Lamin C is required to establish genome organization after mitosis

Abstract

Background The dynamic 3D organization of the genome is central to gene regulation and development. The nuclear lamina influences genome organization through the tethering of lamina-associated domains (LADs) to the nuclear periphery. Evidence suggests that lamins A and C are the predominant lamins involved in the peripheral association of LADs, potentially serving different roles. Results Here, we examine chromosome architecture in mouse cells in which lamin A or lamin C are downregulated. We find that lamin C, and not lamin A, is required for the 3D organization of LADs and overall chromosome organization. Striking differences in localization are present as cells exit mitosis and persist through early G1 and are linked to differential phosphorylation. Whereas lamin A associates with the nascent nuclear envelope (NE) during telophase, lamin C remains in the interior, surrounding globular LAD aggregates enriched on euchromatic regions. Lamin C association with the NE is delayed until several hours into G1 and correlates temporally and spatially with the post-mitotic NE association of LADs. Post-mitotic LAD association with the NE, and global 3D genome organization, is perturbed only in cells depleted of lamin C, and not lamin A. Conclusions Lamin C regulates LAD dynamics during exit from mitosis and is a key regulator of genome organization in mammalian cells. This reveals an unexpectedly central role for lamin C in genome organization, including inter-chromosomal LAD-LAD segregation and LAD scaffolding at the NE, raising intriguing questions about the individual and overlapping roles of lamin A/C in cellular function and disease.

Nuclear lamin isoforms differentially contribute to LINC complex-dependent nucleocytoskeletal coupling and whole-cell mechanics

“…Therefore, we investigated the effects of deleting specific lamin isoforms on nuclear and cellular morphology in MEFs using our previously described homozygous lamin knockout (KO) MEF lines LA-/LC- (here abbreviated as LA/C-), LB1-, and LB2-, as well as their wild type (WT) MEF controls ( 12 ). Since recent studies suggest that LA and LC perform distinct roles in establishing the cellular mechanophenotype ( 13 – 15 ), we also probed the specific contributions of these alternative splice variants in MEFs stably expressing short hairpin RNAs that knockdown (KD) either endogenous LA (LA KD) or LC (LC KD) ( SI Appendix , Fig. S1 A and B ).…”

“…It has been shown that heterochromatin regulates small mechanical deformations of the nucleus ( 43 ). Given the small deformations (< 1 μm) inherent in the AFM and OT assays, we speculated that the observed decreases in nuclear stiffness in the lamin KO and KD MEFs may be, at least in part, related to changes in their levels of the heterochromatic markers H3K9me2, H3K9me3, and H3K27me3, which have been shown to be associated with nuclear stiffness ( 15 , 44 , 51 ). Therefore, we determined the levels of these heterochromatic markers in lamin KO and KD MEFs using quantitative immunofluorescence and Western blot analyses ( SI Appendix , Fig.…”

Mechanisms of A-Type Lamin Targeting to Nuclear Ruptures Are Disrupted in LMNA- and BANF1-Associated Progerias

“…To demonstrate if it was indeed the mobile population of A-type lamins that localizes to the ruptures, the nucleoplasmic pool of GFP-tagged A-type lamins was photobleached prior to rupture. This resulted in a loss of GFP-LaA accumulation at the rupture and greatly diminished the amount of the more mobile [ 75 , 78 ] GFP-LaC ( Figure 1 D). To visualize endogenous mobile A-type lamin behavior during a nuclear rupture, we mechanically-induced nuclear rupture on MCF10A cells via a cell compression chamber ( Supplementary Figure S3A ) and stained with 1E4, a mouse monoclonal antibody that preferentially recognizes nucleoplasmic A-type lamins [ 79 ].…”

“…To visualize endogenous mobile A-type lamin behavior during a nuclear rupture, we mechanically-induced nuclear rupture on MCF10A cells via a cell compression chamber ( Supplementary Figure S3A ) and stained with 1E4, a mouse monoclonal antibody that preferentially recognizes nucleoplasmic A-type lamins [ 79 ]. We also utilized an antibody that detects A-type lamins phosphorylated at serine-22 (LaA/C pS22), normally generated during early mitotic dissolution of the lamina, but also found in the nucleoplasm during interphase [ 78 ]. We found the mobile, nucleoplasmic pool of endogenous A-type lamins accumulate at rupture sites ( Supplementary Figure S3B ), and A-type lamins targeting to the rupture site are phosphorylated at Ser-22 ( Supplementary Figure S3C ).…”

Effects of mutant lamins on nucleo-cytoskeletal coupling in Drosophila models of LMNA muscular dystrophy

“…The mutant lamins that did not alter nuclear mechanics might play a role in gene expression. Genomes are rich with lamin-associated domains (LADs) in which sections of chromosomes are in close opposition with the lamin meshwork ( Mohanta et al, 2021 ; Wong et al, 2021 ). In fact, in some cell types, LADs represent up to half of the genome ( Briand & Collas, 2020 ; Mohanta et al, 2021 ; Rullens & Kind, 2021 ).…”