Drosophila ALS/FTD models, we identify SG-associated modifiers of neurotoxicity in vivo. Altogether, our results highlight SG proteins as central to understanding and ultimately targeting neurodegeneration.

Gastric cancer is one of the most common malignancies worldwide. Recent studies have shown that microRNAs play crucial roles in regulating cellular proliferation process in gastric cancer. MicroRNA-29c (miR-29c) acts as a tumor suppressor in different kinds of tumors.

In higher eukaryotes, maternally provided gene products drive the initial stages of embryogenesis until the zygotic transcriptional program takes over, a developmental process called the midblastula transition (MBT). In addition to zygotic genome activation, the MBT involves alterations in cell-cycle length and the implementation of DNA damage/replication checkpoints that serve to monitor genome integrity. Previous work has shown that mutations affecting histone mRNA metabolism or DNA replication checkpoint factors severely impact developmental progression through the MBT, prompting us to characterize and contrast the transcriptomic impact of these genetic perturbations. In this study, we define gene expression profiles that mark early embryogenesis in Drosophila through transcriptomic analyses of developmentally staged (early syncytial versus late blastoderm) and biochemically fractionated (nuclear versus cytoplasmic) wild-type (wt) embryos. We then compare the transcriptomic profiles of loss-of-function mutants of the Chk1/Grapes replication checkpoint kinase and the stem loop binding protein (SLBP), a key regulator of replication-dependent histone mRNAs. Our analysis of RNA spatial and temporal distribution during embryogenesis offers new insights into the dynamics of early embryogenesis. In addition, we find that grp and Slbp mutant embryos display profound and highly similar defects in gene expression, most strikingly in zygotic gene expression, compromising the transition from a maternal to a zygotic regulation of development.

Gastric cancer is the third leading cause of cancer-related mortality worldwide. Chemotherapy is frequently used for gastric cancer treatment. Most patients with advanced gastric cancer eventually succumb to the disease despite some patients responded initially to chemotherapy. Thus, identifying molecular mechanisms responsible for cancer relapse following chemotherapy will help design new ways to treat gastric cancer. In this study, we revealed that the residual cancer cells following treatment with chemotherapeutic reagent cisplatin have elevated expression of hedgehog target genes GLI1, GLI2 and PTCH1, suggestive of hedgehog signaling activation. We showed that GLI1 knockdown sensitized gastric cancer cells to CDDP whereas ectopic GLI1 expression decreased the sensitivity. Further analyses indicate elevated GLI1 expression is associated with an increase in tumor sphere formation, side population and cell surface markers for putative cancer stem cells. We have evidence to support that GLI1 is critical for maintenance of putative cancer stem cells through direct regulation of ABCG2. In fact, GLI1 protein was shown to be associated with the promoter fragment of ABCG2 through a Gli-binding consensus site in gastric cancer cells. Disruption of ABCG2 function, through ectopic expression of an ABCG2 dominant negative construct or a specific ABCG2 inhibitor, increased drug sensitivity of cancer cells both in culture and in mice. The relevance of our studies to gastric cancer patient care is reflected by our discovery that high ABCG2 expression was associated with poor survival in the gastric cancer patients who underwent chemotherapy. Taken together, we have identified a molecular mechanism by which gastric cancer cells gain chemotherapy resistance.

The human and mouse genomes contain instructions that specify RNAs and proteins and govern the timing, magnitude, and cellular context of their production. To better delineate these elements, phase III of the Encyclopedia of DNA Elements (ENCODE) Project has expanded analysis of the cell and tissue repertoires of RNA transcription, chromatin structure and modification, DNA methylation, chromatin looping, and occupancy by transcription factors and RNA-binding proteins. Here we summarize these efforts, which have produced 5,992 new experimental datasets, including systematic determinations across mouse fetal development. All data are available through the ENCODE data portal (https://www.encodeproject.org), including phase II ENCODE1 and Roadmap Epigenomics2 data. We have developed a registry of 926,535 human and 339,815 mouse candidate cis-regulatory elements, covering 7.9 and 3.4% of their respective genomes, by integrating selected datatypes associated with gene regulation, and constructed a web-based server (SCREEN; http://screen.encodeproject.org) to provide flexible, user-defined access to this resource. Collectively, the ENCODE data and registry provide an expansive resource for the scientific community to build a better understanding of the organization and function of the human and mouse genomes.

