SFPQ•NONO and XLF function separately and together to promote DNA double-strand break repair via canonical nonhomologous end joining.

• Type of Material: Article
• Authors: Lahcen Jaafar, Emory University; Shuyi Li, Emory University; William Dynan, Emory University
• Language: English
• Date: 2016-12-06
• Publisher: Oxford University Press (OUP): Policy C - Option B
• Publication Version: Final Published Version
• Copyright Statement: © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.
• License: Creative Commons Attribution-NonCommercial 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.1093/nar/gkw1209
• Title of Journal or Parent Work: Nucleic Acids Research
• ISSN: 0305-1048
• Grant/Funding Information: This work was partially funded from Emory University's Open Access Publishing Fund.; Funding for open access charge: Emory University.; US National Institutes of Health [2 R01 CA98239].
• Research Categories: Health Sciences, Oncology; Health Sciences, Radiology; Health Sciences, General

Abstract

A complex of two related mammalian proteins, SFPQ and NONO, promotes DNA double-strand break repair via the canonical nonhomologous end joining (c-NHEJ) pathway. However, its mechanism of action is not fully understood. Here we describe an improved SFPQ•NONO-dependent in vitro end joining assay. We use this system to demonstrate that the SFPQ•NONO complex substitutes in vitro for the core c-NHEJ factor, XLF. Results are consistent with a model where SFPQ•NONO promotes sequence-independent pairing of DNA substrates, albeit in a way that differs in detail from XLF. Although SFPQ•NONO and XLF function redundantly in vitro, shRNA-mediated knockdown experiments indicate that NONO and XLF are both required for efficient end joining and radioresistance in cell-based assays. In addition, knockdown of NONO sensitizes cells to the interstrand crosslinking agent, cisplatin, whereas knockdown of XLF does not, and indeed suppresses the effect of NONO deficiency. These findings suggest that each protein has one or more unique activities, in addition to the DNA pairing revealed in vitro, that contribute to DNA repair in the more complex cellular milieu. The SFPQ•NONO complex contains an RNA binding domain, and prior work has demonstrated diverse roles in RNA metabolism. It is thus plausible that the additional repair function of NONO, revealed in cell-based assays, could involve RNA interaction.