Spatiotemporal Mislocalization of Nuclear Membrane-Associated Proteins in γ-Irradiation-Induced Senescent Cells.

Alena Svobodová Kovaříková,Emilie Lukášová,Eva Bártová,Aleš Kovařík

doi:10.3390/cells9040999

Alena Svobodová Kovaříková, Emilie Lukášová + Show 2 more

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https://doi.org/10.3390/cells9040999

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Abstract

Cellular senescence, induced by genotoxic or replication stress, is accompanied by defects in nuclear morphology and nuclear membrane-heterochromatin disruption. In this work, we analyzed cytological and molecular changes in the linker of nucleoskeleton and cytoskeleton (LINC) complex proteins in senescence triggered by γ-irradiation. We used human mammary carcinoma and osteosarcoma cell lines, both original and shRNA knockdown clones targeting lamin B receptor (LBR) and leading to LBR and lamin B (LB1) reduction. The expression status and integrity of LINC complex proteins (nesprin-1, SUN1, SUN2), lamin A/C, and emerin were analyzed by immunodetection using confocal microscopy and Western blot. The results show frequent mislocalization of these proteins from the nuclear membrane to cytoplasm and micronuclei and, in some cases, their fragmentation and amplification. The timing of these changes clearly preceded the onset of senescence. The LBR deficiency triggered neither senescence nor changes in the LINC protein distribution before irradiation. However, the cytological changes following irradiation were more pronounced in shRNA knockdown cells compared to original cell lines. We conclude that mislocalization of LINC complex proteins is a significant characteristic of cellular senescence phenotypes and may influence complex events at the nuclear membrane, including trafficking and heterochromatin attachment.

Highlights

The nuclear envelope in an interphase cell is anchored internally to the scaffold of the nucleoskeleton and chromatin and externally to the cytoskeleton
We recently found that DNA damage induced by γ-irradiation or replication stress (RS) in cancer cells leads to downregulation of the lamin B receptor (LBR) and lamin B1 (LB1)
While the effects of irradiation on DNA and chromatin topology are relatively well described, little is known about the impact of γ-radiation on nuclear membrane integrity and its protein components

Summary

Introduction

The nuclear envelope in an interphase cell is anchored internally to the scaffold of the nucleoskeleton and chromatin and externally to the cytoskeleton. The complex of proteins called the LINC (linker of nucleoskeleton and cytoskeleton) complex connects the perinuclear space with the nucleoskeleton and cytoskeleton [1,2,3,4]. This LINC complex plays a role in maintaining nuclear morphology, nuclear positioning, gene expression, and cell signaling [4,5,6,7].

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