Increasing evidence suggests that transcriptional control and chromatin activities at large involve regulatory RNAs, which likely enlist specific RNA-binding proteins (RBPs). Although multiple RBPs have been implicated in transcription control, it has remained unclear how extensively RBPs directly act on chromatin. We embarked on a large-scale RBP ChIP-seq analysis, revealing widespread RBP presence in active chromatin regions in the human genome. Like transcription factors (TFs), RBPs also show strong preference for hotspots in the genome, particularly gene promoters, where their association is frequently linked to transcriptional output. Unsupervised clustering reveals extensive co-association between TFs and RBPs, as exemplified by YY1, a known RNA-dependent TF, and RBM25, an RBP involved in splicing regulation. Remarkably, RBM25 depletion attenuates all YY1-dependent activities, including chromatin binding, DNA looping, and transcription. We propose that various RBPs may enhance network interaction through harnessing regulatory RNAs to control transcription.

Background: Fluorouracil-based chemotherapy is recommended by the main clinical guidelines for post-operative gastric cancer (GC) patient's chemotherapy treatment, this study aim to establish relate model to predict patients' susceptibility to fluorouracil-based chemotherapy to prevent patients' unnecessary exposure to chemotherapy treatments and improve patients' treatment. Methods: Data from Gene Expression Omnibus (GEO) database, Cancer Cell Line Encyclopedia (CCLE) database, Cancer Therapeutics Response Portal (CTRP) and The Cancer Genome Atlas (TCGA) were used. A predictive model was built based on univariate and multivariate Cox analysis and visualized by nomogram. Survival analysis was performed using Kaplan-Meier and log-rank test. Results: A total of 514 differentially expressed genes (DEGs) were identified between fluorouracil-resistant cell lines and fluorouracil-sensitive cell lines based on CCLE database. A total of 300 patients who had radical gastrectomy were recruited, of which 144 received fluorouracil-based chemotherapy and 156 were untreated. Three biomarkers (CTF1, BTN3A3, ADAD2) were finally selected by univariate and multivariate Cox regression analysis to establish the predictive models visualized by nomogram. This model could precisely predict both the Disease free survival (DFS) and Overall survival (OS) of patients treated with fluorouracil-based chemotherapy after surgery compared to untreated GC patients validated by both GEO database and TCGA database. Conclusion: Our data established three genes-based predictive model which might predict GC patients' susceptibility to fluorouracil and help clinicians develop personalized treatment.

RNA binding proteins (RBPs) are central regulators of gene expression implicated in all facets of RNA metabolism. As such, they play key roles in cellular physiology and disease etiology. Since different steps of post-transcriptional gene expression tend to occur in specific regions of the cell, including nuclear or cytoplasmic locations, defining the subcellular distribution properties of RBPs is an important step in assessing their potential functions. Here, we present the RBP Image Database, a resource that details the subcellular localization features of 301 RBPs in the human HepG2 and HeLa cell lines, based on the results of systematic immuno-fluorescence studies conducted using a highly validated collection of RBP antibodies and a panel of 12 markers for specific organelles and subcellular structures. The unique features of the RBP Image Database include: (i) hosting of comprehensive representative images for each RBP-marker pair, with ∼250,000 microscopy images; (ii) a manually curated controlled vocabulary of annotation terms detailing the localization features of each factor; and (iii) a user-friendly interface allowing the rapid querying of the data by target or annotation. The RBP Image Database is freely available at https://rnabiology.ircm.qc.ca/RBPImage/.

Vacuolar protein sorting-associated protein 35 (VPS35) is a core component of the retromer complex which mediates intracellular protein transport. It is well known that dysfunctional VPS35 functions in the accumulation of pathogenic proteins. In our previous study, VPS35 was found to be a potential gene related to poor prognosis in gastric cancer. However, the biological functions of VPS35 in gastric cancer remain unclear.