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short hairpin rna shrna targeting c myc  (Addgene inc)


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    Addgene inc short hairpin rna shrna targeting c myc
    KSHV dysregulated NSUN2/1 expression via <t>c-Myc.</t> ( A and B ) Transcription factors (TFs) that regulate NSUN2/1 gene expression were predicted by analyzing public-domain ChIP-seq datasets. Venn diagram (A) showed the overlapped TFs across the indicated datasets. Six TF candidates including c-Myc (B) were listed. ( C ) Integrative genomics viewer (IGV) visualization of c-Myc occupancy near the NSUN2/1 promoter regions using the public-domain c-Myc ChIP-seq dataset of five lymphoma B cell lines ( GSE30726 ). ( D ) c-Myc binding motif near the promoter regions of NSUN2/1 was illustrated. ( E ) TREx.BCBL1.Rta cells were subjected to ChIP-PCR analysis for quantification of c-Myc association with the promoter regions of NSUN2/1 using an antibody recognizing c-Myc protein for its immunoprecipitation or a control IgG antibody, followed by qPCR analysis using three sets of primers (Set 1–3) targeting NSUN2 or NSUN1 promoter. ( F ) Public-domain <t>RNA-seq</t> data of KSHV-infected cell lines were collected and reanalyzed using the customized pipeline to identify the differentially expressed genes (adjust P -value < 0.05 as cutoff). The distinct gene expression level of c-Myc due to KSHV lytic reactivation was illustrated. ( G ) TREx.BCBL1.Rta, iSLK.BAC16, and iSLK.r219 cells were treated with Dox or mock, followed by protein immunoblotting analysis of c-Myc using its specific antibody. GAPDH was used as a loading control. ( H ) TREx.BCBL1.Rta cells were transduced with the <t>shRNA</t> targeting c-Myc or non-targeting control, followed by protein immunoblotting analysis of c-Myc, NSUN2, NSUN1, and KSHV K8.1. ( I and J ) TREx.BCBL1.Rta cells were treated with c-Myc inhibitors, EN4 (H) or 10074-G5 (I), at a series of concentrations or mock, followed by protein immunoblotting analysis of c-Myc, NSUN2, and NSUN1 using their specific antibodies. ( K ) TIME cells were transiently transfected with siRNAs (si1, si2) targeting c-Myc or siNT, followed by inoculation with KSHV.BAC16 viruses. These cells were harvested for nuclei staining with DAPI. GFP fluorescence signal indicating KSHV-infected cells was quantified (**** P <0.0001). ( L ) TIME cells were treated with the c-Myc inhibitor EN4 at the indicated concentrations, followed by transfection of NSUN1 or NSUN2 cDNAs. These cells were inoculated with KSHV.BAC16 viruses and lysed for protein immunoblotting assays of KSHV K8.1 protein.
    Short Hairpin Rna Shrna Targeting C Myc, supplied by Addgene inc, used in various techniques. Bioz Stars score: 93/100, based on 18 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 93 stars, based on 18 article reviews
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    Images

    1) Product Images from "m5C RNA methylation is dysregulated by oncogenic herpesviruses via c-Myc signaling to counteract host antiviral factors"

    Article Title: m5C RNA methylation is dysregulated by oncogenic herpesviruses via c-Myc signaling to counteract host antiviral factors

