TP63 is a member of the p53 gene family that encodes for up to ten different TA- and ΔN- isoforms through differential promoter usage and alternative C-terminal splicing. The TA isoforms (TA-p63α, β, γ, δ and ϵ) contain the N-terminal transactivation domain (TA1), whereas the ΔN isoforms (ΔN-p63α, β, γ, δ and ϵ) are transcribed from an internal promoter (P2) and lack the TA1 domain. A second C-terminal transactivation domain (TA2) present in all p63α and β isoforms has been reported. Besides being a master regulator of gene expression for squamous epithelial proliferation, differentiation and maintenance (germline TP63 mutations are causative for a subset of human ectodermal dysplasia syndromes -EDs-), TA- and ΔN-p63 isoforms play an important role in tumorigenesis. More recently, p63 was shown to modulate apoptosis in the female and male germ line in response to DNA damage. All isoforms share a large, immunoglobulin-like folded DNA binding domain that is responsible for binding to sequence-specific response elements (REs), whose overall consensus sequence is similar to the p53 RE. Since the ΔN-p63 isoforms lack the N-terminal transactivation domain, it was originally proposed that these proteins might act primarily as oncogenes through dominant-negative mechanisms. However, different studies indicate that ΔN-p63 protein itself can be transcriptionally active. For example, ΔN-p63α may directly contribute to tumorigenesis by up-regulating the chaperone protein Hsp70, which displays proliferative and anti-apoptotic functions or by repressing pro-apoptotic genes. Transcriptional activation of specific genes by ΔN-p63α, namely VDR and Id-3, has been also associated with an anti-tumorigenic role, i.e. a decrease in cell invasion. Using a defined functional assay in yeast where p63 isoform and RE sequence are the only variables (more than 80 different REs were tested), as well as mammalian-transcription assays (gene reporter assays, qPCR measurements, western blotting), we demonstrated that human TA- and ΔN-p63α proteins exhibited differences in transactivation specificity. In fact 21 REs were identified that exhibited higher or selective responsiveness to ΔN-p63α. These differences were not observed with the related p73 and p53 proteins isoforms and were dependent on specific features of the RE sequences. Based on gene annotations, we propose that cis-element sequence features might have been selected along with evolutionarily conserved, intrinsic differences in cooperative DNA binding of p63 proteins, to establish tighter control of the apoptotic processes.

Abstract 3402: ΔN-p63α and TA-p63α exhibit intrinsic differences in transactivation specificities that depend on distinct features of DNA target sites

CAMPOMENOSI, PAOLA;
2014-01-01

Abstract

TP63 is a member of the p53 gene family that encodes for up to ten different TA- and ΔN- isoforms through differential promoter usage and alternative C-terminal splicing. The TA isoforms (TA-p63α, β, γ, δ and ϵ) contain the N-terminal transactivation domain (TA1), whereas the ΔN isoforms (ΔN-p63α, β, γ, δ and ϵ) are transcribed from an internal promoter (P2) and lack the TA1 domain. A second C-terminal transactivation domain (TA2) present in all p63α and β isoforms has been reported. Besides being a master regulator of gene expression for squamous epithelial proliferation, differentiation and maintenance (germline TP63 mutations are causative for a subset of human ectodermal dysplasia syndromes -EDs-), TA- and ΔN-p63 isoforms play an important role in tumorigenesis. More recently, p63 was shown to modulate apoptosis in the female and male germ line in response to DNA damage. All isoforms share a large, immunoglobulin-like folded DNA binding domain that is responsible for binding to sequence-specific response elements (REs), whose overall consensus sequence is similar to the p53 RE. Since the ΔN-p63 isoforms lack the N-terminal transactivation domain, it was originally proposed that these proteins might act primarily as oncogenes through dominant-negative mechanisms. However, different studies indicate that ΔN-p63 protein itself can be transcriptionally active. For example, ΔN-p63α may directly contribute to tumorigenesis by up-regulating the chaperone protein Hsp70, which displays proliferative and anti-apoptotic functions or by repressing pro-apoptotic genes. Transcriptional activation of specific genes by ΔN-p63α, namely VDR and Id-3, has been also associated with an anti-tumorigenic role, i.e. a decrease in cell invasion. Using a defined functional assay in yeast where p63 isoform and RE sequence are the only variables (more than 80 different REs were tested), as well as mammalian-transcription assays (gene reporter assays, qPCR measurements, western blotting), we demonstrated that human TA- and ΔN-p63α proteins exhibited differences in transactivation specificity. In fact 21 REs were identified that exhibited higher or selective responsiveness to ΔN-p63α. These differences were not observed with the related p73 and p53 proteins isoforms and were dependent on specific features of the RE sequences. Based on gene annotations, we propose that cis-element sequence features might have been selected along with evolutionarily conserved, intrinsic differences in cooperative DNA binding of p63 proteins, to establish tighter control of the apoptotic processes.
Ciribilli, Yari; Bisio, Alessandra; Monti, Paola; Foggetti, Giorgia; Raimondi, Ivan; Campomenosi, Paola; Menichini, Paola; Fronza, Gilberto; Inga, Alberto
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11383/2024195
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