Publications (35) View all
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Article: Silencing the YB-1 Gene Inhibits Cell Migration in Gastric Cancer In Vitro.
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ABSTRACT: The Y-Box-Binding Protein-1 (YB-1) is known to regulate the processes of transcription, translation, cellular response to drug treatment and viral infection as well as DNA repair among others. As gastric cancer is a common cancer with a high incidence in countries in Asia, we evaluated the association of YB-1 with the malignant potential of gastric cancer cells in vitro. YB-1 mRNA expression levels were first determined by real-time RT-PCR in two adherent gastric cancer cell lines (viz., MKN7 and NUGC3 gastric cancer cells) and a normal GES-1 gastric epithelial cell line. Poorly differentiated NUGC3 gastric cancer cells were found to have the highest YB-1 gene expression among the adherent cells. YB-1 gene expression was also observed to be higher in non-adherent SNU5 gastric cancer cells compared to more aggressive SNU16 cells. Silencing of the YB-1 gene by siRNA in NUGC3 cells was associated with a significant reduction of the YB-1 protein by more than 55% as verified by Western blot analysis. Down-regulation of YB-1 protein expression was further demonstrated qualitatively by immunocytochemistry and immunofluorescence staining. Silencing of the YB-1 gene induced significant inhibition of cell migration in NUGC3 cells by 60% but did not influence cell invasion. Although epithelial-mesenchymal-transition (EMT) is known to be associated with the migratory phenotype in cancer cells, there was no change in the expression of EMT genes when YB-1 expression was modulated. YB-1 appears to have an integral role in cancer cell migration, a process which is important for gastric cancer metastasis. Anat Rec, 2013. © 2013 Wiley Periodicals, Inc.The Anatomical Record Advances in Integrative Anatomy and Evolutionary Biology 04/2013; · 1.47 Impact Factor -
Article: The NS1 protein of influenza A virus interacts with cellular processing bodies (P-bodies) and stress granules through RNA-associated protein 55 (RAP55) during virus infection.
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ABSTRACT: The non-structural protein (NS1) of influenza A virus exhibits multiple functions in the virus life cycle. Proteomics screening for cellular proteins which interact with NS1 identified the cellular protein, RAP55, which is one of the components of cellular processing bodies (P-bodies) and stress granules. To verify whether NS1 interacts with cellular P-bodies, interactions between NS1 and RAP55 and other P-body associated proteins (Ago1, Ago2 and DCP1a) were confirmed using co-immunoprecipitation and cellular co-localization assays. Over-expression of RAP55 induces RAP55-associated stress granule formation and suppresses virus replication. Knockdown of RAP55 with siRNA or expression of a dominant negative mutant RAP55 with defective interaction with P-bodies blocks NS1 co-localization to P-bodies in cells. Expression of NS1 inhibits RAP55 expression and formation of RAP55-associated P-bodies/stress granules. Viral nucleoprotein (NP) was found to be targeted to stress granules in the absence of NS1 but localized to P-bodies when NS1 was co-expressed. Restriction of virus replication via P-bodies occurs in the early phases of infection, as the numbers of RAP55-associated P-bodies in cells diminish over the course of virus infection. NS1 interaction with RAP55-associated P-bodies/stress granules is RNA binding associated and mediated via a PKR-interacting viral element. Mutations introduced into either RNA binding sites, namely R38/K41, or PKR interaction sites, namely I123/M124/K126/N127, cause NS1 proteins to lose the ability to interact with RAP55 and to inhibit stress granules. These results reveal an interplay between virus and host during virus replication in which NP is targeted to P-bodies/stress granules, while NS1 counteracts this host restriction mechanism.Journal of Virology 09/2012; · 5.40 Impact Factor -
Article: YB-1 Functions as a Porter To Lead Influenza Virus Ribonucleoprotein Complexes to Microtubules.
