Nucleoplasmic lamin C rapidly accumulates at sites of nuclear envelope rupture with BAF and cGAS

Abstract

In mammalian cell nuclei, the nuclear lamina (NL) underlies the nuclear envelope (NE) to maintain nuclear structure. The nuclear lamins, the major structural components of the NL, are involved in the protection against NE rupture induced by mechanical stress. However, the specific role of the lamins in repair of NE ruptures has not been fully determined. Our analyses using immunofluorescence and live-cell imaging revealed that the nucleoplasmic pool of lamin C rapidly accumulated at sites of NE rupture induced by laser microirradiation in mouse embryonic fibroblasts. The accumulation of lamin C at the rupture sites required both the immunoglobulin-like fold domain that binds to barrier-to-autointegration factor (BAF) and a nuclear localization signal. The accumulation of nuclear BAF and cytoplasmic cyclic GMP-AMP synthase (cGAS) at the rupture sites was in part dependent on lamin A/C. These results suggest that nucleoplasmic lamin C, BAF, and cGAS concertedly accumulate at sites of NE rupture for rapid repair.

Delayed localization of A-type lamins to the rupture sites in Hutchinson–Gilford progeria syndrome

“…1 C, D, F and G). Because LC rapidly accumulates at the rupture sites and the depletion slows the nuclear reentry of NLS-sfCherry at the early time points (within 10 min) (Kono et al ., 2022), PG expression could slow LC accumulation. To test this idea, endogenous LA, LC and PG were directly labeled with a LA/C-specific genetically encoded probe, designed ankyrin repeat protein (DARPin)-LaA_6 (Zwerger et al ., 2015), fused with super-folder GFP (sfGFP), sfGFP-DARPin-LA6 in WT and G609G/+ MEFs expressing NLS-sfCherry (Kono et al ., 2022) (Fig.…”

“…2 A). Because endogenous LA and PG did not localize to the rupture sites within 180 s after the induction of NE rupture (Kono et al ., 2022) (Fig. 1 B and C, Fig.…”

“…Because nucleoplasmic LC diffuses to the rupture sites (Kono et al ., 2022), the diffusion fraction could be decreased in G609G/+ cells compared to WT cells. To test this idea, the diffusion fraction and time of mEmerald-LC were quantitatively measured by fluorescence correlation spectroscopy (FCS) (Fig.…”

“…Recently, we found that LC but not LA was rapidly recruited from the nucleoplasm to sites of NE rupture induced by laser microirradiation along with BAF in immortalized MEFs (Kono et al ., 2022). However, the molecular mechanism by which LA and LC differ in localization/accumulation kinetics at the rupture sites still remains to be elucidated.…”

An ESCRT grommet cooperates with a diffusion barrier to maintain nuclear integrity

“…Another example might include the rapid recruitment of barrier to autointegration factor (BAF) to exposed chromatin at induced NE ruptures (Halfmann et al, 2019). As BAF forms a cross-linked network that can restrict the passage of macromolecules 500 kDa/49 nm in size (Samwer et al, 2017), it is possible BAF together with other factors (including lamin C (Kono et al, 2022)) might plug NE holes quickly before the membranes themselves are later sealed through a mechanism involving LEM protein(s) and ESCRTs. A similar function could be served by galectin, which appears capable of forming biomolecular condensates (Chiu et al, 2020) and recruiting ESCRT-III (Jia et al, 2020) to prevent leakage of lysosomal hydrolases during ESCRT repair of lysosome membranes (Radulovic et al, 2018; Skowyra et al, 2018).…”