Journal: Journal of Biological Chemistry

Article Title: Phosphorylated TP63 Induces Transcription of RPN13, Leading to NOS2 Protein Degradation

doi: 10.1074/jbc.m110.158642

Figure Lengend Snippet: FIGURE 1. Cisplatin induces RPN13 expression at the RNA level. Wild- type Np63 and Np63-S385G cells were treated with control medium () or 10 g/ml cisplatin (CIS; ) for 12 h. GAPDH was used as a loading control. A, RT-PCR analysis for RPN13 transcription. B, qPCR. Values for RPN13 (in relative units (RU)) were normalized to values for GAPDH, and values obtained from the control untreated samples were designated as 1. Experiments were performed in triplicate. FIGURE 2. Schematic representation of the human RPN13 gene pro- moter. The sequence of the human 1500-bp RPN13 promoter was found on the UCSC Genome Bioinformatics human genome web site, and certain potential TF-responsive elements (RE) were defined using TFSEARCH soft- ware. TF sequences are shown in boldface. The TSS is shown as an uppercase letter. The following responsive elements were located in the RPN13 pro- moter: TP63 (1376/1354, 1231/1216, 1189/1167, and 500/ 481), NF-Y/DDIT3 (1267/1246, 93/71, and 65/37), NF-B (995/ 985 and 724/716), STAT (857/843), and GAS (gamma-activated site; 971/862).

Article Snippet: Antibodies—We used a rabbit anti- Np63 polyclonal antibody (Ab-1, EMD Chemicals); a rabbit anti-TP63 (TP73L) monoclonal antibody (clone Y289, NB110-57309) and a rabbit anti-DDIT3 (DNA damage-inducible transcript 3) polyclonal antibody (NB100-78344) (Novus Biologicals); mouse monoclonal antibodies against -actin (Sigma) and NF-YA (Rockland Immunochemicals); and rabbit polyclonal antibodies against DNA topoisomerase II (TOP2A, ab74715), RPN13 (ab91567), NF- B p65 subunit (ab32536), NF- B p50 subunit (ab7549), STAT3 (signal transduction activator of transcription 3; ab32500), and UCH37 (ab38528) (Abcam).

Techniques: Expressing, Control, Reverse Transcription Polymerase Chain Reaction, Sequencing

Journal: Journal of Biological Chemistry

Article Title: Phosphorylated TP63 Induces Transcription of RPN13, Leading to NOS2 Protein Degradation

doi: 10.1074/jbc.m110.158642

Figure Lengend Snippet: FIGURE 1. Cisplatin induces RPN13 expression at the RNA level. Wild- type Np63 and Np63-S385G cells were treated with control medium () or 10 g/ml cisplatin (CIS; ) for 12 h. GAPDH was used as a loading control. A, RT-PCR analysis for RPN13 transcription. B, qPCR. Values for RPN13 (in relative units (RU)) were normalized to values for GAPDH, and values obtained from the control untreated samples were designated as 1. Experiments were performed in triplicate. FIGURE 2. Schematic representation of the human RPN13 gene pro- moter. The sequence of the human 1500-bp RPN13 promoter was found on the UCSC Genome Bioinformatics human genome web site, and certain potential TF-responsive elements (RE) were defined using TFSEARCH soft- ware. TF sequences are shown in boldface. The TSS is shown as an uppercase letter. The following responsive elements were located in the RPN13 pro- moter: TP63 (1376/1354, 1231/1216, 1189/1167, and 500/ 481), NF-Y/DDIT3 (1267/1246, 93/71, and 65/37), NF-B (995/ 985 and 724/716), STAT (857/843), and GAS (gamma-activated site; 971/862).

Article Snippet: Antibodies—We used a rabbit anti- Np63 polyclonal antibody (Ab-1, EMD Chemicals); a rabbit anti-TP63 (TP73L) monoclonal antibody (clone Y289, NB110-57309) and a rabbit anti-DDIT3 (DNA damage-inducible transcript 3) polyclonal antibody (NB100-78344) (Novus Biologicals); mouse monoclonal antibodies against -actin (Sigma) and NF-YA (Rockland Immunochemicals); and rabbit polyclonal antibodies against DNA topoisomerase II (TOP2A, ab74715), RPN13 (ab91567), NF- B p65 subunit (ab32536), NF- B p50 subunit (ab7549), STAT3 (signal transduction activator of transcription 3; ab32500), and UCH37 (ab38528) (Abcam).

Techniques: Expressing, Control, Reverse Transcription Polymerase Chain Reaction, Sequencing

Journal: PLoS ONE

Article Title: Keratinocyte Progenitor Cells Reside in Human Subcutaneous Adipose Tissue

doi: 10.1371/journal.pone.0118402

Figure Lengend Snippet: ASCs cultured in chambered slides were fixed and labeled with antibodies against p63 and DSG3. Undifferentiated ASCs and NHEKs (positive control) expressed (a) p63 and (b) DSG3 (both visible by green fluorescence). Scale bars, 20 μm. (c) The expression of p63 and DSG3 in ASCs and NHEKs was detected by flow cytometry.

Article Snippet: The expression of p63 and DSG3 (using primers Hs00978343_m1 and Hs00951897_m1, respectively; Applied Biosystems) was normalized to β-actin levels, and the comparative cycle threshold (Ct) method using the formula 2 -ΔΔCt was used to calculate relative mRNA levels.

Techniques: Cell Culture, Labeling, Positive Control, Fluorescence, Expressing, Flow Cytometry

Journal: PLoS ONE

Article Title: Keratinocyte Progenitor Cells Reside in Human Subcutaneous Adipose Tissue

doi: 10.1371/journal.pone.0118402

Figure Lengend Snippet: The expression of p63 and DSG3 was measured by (a) quantitative real-time PCR (relative to glyceraldehyde-3-phosphate dehydrogenase) and (b) by western blotting (relative to β-actin) in undifferentiated ASCs and those that had differentiated into adipocytes. Both markers were expressed at lower levels after differentiation.

Article Snippet: The expression of p63 and DSG3 (using primers Hs00978343_m1 and Hs00951897_m1, respectively; Applied Biosystems) was normalized to β-actin levels, and the comparative cycle threshold (Ct) method using the formula 2 -ΔΔCt was used to calculate relative mRNA levels.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Western Blot

Journal: PLoS ONE

Article Title: Keratinocyte Progenitor Cells Reside in Human Subcutaneous Adipose Tissue

doi: 10.1371/journal.pone.0118402

Figure Lengend Snippet: The p63 transcript (but not that of DSG3 ) was detected in human subcutaneous adipose tissue by RT-PCR but at a lower level than in NHEKs.

Article Snippet: The expression of p63 and DSG3 (using primers Hs00978343_m1 and Hs00951897_m1, respectively; Applied Biosystems) was normalized to β-actin levels, and the comparative cycle threshold (Ct) method using the formula 2 -ΔΔCt was used to calculate relative mRNA levels.

Techniques: Reverse Transcription Polymerase Chain Reaction