    Journal: Nucleic Acids Research

    doi: 10.1093/nar/gkag251

    KSHV dysregulated NSUN2/1 expression via c-Myc. ( A and B ) Transcription factors (TFs) that regulate NSUN2/1 gene expression were predicted by analyzing public-domain ChIP-seq datasets. Venn diagram (A) showed the overlapped TFs across the indicated datasets. Six TF candidates including c-Myc (B) were listed. ( C ) Integrative genomics viewer (IGV) visualization of c-Myc occupancy near the NSUN2/1 promoter regions using the public-domain c-Myc ChIP-seq dataset of five lymphoma B cell lines ( GSE30726 ). ( D ) c-Myc binding motif near the promoter regions of NSUN2/1 was illustrated. ( E ) TREx.BCBL1.Rta cells were subjected to ChIP-PCR analysis for quantification of c-Myc association with the promoter regions of NSUN2/1 using an antibody recognizing c-Myc protein for its immunoprecipitation or a control IgG antibody, followed by qPCR analysis using three sets of primers (Set 1–3) targeting NSUN2 or NSUN1 promoter. ( F ) Public-domain RNA-seq data of KSHV-infected cell lines were collected and reanalyzed using the customized pipeline to identify the differentially expressed genes (adjust P -value < 0.05 as cutoff). The distinct gene expression level of c-Myc due to KSHV lytic reactivation was illustrated. ( G ) TREx.BCBL1.Rta, iSLK.BAC16, and iSLK.r219 cells were treated with Dox or mock, followed by protein immunoblotting analysis of c-Myc using its specific antibody. GAPDH was used as a loading control. ( H ) TREx.BCBL1.Rta cells were transduced with the shRNA targeting c-Myc or non-targeting control, followed by protein immunoblotting analysis of c-Myc, NSUN2, NSUN1, and KSHV K8.1. ( I and J ) TREx.BCBL1.Rta cells were treated with c-Myc inhibitors, EN4 (H) or 10074-G5 (I), at a series of concentrations or mock, followed by protein immunoblotting analysis of c-Myc, NSUN2, and NSUN1 using their specific antibodies. ( K ) TIME cells were transiently transfected with siRNAs (si1, si2) targeting c-Myc or siNT, followed by inoculation with KSHV.BAC16 viruses. These cells were harvested for nuclei staining with DAPI. GFP fluorescence signal indicating KSHV-infected cells was quantified (**** P <0.0001). ( L ) TIME cells were treated with the c-Myc inhibitor EN4 at the indicated concentrations, followed by transfection of NSUN1 or NSUN2 cDNAs. These cells were inoculated with KSHV.BAC16 viruses and lysed for protein immunoblotting assays of KSHV K8.1 protein.
    Figure Legend Snippet: KSHV dysregulated NSUN2/1 expression via c-Myc. ( A and B ) Transcription factors (TFs) that regulate NSUN2/1 gene expression were predicted by analyzing public-domain ChIP-seq datasets. Venn diagram (A) showed the overlapped TFs across the indicated datasets. Six TF candidates including c-Myc (B) were listed. ( C ) Integrative genomics viewer (IGV) visualization of c-Myc occupancy near the NSUN2/1 promoter regions using the public-domain c-Myc ChIP-seq dataset of five lymphoma B cell lines ( GSE30726 ). ( D ) c-Myc binding motif near the promoter regions of NSUN2/1 was illustrated. ( E ) TREx.BCBL1.Rta cells were subjected to ChIP-PCR analysis for quantification of c-Myc association with the promoter regions of NSUN2/1 using an antibody recognizing c-Myc protein for its immunoprecipitation or a control IgG antibody, followed by qPCR analysis using three sets of primers (Set 1–3) targeting NSUN2 or NSUN1 promoter. ( F ) Public-domain RNA-seq data of KSHV-infected cell lines were collected and reanalyzed using the customized pipeline to identify the differentially expressed genes (adjust P -value < 0.05 as cutoff). The distinct gene expression level of c-Myc due to KSHV lytic reactivation was illustrated. ( G ) TREx.BCBL1.Rta, iSLK.BAC16, and iSLK.r219 cells were treated with Dox or mock, followed by protein immunoblotting analysis of c-Myc using its specific antibody. GAPDH was used as a loading control. ( H ) TREx.BCBL1.Rta cells were transduced with the shRNA targeting c-Myc or non-targeting control, followed by protein immunoblotting analysis of c-Myc, NSUN2, NSUN1, and KSHV K8.1. ( I and J ) TREx.BCBL1.Rta cells were treated with c-Myc inhibitors, EN4 (H) or 10074-G5 (I), at a series of concentrations or mock, followed by protein immunoblotting analysis of c-Myc, NSUN2, and NSUN1 using their specific antibodies. ( K ) TIME cells were transiently transfected with siRNAs (si1, si2) targeting c-Myc or siNT, followed by inoculation with KSHV.BAC16 viruses. These cells were harvested for nuclei staining with DAPI. GFP fluorescence signal indicating KSHV-infected cells was quantified (**** P <0.0001). ( L ) TIME cells were treated with the c-Myc inhibitor EN4 at the indicated concentrations, followed by transfection of NSUN1 or NSUN2 cDNAs. These cells were inoculated with KSHV.BAC16 viruses and lysed for protein immunoblotting assays of KSHV K8.1 protein.

    Techniques Used: Expressing, Gene Expression, ChIP-sequencing, Binding Assay, Immunoprecipitation, Control, RNA Sequencing, Infection, Western Blot, Transduction, shRNA, Transfection, Staining, Fluorescence

