ΔNp63α transcriptionally represses p53 target genes involved in the radiation-induced DNA damage response

Abstract

Background The DNA damage response (DDR) is a mechanism that protects cells against radiation-induced oxidative DNA damage by causing cell cycle arrest and apoptosis. TP63 is a member of the tumour suppressor TP53 gene family, and ΔNp63α, a TP63 splicing variant, is constitutively expressed in the stem cell-containing basal layer of stratified epithelial tissues, including the mammary gland, where it plays a critical role in stemness and tissue development. ΔNp63α has been reported to transcriptionally inhibit the tumour suppression protein p53. This p53-repressive activity may cause genomic instability in epithelial stem cells exposed to radiation. In this study, we analysed the inhibitory effect of ΔNp63α on radiation-induced DDR. Methods To elucidate the role of the p53-repressive effect of ΔNp63α in radiation response, we performed a p63-siRNA knockdown experiment using human mammary epithelial cells (HMECs) expressing ΔNp63α and then performed ectopic and entopic expression experiments using human induced pluripotent stem cells (hiPSCs). After irradiation, the expression of DDR-related genes and proteins in ΔNp63α-expressing and control cells was analysed by RT–qPCR, Western blotting, and flow cytometry. Results The mRNA/protein expression levels of BAX and p21 were significantly increased in p63-siRNA-treated HMECs (sip63) after X-ray irradiation (4 Gy, 0.7 Gy/min) but not in scramble-siRNA treated HMECs (scr). Transcriptomic analysis showed decreased RNA expression of cell cycle-related genes and increased expression of programmed cell death-related genes in sip63 cells compared to scr cells. Furthermore, flow cytometric analysis revealed an increase in apoptotic cells and a decrease in 5-ethynyl-2´-deoxyuridine uptake in sip63 cells compared to scr cells. On the other hand, both the ectopic and entopic expression of ΔNp63α in apoptosis-sensitive hiPSCs reduced the expression levels of BAX after irradiation and significantly decreased the number of apoptotic cells induced by radiation. Conclusion Taken together, these results indicate that ΔNp63α represses p53-related radiation-induced DDR, thereby potentially causing genomic instability in epithelial stem cells.

ΔNp63 Regulates Radioresistance in Human Head and Neck Squamous Carcinoma Cells

“…Kudo et al recently showed that ΔNp63 knockdown increases radiation-induced apoptosis in normal mammary epithelial cells expressing ΔNp63 [ 22 ]. In addition, they reported that ΔNp63α represses the p53-related radiation-induced DNA damage response, thereby inducing radioresistance of mammary epithelial stem cells [ 22 ]. Here, we found that ΔNp63 knockdown enhanced radiation-induced apoptosis in HNSCC cells.…”

“…Bretz et al reported that ΔNp63 activates the Fanconi anemia DNA repair pathway and limits the efficacy of DNA-damaging agent cisplatin treatment in SCC [ 21 ]. Recently, Kudo et al also reported that ΔNp63α represses the p53-related DNA damage response after irradiation [ 22 ].…”

p63: a crucial player in epithelial stemness regulation

“…TAp63 protein is instead expressed in luminal cells [ 78 ]. The expression of ΔNp63 in mammary progenitor cells serves to maintain their undifferentiated state and quiescence and therefore to promote stem cell activity [ 31 , 74 , 79 – 81 ]. In addition, it has been reported that ΔNp63 can reprogram adult luminal into basal cells by promoting an intermediate multipotent-like state, thus supporting the concept of reversible plasticity in epithelial cell fate [ 71 , 82 ].…”