lentiviral expression plasmids psin ef2 sox2 pur plasmid nr 16577 addgene to express homo sapiens sry sex determining region y box 2  (Addgene inc)

Journal: BMC Cancer

Article Title: Isorhapontigenin inhibition of basal muscle-invasive bladder cancer attributed to its downregulation of SNHG1 and DNMT3b

doi: 10.1186/s12885-024-12490-5

Figure Lengend Snippet: SOX2 suppression mediated ISO-induced SNHG1 downregulation in BMIBC cells. (A) Wild-type SNHG1 promoter-driven luciferase reporter was cotransfected with pRL-TK into 5637 or UM-UC-3 cells; luciferase activity was measured 24 h post-transfection. TK served as an internal control. *Significantly different from the control group, p < 0.05. (B) Predicted transcription factor-binding sites in the SNHG1 promoter. (C) Transcription factor expression in 5637 cells treated with ISO at indicated times. (D) Schematic of SOX2 binding site in wild-type and mutant SNHG1 promoters. (E) 5637 cells cotransfected with wild-type or mutant SNHG1 promoter reporters and pRL-TK. (F) Western blot analysis of SOX2 expression in 5637 (SOX2) and 5637 (Vector) cells; GAPDH as control. (G) Western blot analysis of indicated proteins in cells with/without ISO treatment; GAPDH for normalization. (H) Relative SNHG1 levels in 5637 (SOX2) and 5637 (Vector) cells with 20 µM ISO or 0.1% DMSO were assessed by RT-qPCR. (I & J) Invasion abilities of 5637 (Vector) and 5637 (SOX2) cells with 20 µM ISO or 0.1% DMSO (I); relative invasion plotted (J). * p < 0.05, # p < 0.05, ♣ p < 0.05 vs. respective controls. (K & L) Colonies of 5637 (SOX2) and 5637 (Vector) cells in soft agar with 20 µM ISO or 0.1% DMSO (K). Colony counts shown in (L). * p < 0.05, # p < 0.05, ♣ p < 0.05 vs. respective controls

Article Snippet: The SOX2 overexpression construct pSin-EF2-SOX2 and the Myc-DNMT3b overexpression construct were purchased from Addgene (Cambridge, MA, USA).

Techniques: Luciferase, Activity Assay, Transfection, Control, Binding Assay, Expressing, Mutagenesis, Western Blot, Plasmid Preparation, Quantitative RT-PCR

Journal: BMC Cancer

Article Title: Isorhapontigenin inhibition of basal muscle-invasive bladder cancer attributed to its downregulation of SNHG1 and DNMT3b

doi: 10.1186/s12885-024-12490-5

Figure Lengend Snippet: miR-129 induced by ISO destabilizes SOX2 mRNA by interacting with its 3′UTR. A Relative SOX2 mRNA levels in 5637 cells treated with 20 μM ISO at indicated times by RT-qPCR. ( B ) Wild-type SOX2 promoter-driven luciferase reporter cotransfected with pRL-TK into 5637 cells. ( C ) 5637 cells were cotransfected with wild-type SOX2 mRNA 3′-UTR luciferase reporters and pRL-TK, followed by treatment with 20 μM ISO. TK served as the internal control. Luciferase activity is presented relative to SOX2 promoter activity. * p < 0.05 vs. 0.1% DMSO group. Bars represent mean ± SD from three independent trials. ( D ) Potential miRNA binding sites in SOX2 mRNA 3′-UTR from TargetScan database. ( E ) Relative miRNA levels in 5637 cells treated with 20 μM ISO at indicated times by RT-qPCR. ( F ) Relative miR-129 levels in UM-UC-3 cells with/without 20 μM ISO treatment. ( G ) miR-129 binding site in SOX2 mRNA 3′-UTR and mutants. ( H ) 5637 cells were cotransfected with wild-type or mutant SOX2 mRNA 3′-UTR luciferase reporters and pRL-TK, followed by treatment with 0.1% DMSO or 20 μM ISO for 24 h. Relative SOX2 mRNA 3′-UTR activity is presented. Data are shown as mean ± SD from three independent tests. * p < 0.05, ♣ p < 0.05 among corresponding groups

Article Snippet: The SOX2 overexpression construct pSin-EF2-SOX2 and the Myc-DNMT3b overexpression construct were purchased from Addgene (Cambridge, MA, USA).

Techniques: Quantitative RT-PCR, Luciferase, Control, Activity Assay, Binding Assay, Mutagenesis

Journal: BMC Cancer

Article Title: Isorhapontigenin inhibition of basal muscle-invasive bladder cancer attributed to its downregulation of SNHG1 and DNMT3b

doi: 10.1186/s12885-024-12490-5

Figure Lengend Snippet: Mechanistic insights into ISO-mediated suppression of BMIBC cell invasion and growth. ISO treatment suppresses SNHG1 transcription through the DNMT3b/miR-129/SOX2 axis, which leads to a reduction in MMP-2 and MMP-9 protein levels and an increase in PTEN protein expression, ultimately attenuating invasion and anchorage-independent growth of BIMBC cells

Article Snippet: The SOX2 overexpression construct pSin-EF2-SOX2 and the Myc-DNMT3b overexpression construct were purchased from Addgene (Cambridge, MA, USA).

Techniques: Expressing