title: In silico synchronization reveals regulators of nuclear ruptures in lamin A/C deficient model cells.
abstract: The nuclear lamina is a critical regulator of nuclear structure and function. Nuclei from laminopathy patient cells experience repetitive disruptions of the nuclear envelope, causing transient intermingling of nuclear and cytoplasmic components. The exact causes and consequences of these events are not fully understood, but their stochastic occurrence complicates in-depth analyses. To resolve this, we have established a method that enables quantitative investigation of spontaneous nuclear ruptures, based on co-expression of a firmly bound nuclear reference marker and a fluorescent protein that shuttles between the nucleus and cytoplasm during ruptures. Minimally invasive imaging of both reporters, combined with automated tracking and in silico synchronization of individual rupture events, allowed extracting information on rupture frequency and recovery kinetics. Using this approach, we found that rupture frequency correlates inversely with lamin A/C levels, and can be reduced in genome-edited LMNA knockout cells by blocking actomyosin contractility or inhibiting the acetyl-transferase protein NAT10. Nuclear signal recovery followed a kinetic that is co-determined by the severity of the rupture event, and could be prolonged by knockdown of the ESCRT-III complex component CHMP4B. In conclusion, our approach reveals regulators of nuclear rupture induction and repair, which may have critical roles in disease development.
authors.affiliation: Laboratory of Cell Biology and Histology, Department of Veterinary Sciences, University of Antwerp, Antwerp, Belgium.
authors.forename: J
authors.initials: J
authors.lastname: Robijns
authors.affiliation: Laboratory of Cell Biology and Histology, Department of Veterinary Sciences, University of Antwerp, Antwerp, Belgium.
authors.forename: F
authors.initials: F
authors.lastname: Molenberghs
authors.affiliation: Laboratory of Cell Biology and Histology, Department of Veterinary Sciences, University of Antwerp, Antwerp, Belgium.|Cell Systems and Imaging Research Group (CSI), Department of Molecular Biotechnology, Ghent University, Ghent, Belgium.
authors.forename: T
authors.initials: T
authors.lastname: Sieprath
authors.affiliation: Laboratory of Cell Biology and Histology, Department of Veterinary Sciences, University of Antwerp, Antwerp, Belgium.|Cell Systems and Imaging Research Group (CSI), Department of Molecular Biotechnology, Ghent University, Ghent, Belgium.
authors.forename: T D J
authors.initials: TD
authors.lastname: Corne
authors.affiliation: Laboratory of Cell Biology and Histology, Department of Veterinary Sciences, University of Antwerp, Antwerp, Belgium.
authors.forename: M
authors.initials: M
authors.lastname: Verschuuren
authors.affiliation: Laboratory of Cell Biology and Histology, Department of Veterinary Sciences, University of Antwerp, Antwerp, Belgium.|Cell Systems and Imaging Research Group (CSI), Department of Molecular Biotechnology, Ghent University, Ghent, Belgium.
authors.forename: W H
authors.initials: WH
authors.lastname: De Vos
mesh_terms: D000205:Actomyosin / Q000378:metabolism; D045744:Cell Line, Tumor; D003198:Computer Simulation; D056827:Endosomal Sorting Complexes Required for Transport / Q000235:genetics / Q000378:metabolism; D006801:Humans; D034904:Lamin Type A / Q000235:genetics* / Q000378:metabolism; D063213:N-Terminal Acetyltransferase E / Q000235:genetics / Q000378:metabolism; D063206:N-Terminal Acetyltransferases; D009685:Nuclear Envelope / Q000378:metabolism* / Q000648:ultrastructure
chemical_list: C478397:CHMP4B protein, human; D056827:Endosomal Sorting Complexes Required for Transport; C494847:LMNA protein, human; D034904:Lamin Type A; D000205:Actomyosin; D063213:N-Terminal Acetyltransferase E; D063206:N-Terminal Acetyltransferases; C522989:NAT10 protein, human