    EBV also reduced expression of NSUN2/1 that restrict its lytic infection. ( A ) BJAB cells were inoculated with EBV.BX viruses. Cell lysates were collected at 48 h post infection and were followed by protein immunoblotting analysis using specific antibodies recognizing NSUN2 and NSUN1. ( B ) Akata and Akata BX cells were treated with a human IgG antibody to induce EBV lytic reactivation, followed by protein immunoblotting analysis using specific antibodies recognizing NSUN2, NSUN1, and c-Myc. GAPDH was used as a loading control. ( C ) Public-domain RNA-seq data of EBV-infected cell lines were collected and reanalyzed using the customized pipeline to identify the differentially expressed genes (adjust P -value < 0.05 as cutoff). The distinct gene expression level of NSUN2/1 due to EBV lytic reactivation was illustrated. ( D and E ) Akata cells were transiently transfected with siRNAs targeting NSUN2/1 (D) or TRIM25 (E), or siNT. The RNAs were extracted and subjected to RT-qPCR analysis of EBV viral genes (BZLF1, BMRF1), which are normalized to GAPDH. ( F–H ) AGS.BX1 cells were transfected with siRNA targeting NSUN1 (F), NSUN2 (G), or TRIM25 (H). EBV viral gene expression was measured by RT-qPCR. Results of at least three biological replicates were presented as mean ± SD (* P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001, Student’s t -test). ( I ) A model that human gamma-herpesviruses (KSHV, EBV) downregulate NSUN2/1 and decrease m5C modification of TRIM25 mRNA to favor their lytic replication. c-Myc is downregulated due to KSHV/EBV lytic replication, which reduces NSUN2/1 expression. Coordinately, NSUN2/1-mediated m5C modification of TRIM25 mRNA is inhibited, which impairs its stability. As TRIM25 is a key E3 ubiquitin ligase in RIG-I signal transduction, its inhibition disrupts RIG-I mediated antiviral sensing and thus favors KSHV/EBV lytic replication.
    Figure Legend Snippet: EBV also reduced expression of NSUN2/1 that restrict its lytic infection. ( A ) BJAB cells were inoculated with EBV.BX viruses. Cell lysates were collected at 48 h post infection and were followed by protein immunoblotting analysis using specific antibodies recognizing NSUN2 and NSUN1. ( B ) Akata and Akata BX cells were treated with a human IgG antibody to induce EBV lytic reactivation, followed by protein immunoblotting analysis using specific antibodies recognizing NSUN2, NSUN1, and c-Myc. GAPDH was used as a loading control. ( C ) Public-domain RNA-seq data of EBV-infected cell lines were collected and reanalyzed using the customized pipeline to identify the differentially expressed genes (adjust P -value < 0.05 as cutoff). The distinct gene expression level of NSUN2/1 due to EBV lytic reactivation was illustrated. ( D and E ) Akata cells were transiently transfected with siRNAs targeting NSUN2/1 (D) or TRIM25 (E), or siNT. The RNAs were extracted and subjected to RT-qPCR analysis of EBV viral genes (BZLF1, BMRF1), which are normalized to GAPDH. ( F–H ) AGS.BX1 cells were transfected with siRNA targeting NSUN1 (F), NSUN2 (G), or TRIM25 (H). EBV viral gene expression was measured by RT-qPCR. Results of at least three biological replicates were presented as mean ± SD (* P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001, Student’s t -test). ( I ) A model that human gamma-herpesviruses (KSHV, EBV) downregulate NSUN2/1 and decrease m5C modification of TRIM25 mRNA to favor their lytic replication. c-Myc is downregulated due to KSHV/EBV lytic replication, which reduces NSUN2/1 expression. Coordinately, NSUN2/1-mediated m5C modification of TRIM25 mRNA is inhibited, which impairs its stability. As TRIM25 is a key E3 ubiquitin ligase in RIG-I signal transduction, its inhibition disrupts RIG-I mediated antiviral sensing and thus favors KSHV/EBV lytic replication.

    Techniques Used: Expressing, Infection, Western Blot, Control, RNA Sequencing, Gene Expression, Transfection, Quantitative RT-PCR, Modification, Ubiquitin Proteomics, Transduction, Inhibition



