Overexpression of the scaffold WD40 protein WRAP53β enhances the repair of and cell survival from DNA double-strand breaks

Abstract

Altered expression of the multifunctional protein WRAP53β (WD40 encoding RNA Antisense to p53), which targets repair factors to DNA double-strand breaks and factors involved in telomere elongation to Cajal bodies, is linked to carcinogenesis. While loss of WRAP53β function has been shown to disrupt processes regulated by this protein, the consequences of its overexpression remain unclear. Here we demonstrate that overexpression of WRAP53β disrupts the formation of and impairs the localization of coilin to Cajal bodies. At the same time, the function of this protein in the repair of DNA double-strand breaks is enhanced. Following irradiation, cells overexpressing WRAP53β exhibit more rapid clearance of phospho-histone H2AX (γH2AX), and more efficient homologous recombination and non-homologous end-joining, in association with fewer DNA breaks. Moreover, in these cells the ubiquitylation of damaged chromatin, which is known to facilitate the recruitment of repair factors and subsequent repair, is elevated. Knockdown of the ubiquitin ligase involved, ring-finger protein 8 (RNF8), which is recruited to DNA breaks by WRAP53β, attenuated this effect, suggesting that overexpression of WRAP53β leads to more rapid repair, as well as improved cell survival, by enhancing RNF8-mediated ubiquitylation at DNA breaks. Our present findings indicate that WRAP53β and RNF8 are rate-limiting factors in the repair of DNA double-strand breaks and raise the possibility that upregulation of WRAP53β may contribute to genomic stability in and survival of cancer cells.

Phosphorylation of the Cajal body protein WRAP53β by ATM promotes its involvement in the DNA damage response

“…WRAP53β has recently been shown to be a rate-limiting factor in the repair of double-strand breaks and overexpression of this protein results in more efficient HR and NHEJ repair and fewer DNA breaks 15 . Therefore, we asked whether phosphorylation of WRAP53β influences HR and NHEJ employing reporter assays that measure restoration of a functional GFP protein by these processes.…”

Biallelic mutations in WRAP53 result in dysfunctional telomeres, Cajal bodies and DNA repair, thereby causing Hoyeraal–Hreidarsson syndrome

“…The inability of mutant WRAP53β to accumulate in Cajal bodies and/or bind known interaction partners prevented various functions of this protein at this site. First, high overexpression of the mutant forms did not result in the collapse of Cajal bodies and/or inability of this organelle to form, in contrast to high overexpression of the wild-type protein 26,32 . Second, the mutant proteins no longer interacted with SMN, coilin, scaRNAs or telomerase, interactions known to be required for the localization of these factors to Cajal bodies [24][25][26] .…”

“…Proper assembly of WRAP53β in Cajal bodies is essential for their maintenance, as demonstrated by the findings that either loss or overexpression of WRAP53β causes these organelles to collapse, after which they cannot re-form 24,26,31,32 . Accordingly, overexpression of exogenous wild-type WRAP53β here led to the disappearance and/or prevented the formation of Cajal bodies in approximately half of the cells transfected ( Fig.…”

Suppression of TCAB1 Expression Induced Cellular Senescence by Lessening Proteasomal Degradation of p21 in Cancer Cells

“…It was reported that TCAB1 RNAi in U2OS, H1299 and HeLa cells increased the number of phospho-histone H2AX (γ-H2AX) foci in nonirradiated cells, indicating TCAB1 protects cells against accumulation of spontaneous DNA damage [14]. Cells overexpressing TCAB1 exhibited more rapid clearance of γ-H2AX, in association with homologous recombination and non-homologous end-joining at sites of less DNA breaks [26]. Reduced expression of TCAB1 in ovarian tumors correlated with attenuated DNA damage response and poor patient survival [19].…”