Journal: Scientific reports

Article Title: Syndecan-2 regulates PAD2 to exert antifibrotic effects on RA-ILD fibroblasts.

doi: 10.1038/s41598-022-06678-7

Figure Lengend Snippet: Figure 3. Syndecan-2 (SDC2) inhibits α-SMA and PAD2 expression by regulating CD148/PI3K/Akt signaling in RA-ILD fibroblasts. Control (n = 4) and RA-ILD (n = 4) fibroblasts were transfected with scramble (scr) or shCD148 via lentivirus as described in the Methods. Stably transfected cells were treated with syndecan-2 (10 μg/ml) for 24 h. After treatment, cells were subjected to (A) Western blot to measure phospho-Akt, α-SMA, and PAD2 expression, (B) RT-PCR to measure mRNA levels of PADI2 (PAD2), or (C) a Sp1 transcriptional activity assay as described in the Methods. (D) RA-ILD fibroblasts were transfected via lentivirus with empty vector or pLenti-GIII-CMV-C-term-HA plasmid carrying the PADI2 (PAD2-over). Stably transfected cells were treated with syndecan-2 (10 μM) for 24 h. After treatment, cells were subjected to RT-PCR to measure mRNA levels of COL1A1 and PADI2. Data are mean ± SEM. P < 0.05; significant comparisons by one-way ANOVA: * vs. control, † vs. RA-ILD fibroblasts + Scr, # vs. RA-ILD fibroblasts + Scr + SDC2. Abbreviations: α-SMA = alpha- smooth muscle actin, ILD = interstitial lung disease, p-Akt = phosphorylated Akt, PAD2 = peptidylarginine deiminase 2, PI3K = phosphoinositide 3-kinase, RA = rheumatoid arthritis, scr = scrambled short hairpin RNA, SDC2 = syndecan-2, shCD148 = CD148 short hairpin RNA, shSp1 = Sp1 short hairpin RNA.

Article Snippet: Sp1 transcriptional activity was measured using a commercially available kit from RayBiotech (Peachtree Corners, GA) according to the manufacturer’s instructions.

Techniques: Expressing, Control, Transfection, Stable Transfection, Western Blot, Reverse Transcription Polymerase Chain Reaction, Activity Assay, Plasmid Preparation, shRNA

Journal:

Article Title: Transcription Factors Pax6 and AP-2? Interact To Coordinate Corneal Epithelial Repair by Controlling Expression of Matrix Metalloproteinase Gelatinase B

doi: 10.1128/MCB.24.1.245-257.2004

Figure Lengend Snippet: Pax6 binding is found at two sites on the gelB promoter. (A) EMSA with GB(−467) as a probe. The arrow indicates a shifted band that is removed by a Pax6 PD consensus sequence (PDcon) and Pax6 antibody but not control oligonucleotides [GB(−487) and SP1con] or control antibodies (NF-κB and SP1). (B) DNase footprinting assay with recombinant Pax6/GST PD protein (PD) protected bases −492 to −465 of the gelB promoter that were not protected by GST alone. This sequence corresponds to the GB(−467) oligonucleotide sequence. (C) EMSA with GB(−359) as a probe. The arrow indicates a shifted band that is removed by antibody to Pax6 and AP-2, but not NF-κB. (D) Representation of the gelB promoter showing previously identified transcription factor binding sites and the locations of the new Pax6 elements.

Article Snippet: Probes containing the canonical DNA-binding sites for transcription factor Sp1 was purchased from Promega (Madison, Wis.).

Techniques: Binding Assay, Sequencing, DNase Footprinting Assay, Recombinant

Journal: BMC Medical Genetics

Article Title: SNP55, a new functional polymorphism of MDM2 -P2 promoter, contributes to allele-specific expression of MDM2 in endometrial cancers

doi: 10.1186/s12881-015-0216-8

Figure Lengend Snippet: The effect of SNP55 on MDM2 -P2 promoter activity. a The pGL4 luciferase reporter plasmids containing either SNP55T or SNP55C were co-transfected with pcDNA-LacZ (control) and pcDNA-Sp1 (N-terminus myc-tagged). b Western blot analysis confirming a gradient increase of nuclear factor kappa-B (NFκB) p50. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) is shown as an internal control. c The amount of pcDNA- nuclear factor kappa-B (NFκB) p50 (0–0.8 μg) was gradually increased and co-transfected with the SNP55C reporter plasmid. d The amount of pcDNA- nuclear factor kappa-B (NFκB) p50 (0–0.8 μg) was gradually increased and co-transfected with the SNP55T reporter plasmid

Article Snippet: Transcription factor Sp1 (Proteinone), extracted from HeLa cells, or recombinant NFκB p50 (Enzo) was prepared at a concentration of 1 μg/ml, in advance.

Techniques: Activity Assay, Luciferase, Transfection, Control, Western Blot, Plasmid Preparation

Journal: BMC Medical Genetics

Article Title: SNP55, a new functional polymorphism of MDM2 -P2 promoter, contributes to allele-specific expression of MDM2 in endometrial cancers

doi: 10.1186/s12881-015-0216-8

Figure Lengend Snippet: Biomolecular interaction analysis (BIA) using Biacore. a Biosensor figure of relative quantitative response with transcription factor Sp1, purified from HeLa cells. b Comparison of a response unit (RU) value at 600 s for ( a ). c Biosensor figure of the relative quantitative response with recombinant nuclear factor kappa-B (NFκB) p50. d Comparison of the RU value at 600 s for ( c )

Article Snippet: Transcription factor Sp1 (Proteinone), extracted from HeLa cells, or recombinant NFκB p50 (Enzo) was prepared at a concentration of 1 μg/ml, in advance.

Techniques: Purification, Comparison, Recombinant