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    Addgene inc self targeting gfp construct
    KSHV dysregulated NSUN2/1 expression via <t>c-Myc.</t> ( A and B ) Transcription factors (TFs) that regulate NSUN2/1 gene expression were predicted by analyzing public-domain ChIP-seq datasets. Venn diagram (A) showed the overlapped TFs across the indicated datasets. Six TF candidates including c-Myc (B) were listed. ( C ) Integrative genomics viewer (IGV) visualization of c-Myc occupancy near the NSUN2/1 promoter regions using the public-domain c-Myc ChIP-seq dataset of five lymphoma B cell lines ( GSE30726 ). ( D ) c-Myc binding motif near the promoter regions of NSUN2/1 was illustrated. ( E ) TREx.BCBL1.Rta cells were subjected to ChIP-PCR analysis for quantification of c-Myc association with the promoter regions of NSUN2/1 using an antibody recognizing c-Myc protein for its immunoprecipitation or a control IgG antibody, followed by qPCR analysis using three sets of primers (Set 1–3) targeting NSUN2 or NSUN1 promoter. ( F ) Public-domain <t>RNA-seq</t> data of KSHV-infected cell lines were collected and reanalyzed using the customized pipeline to identify the differentially expressed genes (adjust P -value < 0.05 as cutoff). The distinct gene expression level of c-Myc due to KSHV lytic reactivation was illustrated. ( G ) TREx.BCBL1.Rta, iSLK.BAC16, and iSLK.r219 cells were treated with Dox or mock, followed by protein immunoblotting analysis of c-Myc using its specific antibody. GAPDH was used as a loading control. ( H ) TREx.BCBL1.Rta cells were transduced with the <t>shRNA</t> targeting c-Myc or non-targeting control, followed by protein immunoblotting analysis of c-Myc, NSUN2, NSUN1, and KSHV K8.1. ( I and J ) TREx.BCBL1.Rta cells were treated with c-Myc inhibitors, EN4 (H) or 10074-G5 (I), at a series of concentrations or mock, followed by protein immunoblotting analysis of c-Myc, NSUN2, and NSUN1 using their specific antibodies. ( K ) TIME cells were transiently transfected with siRNAs (si1, si2) targeting c-Myc or siNT, followed by inoculation with KSHV.BAC16 viruses. These cells were harvested for nuclei staining with DAPI. GFP fluorescence signal indicating KSHV-infected cells was quantified (**** P <0.0001). ( L ) TIME cells were treated with the c-Myc inhibitor EN4 at the indicated concentrations, followed by transfection of NSUN1 or NSUN2 cDNAs. These cells were inoculated with KSHV.BAC16 viruses and lysed for protein immunoblotting assays of KSHV K8.1 protein.
    Self Targeting Gfp Construct, supplied by Addgene inc, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    self targeting gfp construct - by Bioz Stars, 2026-06
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    target plasmids  (Azenta)
    86
    Azenta target plasmids
    KSHV dysregulated NSUN2/1 expression via <t>c-Myc.</t> ( A and B ) Transcription factors (TFs) that regulate NSUN2/1 gene expression were predicted by analyzing public-domain ChIP-seq datasets. Venn diagram (A) showed the overlapped TFs across the indicated datasets. Six TF candidates including c-Myc (B) were listed. ( C ) Integrative genomics viewer (IGV) visualization of c-Myc occupancy near the NSUN2/1 promoter regions using the public-domain c-Myc ChIP-seq dataset of five lymphoma B cell lines ( GSE30726 ). ( D ) c-Myc binding motif near the promoter regions of NSUN2/1 was illustrated. ( E ) TREx.BCBL1.Rta cells were subjected to ChIP-PCR analysis for quantification of c-Myc association with the promoter regions of NSUN2/1 using an antibody recognizing c-Myc protein for its immunoprecipitation or a control IgG antibody, followed by qPCR analysis using three sets of primers (Set 1–3) targeting NSUN2 or NSUN1 promoter. ( F ) Public-domain <t>RNA-seq</t> data of KSHV-infected cell lines were collected and reanalyzed using the customized pipeline to identify the differentially expressed genes (adjust P -value < 0.05 as cutoff). The distinct gene expression level of c-Myc due to KSHV lytic reactivation was illustrated. ( G ) TREx.BCBL1.Rta, iSLK.BAC16, and iSLK.r219 cells were treated with Dox or mock, followed by protein immunoblotting analysis of c-Myc using its specific antibody. GAPDH was used as a loading control. ( H ) TREx.BCBL1.Rta cells were transduced with the <t>shRNA</t> targeting c-Myc or non-targeting control, followed by protein immunoblotting analysis of c-Myc, NSUN2, NSUN1, and KSHV K8.1. ( I and J ) TREx.BCBL1.Rta cells were treated with c-Myc inhibitors, EN4 (H) or 10074-G5 (I), at a series of concentrations or mock, followed by protein immunoblotting analysis of c-Myc, NSUN2, and NSUN1 using their specific antibodies. ( K ) TIME cells were transiently transfected with siRNAs (si1, si2) targeting c-Myc or siNT, followed by inoculation with KSHV.BAC16 viruses. These cells were harvested for nuclei staining with DAPI. GFP fluorescence signal indicating KSHV-infected cells was quantified (**** P <0.0001). ( L ) TIME cells were treated with the c-Myc inhibitor EN4 at the indicated concentrations, followed by transfection of NSUN1 or NSUN2 cDNAs. These cells were inoculated with KSHV.BAC16 viruses and lysed for protein immunoblotting assays of KSHV K8.1 protein.
    Target Plasmids, supplied by Azenta, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/target plasmids/product/Azenta
    Average 86 stars, based on 1 article reviews
    target plasmids - by Bioz Stars, 2026-06
    86/100 stars
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    target gene plasmids  (Yeasen Biotechnology)
    86
    Yeasen Biotechnology target gene plasmids
    KSHV dysregulated NSUN2/1 expression via <t>c-Myc.</t> ( A and B ) Transcription factors (TFs) that regulate NSUN2/1 gene expression were predicted by analyzing public-domain ChIP-seq datasets. Venn diagram (A) showed the overlapped TFs across the indicated datasets. Six TF candidates including c-Myc (B) were listed. ( C ) Integrative genomics viewer (IGV) visualization of c-Myc occupancy near the NSUN2/1 promoter regions using the public-domain c-Myc ChIP-seq dataset of five lymphoma B cell lines ( GSE30726 ). ( D ) c-Myc binding motif near the promoter regions of NSUN2/1 was illustrated. ( E ) TREx.BCBL1.Rta cells were subjected to ChIP-PCR analysis for quantification of c-Myc association with the promoter regions of NSUN2/1 using an antibody recognizing c-Myc protein for its immunoprecipitation or a control IgG antibody, followed by qPCR analysis using three sets of primers (Set 1–3) targeting NSUN2 or NSUN1 promoter. ( F ) Public-domain <t>RNA-seq</t> data of KSHV-infected cell lines were collected and reanalyzed using the customized pipeline to identify the differentially expressed genes (adjust P -value < 0.05 as cutoff). The distinct gene expression level of c-Myc due to KSHV lytic reactivation was illustrated. ( G ) TREx.BCBL1.Rta, iSLK.BAC16, and iSLK.r219 cells were treated with Dox or mock, followed by protein immunoblotting analysis of c-Myc using its specific antibody. GAPDH was used as a loading control. ( H ) TREx.BCBL1.Rta cells were transduced with the <t>shRNA</t> targeting c-Myc or non-targeting control, followed by protein immunoblotting analysis of c-Myc, NSUN2, NSUN1, and KSHV K8.1. ( I and J ) TREx.BCBL1.Rta cells were treated with c-Myc inhibitors, EN4 (H) or 10074-G5 (I), at a series of concentrations or mock, followed by protein immunoblotting analysis of c-Myc, NSUN2, and NSUN1 using their specific antibodies. ( K ) TIME cells were transiently transfected with siRNAs (si1, si2) targeting c-Myc or siNT, followed by inoculation with KSHV.BAC16 viruses. These cells were harvested for nuclei staining with DAPI. GFP fluorescence signal indicating KSHV-infected cells was quantified (**** P <0.0001). ( L ) TIME cells were treated with the c-Myc inhibitor EN4 at the indicated concentrations, followed by transfection of NSUN1 or NSUN2 cDNAs. These cells were inoculated with KSHV.BAC16 viruses and lysed for protein immunoblotting assays of KSHV K8.1 protein.
    Target Gene Plasmids, supplied by Yeasen Biotechnology, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/target gene plasmids/product/Yeasen Biotechnology
    Average 86 stars, based on 1 article reviews
    target gene plasmids - by Bioz Stars, 2026-06
    86/100 stars
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    Image Search Results