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ABSTRACT: De novo-synthesized RNAs are under the regulation of multiple posttranscriptional processes by a variety of RNA-binding proteins. The influenza virus genome consists of single-stranded RNAs and exists as viral ribonucleoprotein (vRNP) complexes. After the replication of vRNP in the nucleus, it is exported to the cytoplasm and then reaches the budding site beneath the cell surface in a process mediated by Rab11a-positive recycling endosomes along microtubules. However, the regulatory mechanisms of the postreplicational processes of vRNP are largely unknown. Here we identified, as a novel vRNP-interacting protein, Y-box-binding protein 1 (YB-1), a cellular protein that is involved in regulation of cellular transcription and translation. YB-1 translocated to the nucleus from the cytoplasm and accumulated in PML nuclear bodies in response to influenza virus infection. vRNP assembled into the exporting complexes with YB-1 at PML nuclear bodies. After nuclear export, using YB-1 knockdown cells and in vitro reconstituted systems, YB-1 was shown to be required for the interaction of vRNP exported from the nucleus with microtubules around the microtubule-organizing center (MTOC), where Rab11a-positive recycling endosomes were located. Further, we also found that YB-1 overexpression stimulates the production of progeny virions in an Rab11a-dependent manner. Taking these findings together, we propose that YB-1 is a porter that leads vRNP to microtubules from the nucleus and puts it into the vesicular trafficking system.Journal of Virology 08/2012; 86(20):11086-95. · 5.40 Impact Factor -
Article: PRMT1 is required for RAP55 to localize to processing bodies.
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ABSTRACT: In eukaryotic cells, components of messenger ribonucleoproteins (mRNPs) are often detected in cytoplasmic granules, such as processing bodies (P-bodies) and stress granules (SGs) where translationally repressed mRNAs accumulate. RAP55A, which is an RNA binding component of mRNPs, acts as a translational repressor and localizes to P-bodies and SGs. We found here that a homologous protein RAP55B also localized to P-bodies when expressed in human cultured cells. When RAP55A or RAP55B was highly expressed in the cells, they induced the formation of SG-like large cytoplasmic mRNP granules that contained both P-body and SG components, indicating that RAP55 is important for the assembly of cytoplasmic mRNP granules. In addition, we found that RAP55A associated with protein arginine methyltransferases PRMT1 and PRMT5. Multiple arginine residues of RAP55A were indeed asymmetrically dimethylated in the cell and PRMT1 was shown to be a component of large mRNP granules induced by RAP55A overexpression. Although PRMT1 did not accumulate in P-bodies, siRNA-mediated knockdown of PRMT1 impaired the localization of RAP55A to P-bodies, while other components were still retained in these structures. Thus, our data indicate that RAP55 is important for the assembly of cytoplasmic mRNP granules and that PRMT1 is required for RAP55A to localize to P-bodies.RNA biology 05/2012; 9(5):610-23. · 5.56 Impact Factor -
Article: PRAS40 is a functionally critical target for EWS repression in Ewing sarcoma.
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ABSTRACT: Ewing sarcoma family tumors (ESFT) are highly aggressive and highly metastatic tumors caused by a chromosomal fusion between the Ewing sarcoma protein (EWS) with the transcription factor FLI-1. However, expression of the EWS/FLI-1 chimeric oncogene by itself is insufficient for carcinogenesis, suggesting that additional events are required. Here, we report the identification of the Akt substrate PRAS40 as an EWS target gene. EWS negatively regulates PRAS40 expression by binding the 3' untranslated region in PRAS40 mRNA. ESFT cell proliferation was suppressed by treatment with an Akt inhibitor, and ESFT cell proliferation and metastatic growth were suppressed by siRNA-mediated PRAS40 knockdown. Furthermore, PRAS40 knockdown was sufficient to reverse an increased cell proliferation elicited by EWS knockdown. In support of a pathologic role for PRAS40 elevation in EFST, we documented inverse protein levels of EWS and PRAS40 in ESFT cells. Together, our findings suggest that PRAS40 promotes the development of ESFT and might therefore represent a novel therapeutic target in this aggressive disease.Cancer Research 03/2012; 72(5):1260-9. · 7.86 Impact Factor
