Nuclear actin filaments in DNA repair dynamics

Abstract

Recent development of innovative tools for live imaging of actin filaments (F-actin) enabled the detection of surprising nuclear structures responding to various stimuli, challenging previous models that actin is substantially monomeric in the nucleus. We review these discoveries, focusing on double-strand break (DSB) repair responses. These studies revealed a remarkable network of nuclear filaments and regulatory mechanisms coordinating chromatin dynamics with repair progression and led to a paradigm shift by uncovering the directed movement of repair sites.

Fig. 1 |. Nuclear actin polymerizes in response to several stimuli.

a, Different actin remodelling pathways are shown. Spontaneous actin nucleation is characterized by a fast-growing (+) ‘barbed’ end and a slow-growing (−) ‘pointed’ end, with more efficient addition of G-actin to the (+) end. F-actin formation and disassembly are regulated by actin remodellers, including nucleating, severing, capping, and crosslinking proteins. Arp2/3 promotes nucleation at 70° angles from preexisting filaments and is activated by the WAS family proteins (e.g., Wasp). Spire recruits several actin monomers with its WASP-homology2 domains (WH2), forming a seeding polymer for filament elongation. Formins associate with the (+) end and promote polymerization by bringing actin monomers in close proximity via formin homology 2 domains (FH2). Cofilin stimulates filament severing. CapZ associates with the (+) end, blocking G-actin access and filament elongation. Filamin holds two filaments together, promoting the formation of F-actin networks. b, Nuclear F-actin forms in response to different stimuli. DNA damage induces Arp2/3-dependent nuclear actin filament formation and formin-dependent filament formation for relocalization of heterochromatic DSBs and focus clustering, promoting repair^,,,. Serum stimulation, fibronectin treatment, or cell spreading, promotes MRTF-A activation through formin-dependent nuclear filaments^,. T-cell-receptor activation results in Arp2/3-dependent nuclear filaments promoting cytokine expression and antibody production. Baculoviruses can hijack the host system to produce Arp2/3-dependent filaments for nuclear egress. Cells entering G₁ experience formin-induced actin polymerization, promoting CenpA recruitment and replication initiation^,. G₁ nuclear filaments also mediate nuclear expansion.

Fig. 2 |. Model for the role of F-actin in DSB repair of heterochromatin and euchromatin.

In heterochromatin, DSB detection and processing (resection) occur inside the heterochromatin domain. Mre (MRN complex) and HP1a promote recruitment of Arp2/3 and myosins to DSBs; Arp2/3 activation by Scar and Wash facilitates actin polymerization and filament growth towards the nuclear periphery; Smc5/6 blocks Rad recruitment inside the heterochromatin domain and recruits Unc to activate nuclear myosins. The myosin-Smc5/6-chromatin complex translocates along actin filaments to anchor DSBs to nuclear pores or inner nuclear membrane proteins (INMPs, not shown), where HR repair continues with Rad51 recruitment and strand invasion. Actin filaments are highly dynamic and start disassembling during relocalization. In euchromatin, Mre11 and resection promote the movement of repair sites via Arp2/3 and F-actin, which in turn facilitate resection and HR repair. Actin polymers travel with euchromatic repair sites, possibly generating propelling forces for clustering.

Fig. 3 |. Different actin nucleators and motor proteins contribute to DSB dynamics and repair.

a, In Drosophila cells (that are mostly in S/G₂), directed motion of heterochromatic DSBs to the nuclear periphery relies on F-actin, Arp2/3, the Arp2/3 activators Scar and Wash, the myosin activator Unc45, and Myo1A, Myo1B, and MyoV nuclear myosins. Wasp, Whamy, Dia, and Spire are not required. Arp2/3, F-actin, Unc45 and myosins are also enriched at repair foci, consistent with a direct functionin repair. Clustering of euchromatic DSBs relies on Arp2/3 and not on Unc45^,. b, In mouse G₂ cells, relocalization of heterochromatic DSBs also requires Arp2/3, actin polymerization, and myosins. c, In human S/G₂ cells, dynamics of HR-prone DSBs depend on Arp2/3, Wasp, and F-actin, which are enriched at repair sites, whereas FMN-2 is not required.d, In human cells treated with MMS, actin filaments form in the nuclei and mediate repair, which also requires FMN-2 and Spire1/2, but not Dia1/29.e, In human G₁ cells, clustering of euchromatic DSBs requires FMN-2, and focus movement is not dependent on Arp2/3. (*) refers to experimental systems in which the nuclear function of the indicated components has been directly established. Actin filaments are indicated for studies that directly identified nuclear structures. Components that are not required for filament formation or repair in different contexts are in parenthesis. f, Schematic representation of MSD curves for different types of motion, as indicated (adapted with permission from ref.). g , Schematic representation of a focus track (adapted from ref.), showing mixed types of motion for heterochromatic repair foci that reach the nuclear periphery. Time points characterized by directed and subdiffusive motions are shown.