    Metastasis-related 12-gene signature, and identification of UPK1B and CGB5 as independent prognostic markers associated with advanced metastatic GC. (A) Venn diagram showing the intersection of genes upregulated in N1-3 stage and M1 stage GC tissues. (B) LASSO Cox regression analysis based on TCGA-STAD data identified a metastasis-associated prognostic risk signature. LASSO coefficient profiles of the signature genes in the merged dataset are shown (top) and the coefficient profile plot is generated against the log(λ) sequence (bottom). (C) The high-risk group of patients with GC exhibited a poor prognosis (overall survival). High expression of (D) CGB5 and (E) UPK1B was associated with poor overall survival in patients with GC based on data from the TCGA-STAD cohort. (F) Elevated UPK1B expression predicted poor prognosis (overall survival) of patients with GC in the Kaplan-Meier plotter database. UPK1B expression was increased in (G) M1 compared with M0 and in (H) N2 metastatic stage GC tissues compared with N0 stage. UPK1B, uroplakin 1B; CGB5, chorionic gonadotropin subunit β-5; GC, gastric cancer; LASSO, Least Absolute Shrinkage and Selection Operator; TCGA-STAD, The Cancer Genome Atlas Stomach Adenocarcinoma; HR, hazard ratio; TPM, transcripts per million.

    Journal: Experimental and Therapeutic Medicine

    Article Title: CDX2-UPK1B-PIK3IP1-PI3K/AKT signaling axis regulates gastric cancer cell invasion and migration and influences patient prognosis

    doi: 10.3892/etm.2026.13179

    Figure Lengend Snippet: Metastasis-related 12-gene signature, and identification of UPK1B and CGB5 as independent prognostic markers associated with advanced metastatic GC. (A) Venn diagram showing the intersection of genes upregulated in N1-3 stage and M1 stage GC tissues. (B) LASSO Cox regression analysis based on TCGA-STAD data identified a metastasis-associated prognostic risk signature. LASSO coefficient profiles of the signature genes in the merged dataset are shown (top) and the coefficient profile plot is generated against the log(λ) sequence (bottom). (C) The high-risk group of patients with GC exhibited a poor prognosis (overall survival). High expression of (D) CGB5 and (E) UPK1B was associated with poor overall survival in patients with GC based on data from the TCGA-STAD cohort. (F) Elevated UPK1B expression predicted poor prognosis (overall survival) of patients with GC in the Kaplan-Meier plotter database. UPK1B expression was increased in (G) M1 compared with M0 and in (H) N2 metastatic stage GC tissues compared with N0 stage. UPK1B, uroplakin 1B; CGB5, chorionic gonadotropin subunit β-5; GC, gastric cancer; LASSO, Least Absolute Shrinkage and Selection Operator; TCGA-STAD, The Cancer Genome Atlas Stomach Adenocarcinoma; HR, hazard ratio; TPM, transcripts per million.

    Article Snippet: For gene knockdown experiments, cells were transfected with a GV493 short hairpin RNA (sh) plasmid targeting UPK1B ( ) or a negative control scrambled plasmid (sh-NC), both obtained from GeneChem, Inc.

    Techniques: Generated, Sequencing, Expressing, Selection

    UPK1B drives GC cell invasion and migration in a PI3K/AKT-dependent manner. (A) Gene set enrichment analysis indicated that genes upregulated in the UPK1B-high group were enriched in the PI3K/AKT pathway. (B) Protein levels of UPK1B in GC cell lines. (C) Knockdown of UPK1B reduced PI3K/AKT activation in MKN45 cells. Silencing UPK1B suppressed the (D) migration/invasion capacity and (E) wound closure rate of MKN45 cells. (F) Overexpression of UPK1B enhanced PI3K/AKT pathway activation in AGS cells, which was attenuated by the PI3K inhibitor LY294002. Inhibition of PI3K/AKT signaling reversed UPK1B-induced (G) migration/invasion capacity and (H) wound closure rate of AGS cells. UPK1B, uroplakin 1B; GC, gastric cancer; p-, phosphorylated; sh, short hairpin RNA; NC, negative control; OE, overexpression.

    Journal: Experimental and Therapeutic Medicine

    Article Title: CDX2-UPK1B-PIK3IP1-PI3K/AKT signaling axis regulates gastric cancer cell invasion and migration and influences patient prognosis

    doi: 10.3892/etm.2026.13179

    Figure Lengend Snippet: UPK1B drives GC cell invasion and migration in a PI3K/AKT-dependent manner. (A) Gene set enrichment analysis indicated that genes upregulated in the UPK1B-high group were enriched in the PI3K/AKT pathway. (B) Protein levels of UPK1B in GC cell lines. (C) Knockdown of UPK1B reduced PI3K/AKT activation in MKN45 cells. Silencing UPK1B suppressed the (D) migration/invasion capacity and (E) wound closure rate of MKN45 cells. (F) Overexpression of UPK1B enhanced PI3K/AKT pathway activation in AGS cells, which was attenuated by the PI3K inhibitor LY294002. Inhibition of PI3K/AKT signaling reversed UPK1B-induced (G) migration/invasion capacity and (H) wound closure rate of AGS cells. UPK1B, uroplakin 1B; GC, gastric cancer; p-, phosphorylated; sh, short hairpin RNA; NC, negative control; OE, overexpression.

