Journal: bioRxiv
Article Title: Differential transcriptional activity of ΔNp63β is encoded by an isoform-specific C-terminus
doi: 10.1101/2024.12.03.626646
Figure Lengend Snippet: A) Schematic of TAp63 and ΔNp63 C-terminal TAD mutants. B) Protein expression of C-terminal TAD mutants in pcDNA plasmid constructs transiently transfected in HCT116 TP53 -/- cells. Negative control is an empty pcDNA backbone. C) Reporter assay of TAp63 and D) ΔNp63 C-terminal TAD mutants on a p63 RE (green) and a mutant (grey) p63 RE. E) Protein expression of ΔNp63 C-terminal TAD mutants in lentiviral vectors under 24hr doxycycline induction in HCT116 TP53 -/- cells. Negative control is GUS expressed in pCW57.1 vector. F) QRT-PCR analysis of KRT5 expression by ΔNp63 C-terminal TAD mutants G) QRT-PCR analysis of JAG2 expression by ΔNp63 C-terminal TAD mutants. H) Genome browser view of KRT5 locus displaying p63 and H3K27ac ChIP-seq binding data. I) Genome browser view of JAG2 locus displaying p63 and H3K27ac ChIP-seq binding data. Statistical analysis for qRT-PCR data was done using a a One-way ANOVA test (*: p -value <.05, **: p -value <.01, ***: p -value <.001, ****: p -value < 0.0001, ns = not significant) and a Two-way ANOVA ( ***: p -value <.001, ****: p -value < 0.0001, ns = not significant ) for reporter assay data.
Article Snippet: For pCW57.1, p63 isoforms in pENTR Twist backbone (Twist Biosciences) were cloned via Gateway cloning using LR Clonase enzyme.
Techniques: Expressing, Plasmid Preparation, Construct, Transfection, Negative Control, Reporter Assay, Mutagenesis, Quantitative RT-PCR, ChIP-sequencing, Binding Assay