elifesciences.org RESEARCH ARTICLE DNA damage induces nuclear actin filament assembly by Formin-2 and Spire-1/2 that promotes efficient DNA repair Brittany J Belin 1,2 , Terri Lee 1 , R Dyche Mullins 1,2 * 1 Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, United States; 2 Physiology Course, Marine Biological Laboratory, Woods Hole, United States Abstract Actin filaments assemble inside the nucleus in response to multiple cellular perturbations, including heat shock, protein misfolding, integrin engagement, and serum stimulation. We find that DNA damage also generates nuclear actin filaments—detectable by phalloidin and live-cell actin probes—with three characteristic morphologies: (i) long, nucleoplasmic filaments; (ii) short, nucleolus-associated filaments; and (iii) dense, nucleoplasmic clusters. This DNA damage-induced nuclear actin assembly requires two biologically and physically linked nucleation factors: Formin-2 and Spire-1/Spire-2. Formin-2 accumulates in the nucleus after DNA damage, and depletion of either Formin-2 or actin’s nuclear import factor, importin-9, increases the number of DNA double-strand breaks (DSBs), linking nuclear actin filaments to efficient DSB clearance. Nuclear actin filaments are also required for nuclear oxidation induced by acute genotoxic stress. Our results reveal a previously unknown role for nuclear actin filaments in DNA repair and identify the molecular mechanisms creating these nuclear filaments. DOI: 10.7554/eLife.07735.001 Introduction Actin was first identified in muscle over seventy years ago (Straub, 1942) and has been established as a component of non-muscle cells for nearly half a century (Hatano and Oosawa, 1966). Subsequent work revealed how actin filaments help organize the cytoplasm of all eukaryotic cells, supporting many fundamental biological processes, including: motility, division, phagocytosis, endocytosis, and membrane trafficking. The first reports of actin inside the nucleus of a cell appeared forty years ago (LeStourgeon et al., 1975), and since that time, actin has been found in the nuclei of many different cell types, linked to a variety of nuclear processes (Pederson and Aebi, 2002). Recent work has identified the molecular mechanisms that control the nuclear concentration of actin, uncovering new roles for the actin-binding proteins profilin and cofilin as co-factors for actin’s nucleocytoplasmic transport. The nuclear export factor exportin-6 (XPO6) binds profilin-actin complexes in the nucleus—as well as a handful of other actin-binding proteins—and shuttles them into the cytoplasm (St ¨ uven et al., 2003; Bohnsack et al., 2006). Conversely, nuclear import of actin is regulated by importin-9 (IPO9), which transports cofilin–actin complexes from the cytoplasm into the nucleus (Dopie et al., 2012). In many organisms, the export factor XPO6 is not expressed in the oocytes of, and so the germinal vesicles contain a high concentration of actin (St ¨ uven et al., 2003). In Xenopus laevis oocytes, this germinal vesicle actin forms a filamentous mesh that protects nucleoli from gravity-induced aggregation (Feric and Brangwynne, 2013). Actin filaments associated with germinal vesicles of starfish oocytes facilitate nuclear envelope breakdown and form a contractile net that facilitates chromosome capture during mitosis (L´ en ´ art et al., 2005; Mori et al., 2014). Several studies have also implicated nuclear actin filaments in oocyte transcription (reviewed in Belin and Mullins, 2013). *For correspondence: Dyche.Mullins@ucsf.edu Competing interests: The authors declare that no competing interests exist. Funding: See page 19 Received: 27 March 2015 Accepted: 12 August 2015 Published: 19 August 2015 Reviewing editor: Pekka Lappalainen, University of Helsinki, Finland Copyright Belin et al. This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited. Belin et al. eLife 2015;4:e07735. DOI: 10.7554/eLife.07735 1 of 21