    Article Snippet: For gene knockdown experiments, cells were transfected with a GV493 short hairpin RNA (sh) plasmid targeting UPK1B ( ) or a negative control scrambled plasmid (sh-NC), both obtained from GeneChem, Inc.

    Techniques: Migration, Knockdown, Activation Assay, Over Expression, Inhibition, shRNA, Negative Control

    CDX2 acts as a transcriptional repressor of UPK1B and its high expression is associated with favorable prognosis of patients with GC. (A) Venn diagram showing overlapping predicted transcriptional regulators of UPK1B from ChEA and ChEA3 databases. (B) Knockdown of CDX2 in AGS cells resulted in increased UPK1B (C) mRNA and (D) protein expression. (E) Overexpression of CDX2 in MKN45 cells reduced UPK1B protein levels. Data from (F) The Cancer Genome Atlas Stomach Adenocarcinoma cohort and (G) the Kaplan-Meier plotter database indicated that high CDX2 expression was associated with improved prognosis of patients with GC. UPK1B, uroplakin 1B; GC, gastric cancer; si, small interfering RNA; NC, negative control; OE, overexpression; HR, hazard ratio; CDX2, caudal-related homeobox transcription factor 2; ChEA, ChIP-X Enrichment Analysis.

    Journal: Experimental and Therapeutic Medicine

    Article Title: CDX2-UPK1B-PIK3IP1-PI3K/AKT signaling axis regulates gastric cancer cell invasion and migration and influences patient prognosis

    doi: 10.3892/etm.2026.13179

    Figure Lengend Snippet: CDX2 acts as a transcriptional repressor of UPK1B and its high expression is associated with favorable prognosis of patients with GC. (A) Venn diagram showing overlapping predicted transcriptional regulators of UPK1B from ChEA and ChEA3 databases. (B) Knockdown of CDX2 in AGS cells resulted in increased UPK1B (C) mRNA and (D) protein expression. (E) Overexpression of CDX2 in MKN45 cells reduced UPK1B protein levels. Data from (F) The Cancer Genome Atlas Stomach Adenocarcinoma cohort and (G) the Kaplan-Meier plotter database indicated that high CDX2 expression was associated with improved prognosis of patients with GC. UPK1B, uroplakin 1B; GC, gastric cancer; si, small interfering RNA; NC, negative control; OE, overexpression; HR, hazard ratio; CDX2, caudal-related homeobox transcription factor 2; ChEA, ChIP-X Enrichment Analysis.

    Article Snippet: For gene knockdown experiments, cells were transfected with a GV493 short hairpin RNA (sh) plasmid targeting UPK1B ( ) or a negative control scrambled plasmid (sh-NC), both obtained from GeneChem, Inc.

    Techniques: Expressing, Knockdown, Over Expression, Small Interfering RNA, Negative Control

    UPK1B activates PI3K/AKT signaling by antagonizing the inhibitory regulator PIK3IP1 in gastric cancer cells. (A) Venn diagram showing that PIK3IP1 was identified as a putative UPK1B-interacting partner based on BioGRID and HIPPIE protein-protein interaction databases. (B) UPK1B and PIK3IP1 co-localized in the cytoplasm and plasma membrane of MKN45 cells. (C) Interaction between UPK1B and PIK3IP1 in MKN45 cells. (D) Knockdown of PIK3IP1 in MKN45 cells. (E) Silencing PIK3IP1 in UPK1B-knockdown MKN45 cells restored PI3K/AKT pathway activation. Knockdown of PIK3IP1 reversed the decrease in (F) migration/invasion and (G) wound-healing capacity in UPK1B-silenced MKN45 cells. UPK1B, uroplakin 1B; p-, phosphorylated; si, small interfering RNA; sh, short hairpin RNA; NC, negative control; PIK3IP1, PI3K inhibitor interacting protein 1; HIPPIE, Human Integrated Protein-Protein Interaction Reference; IP, immunoprecipitation.

    Journal: Experimental and Therapeutic Medicine

    Article Title: CDX2-UPK1B-PIK3IP1-PI3K/AKT signaling axis regulates gastric cancer cell invasion and migration and influences patient prognosis

    doi: 10.3892/etm.2026.13179

    Figure Lengend Snippet: UPK1B activates PI3K/AKT signaling by antagonizing the inhibitory regulator PIK3IP1 in gastric cancer cells. (A) Venn diagram showing that PIK3IP1 was identified as a putative UPK1B-interacting partner based on BioGRID and HIPPIE protein-protein interaction databases. (B) UPK1B and PIK3IP1 co-localized in the cytoplasm and plasma membrane of MKN45 cells. (C) Interaction between UPK1B and PIK3IP1 in MKN45 cells. (D) Knockdown of PIK3IP1 in MKN45 cells. (E) Silencing PIK3IP1 in UPK1B-knockdown MKN45 cells restored PI3K/AKT pathway activation. Knockdown of PIK3IP1 reversed the decrease in (F) migration/invasion and (G) wound-healing capacity in UPK1B-silenced MKN45 cells. UPK1B, uroplakin 1B; p-, phosphorylated; si, small interfering RNA; sh, short hairpin RNA; NC, negative control; PIK3IP1, PI3K inhibitor interacting protein 1; HIPPIE, Human Integrated Protein-Protein Interaction Reference; IP, immunoprecipitation.

    Article Snippet: For gene knockdown experiments, cells were transfected with a GV493 short hairpin RNA (sh) plasmid targeting UPK1B ( ) or a negative control scrambled plasmid (sh-NC), both obtained from GeneChem, Inc.

    Techniques: Clinical Proteomics, Membrane, Knockdown, Activation Assay, Migration, Small Interfering RNA, shRNA, Negative Control, Immunoprecipitation

    KSHV dysregulated NSUN2/1 expression via c-Myc. ( A and B ) Transcription factors (TFs) that regulate NSUN2/1 gene expression were predicted by analyzing public-domain ChIP-seq datasets. Venn diagram (A) showed the overlapped TFs across the indicated datasets. Six TF candidates including c-Myc (B) were listed. ( C ) Integrative genomics viewer (IGV) visualization of c-Myc occupancy near the NSUN2/1 promoter regions using the public-domain c-Myc ChIP-seq dataset of five lymphoma B cell lines ( GSE30726 ). ( D ) c-Myc binding motif near the promoter regions of NSUN2/1 was illustrated. ( E ) TREx.BCBL1.Rta cells were subjected to ChIP-PCR analysis for quantification of c-Myc association with the promoter regions of NSUN2/1 using an antibody recognizing c-Myc protein for its immunoprecipitation or a control IgG antibody, followed by qPCR analysis using three sets of primers (Set 1–3) targeting NSUN2 or NSUN1 promoter. ( F ) Public-domain RNA-seq data of KSHV-infected cell lines were collected and reanalyzed using the customized pipeline to identify the differentially expressed genes (adjust P -value < 0.05 as cutoff). The distinct gene expression level of c-Myc due to KSHV lytic reactivation was illustrated. ( G ) TREx.BCBL1.Rta, iSLK.BAC16, and iSLK.r219 cells were treated with Dox or mock, followed by protein immunoblotting analysis of c-Myc using its specific antibody. GAPDH was used as a loading control. ( H ) TREx.BCBL1.Rta cells were transduced with the shRNA targeting c-Myc or non-targeting control, followed by protein immunoblotting analysis of c-Myc, NSUN2, NSUN1, and KSHV K8.1. ( I and J ) TREx.BCBL1.Rta cells were treated with c-Myc inhibitors, EN4 (H) or 10074-G5 (I), at a series of concentrations or mock, followed by protein immunoblotting analysis of c-Myc, NSUN2, and NSUN1 using their specific antibodies. ( K ) TIME cells were transiently transfected with siRNAs (si1, si2) targeting c-Myc or siNT, followed by inoculation with KSHV.BAC16 viruses. These cells were harvested for nuclei staining with DAPI. GFP fluorescence signal indicating KSHV-infected cells was quantified (**** P <0.0001). ( L ) TIME cells were treated with the c-Myc inhibitor EN4 at the indicated concentrations, followed by transfection of NSUN1 or NSUN2 cDNAs. These cells were inoculated with KSHV.BAC16 viruses and lysed for protein immunoblotting assays of KSHV K8.1 protein.

    Journal: Nucleic Acids Research

    Article Title: m5C RNA methylation is dysregulated by oncogenic herpesviruses via c-Myc signaling to counteract host antiviral factors

    doi: 10.1093/nar/gkag251

    Figure Lengend Snippet: KSHV dysregulated NSUN2/1 expression via c-Myc. ( A and B ) Transcription factors (TFs) that regulate NSUN2/1 gene expression were predicted by analyzing public-domain ChIP-seq datasets. Venn diagram (A) showed the overlapped TFs across the indicated datasets. Six TF candidates including c-Myc (B) were listed. ( C ) Integrative genomics viewer (IGV) visualization of c-Myc occupancy near the NSUN2/1 promoter regions using the public-domain c-Myc ChIP-seq dataset of five lymphoma B cell lines ( GSE30726 ). ( D ) c-Myc binding motif near the promoter regions of NSUN2/1 was illustrated. ( E ) TREx.BCBL1.Rta cells were subjected to ChIP-PCR analysis for quantification of c-Myc association with the promoter regions of NSUN2/1 using an antibody recognizing c-Myc protein for its immunoprecipitation or a control IgG antibody, followed by qPCR analysis using three sets of primers (Set 1–3) targeting NSUN2 or NSUN1 promoter. ( F ) Public-domain RNA-seq data of KSHV-infected cell lines were collected and reanalyzed using the customized pipeline to identify the differentially expressed genes (adjust P -value < 0.05 as cutoff). The distinct gene expression level of c-Myc due to KSHV lytic reactivation was illustrated. ( G ) TREx.BCBL1.Rta, iSLK.BAC16, and iSLK.r219 cells were treated with Dox or mock, followed by protein immunoblotting analysis of c-Myc using its specific antibody. GAPDH was used as a loading control. ( H ) TREx.BCBL1.Rta cells were transduced with the shRNA targeting c-Myc or non-targeting control, followed by protein immunoblotting analysis of c-Myc, NSUN2, NSUN1, and KSHV K8.1. ( I and J ) TREx.BCBL1.Rta cells were treated with c-Myc inhibitors, EN4 (H) or 10074-G5 (I), at a series of concentrations or mock, followed by protein immunoblotting analysis of c-Myc, NSUN2, and NSUN1 using their specific antibodies. ( K ) TIME cells were transiently transfected with siRNAs (si1, si2) targeting c-Myc or siNT, followed by inoculation with KSHV.BAC16 viruses. These cells were harvested for nuclei staining with DAPI. GFP fluorescence signal indicating KSHV-infected cells was quantified (**** P <0.0001). ( L ) TIME cells were treated with the c-Myc inhibitor EN4 at the indicated concentrations, followed by transfection of NSUN1 or NSUN2 cDNAs. These cells were inoculated with KSHV.BAC16 viruses and lysed for protein immunoblotting assays of KSHV K8.1 protein.

    Article Snippet: Short hairpin RNA (shRNA) targeting c-Myc was purchased from Addgene (Cat#15662).

    Techniques: Expressing, Gene Expression, ChIP-sequencing, Binding Assay, Immunoprecipitation, Control, RNA Sequencing, Infection, Western Blot, Transduction, shRNA, Transfection, Staining, Fluorescence

    EBV also reduced expression of NSUN2/1 that restrict its lytic infection. ( A ) BJAB cells were inoculated with EBV.BX viruses. Cell lysates were collected at 48 h post infection and were followed by protein immunoblotting analysis using specific antibodies recognizing NSUN2 and NSUN1. ( B ) Akata and Akata BX cells were treated with a human IgG antibody to induce EBV lytic reactivation, followed by protein immunoblotting analysis using specific antibodies recognizing NSUN2, NSUN1, and c-Myc. GAPDH was used as a loading control. ( C ) Public-domain RNA-seq data of EBV-infected cell lines were collected and reanalyzed using the customized pipeline to identify the differentially expressed genes (adjust P -value < 0.05 as cutoff). The distinct gene expression level of NSUN2/1 due to EBV lytic reactivation was illustrated. ( D and E ) Akata cells were transiently transfected with siRNAs targeting NSUN2/1 (D) or TRIM25 (E), or siNT. The RNAs were extracted and subjected to RT-qPCR analysis of EBV viral genes (BZLF1, BMRF1), which are normalized to GAPDH. ( F–H ) AGS.BX1 cells were transfected with siRNA targeting NSUN1 (F), NSUN2 (G), or TRIM25 (H). EBV viral gene expression was measured by RT-qPCR. Results of at least three biological replicates were presented as mean ± SD (* P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001, Student’s t -test). ( I ) A model that human gamma-herpesviruses (KSHV, EBV) downregulate NSUN2/1 and decrease m5C modification of TRIM25 mRNA to favor their lytic replication. c-Myc is downregulated due to KSHV/EBV lytic replication, which reduces NSUN2/1 expression. Coordinately, NSUN2/1-mediated m5C modification of TRIM25 mRNA is inhibited, which impairs its stability. As TRIM25 is a key E3 ubiquitin ligase in RIG-I signal transduction, its inhibition disrupts RIG-I mediated antiviral sensing and thus favors KSHV/EBV lytic replication.

    Journal: Nucleic Acids Research

    Article Title: m5C RNA methylation is dysregulated by oncogenic herpesviruses via c-Myc signaling to counteract host antiviral factors

    doi: 10.1093/nar/gkag251

    Figure Lengend Snippet: EBV also reduced expression of NSUN2/1 that restrict its lytic infection. ( A ) BJAB cells were inoculated with EBV.BX viruses. Cell lysates were collected at 48 h post infection and were followed by protein immunoblotting analysis using specific antibodies recognizing NSUN2 and NSUN1. ( B ) Akata and Akata BX cells were treated with a human IgG antibody to induce EBV lytic reactivation, followed by protein immunoblotting analysis using specific antibodies recognizing NSUN2, NSUN1, and c-Myc. GAPDH was used as a loading control. ( C ) Public-domain RNA-seq data of EBV-infected cell lines were collected and reanalyzed using the customized pipeline to identify the differentially expressed genes (adjust P -value < 0.05 as cutoff). The distinct gene expression level of NSUN2/1 due to EBV lytic reactivation was illustrated. ( D and E ) Akata cells were transiently transfected with siRNAs targeting NSUN2/1 (D) or TRIM25 (E), or siNT. The RNAs were extracted and subjected to RT-qPCR analysis of EBV viral genes (BZLF1, BMRF1), which are normalized to GAPDH. ( F–H ) AGS.BX1 cells were transfected with siRNA targeting NSUN1 (F), NSUN2 (G), or TRIM25 (H). EBV viral gene expression was measured by RT-qPCR. Results of at least three biological replicates were presented as mean ± SD (* P < 0.05, ** P < 0.01, *** P < 0.001, **** P < 0.0001, Student’s t -test). ( I ) A model that human gamma-herpesviruses (KSHV, EBV) downregulate NSUN2/1 and decrease m5C modification of TRIM25 mRNA to favor their lytic replication. c-Myc is downregulated due to KSHV/EBV lytic replication, which reduces NSUN2/1 expression. Coordinately, NSUN2/1-mediated m5C modification of TRIM25 mRNA is inhibited, which impairs its stability. As TRIM25 is a key E3 ubiquitin ligase in RIG-I signal transduction, its inhibition disrupts RIG-I mediated antiviral sensing and thus favors KSHV/EBV lytic replication.

    Article Snippet: Short hairpin RNA (shRNA) targeting c-Myc was purchased from Addgene (Cat#15662).

    Techniques: Expressing, Infection, Western Blot, Control, RNA Sequencing, Gene Expression, Transfection, Quantitative RT-PCR, Modification, Ubiquitin Proteomics, Transduction, Inhibition