A high-content screen reveals new regulators of nuclear membrane stability.

Nuclear membrane rupture is a physiological response to multiple in vivo processes, such as cell migration, that can cause extensive genome instability and upregulate invasive and inflammatory pathways. However, the underlying molecular mechanisms of rupture are unclear and few regulators have been identified. In this study, we developed a reporter that is size excluded from re-compartmentalization following nuclear rupture events. This allows for robust detection of factors influencing nuclear integrity in fixed cells. We combined this with an automated image analysis pipeline in a high-content siRNA screen to identify new proteins that both increase and decrease nuclear rupture frequency in cancer cells. Pathway analysis identified an enrichment of nuclear membrane and ER factors in our hits and we demonstrate that one of these, the protein phosphatase CTDNEP1, is required for nuclear stability. Analysis of known rupture determinants, including an automated quantitative analysis of nuclear lamina gaps, are consistent with CTDNEP1 acting independently of actin and nuclear lamina organization. Our findings provide new insights into the molecular mechanism of nuclear rupture and define a highly adaptable program for rupture analysis that removes a substantial barrier to new discoveries in the field.

Mena regulates nesprin-2 to control actin-nuclear lamina associations, trans-nuclear membrane signalling and gene expression.

Interactions between cells and the extracellular matrix, mediated by integrin adhesion complexes, play key roles in fundamental cellular processes, including the sensing and transduction of mechanical cues. Here, we investigate systems-level changes in the integrin adhesome in patient-derived cutaneous squamous cell carcinoma cells and identify the actin regulatory protein Mena as a key node in the adhesion complex network. Mena is connected within a subnetwork of actin-binding proteins to the LINC complex component nesprin-2, with which it interacts and co-localises at the nuclear envelope. Moreover, Mena potentiates the interactions of nesprin-2 with the actin cytoskeleton and the nuclear lamina. CRISPR-mediated Mena depletion causes altered nuclear morphology, reduces tyrosine phosphorylation of the nuclear membrane protein emerin and downregulates expression of the immunomodulatory gene PTX3 via the recruitment of its enhancer to the nuclear periphery. We uncover an unexpected role for Mena at the nuclear membrane, where it controls nuclear architecture, chromatin repositioning and gene expression. Our findings identify an adhesion protein that regulates gene transcription via direct signalling across the nuclear envelope.

Negative correlation between the nuclear size and nuclear Lamina component Lamin A in intraductal papillary mucinous neoplasms of the pancreas.

Background: The nuclear laminar protein Lamin A and inner nuclear membrane protein Emerin plays important role in sustaining nuclear structure. However, They have not investigated the significance of these proteins for development of pancreatic intraductal papillary mucinous neoplasm (IPMN). Methods: We examined pancreatic IPMN specimens for nuclear morphology and nuclear protein expression pattern of Lamin A and Emerin. Forty-two IPMN specimens were included, with 30 classified as intraductal papillary mucinous adenoma (IPMA) and 12 as intraductal papillary mucinous carcinoma (IPMC). Results: Classification according to histological subtype revealed that 26 specimens were of the gastric subtype (1 IPMC case), 8 were pancreatobiliary (6 IPMC cases), 6 were intestinal (3 IPMC cases), and 2 were oncocytic (all cases were IPMC). The frequency of IPMN subtypes in this study seemed to agree with those in previous reports. We analyzed Feulgen staining sections for nuclear morphological analysis using computer-assisted image analysis. Nuclear area and perimeter were significantly larger in IPMC than in IPMA. Finally, we examined the positive ratios of Lamin A and Emerin in immunohistochemical staining sections by image analysis. We found a negative correlation between the nuclear size and Lamin A-positive ratio, which was significantly lower in IPMC than that in IPMA. However, no significant correlation was observed between nuclear size and Emerin expression was observed, and no differences were found in the Emerin-positive ratio between IPMA and IPMC. Conclusion: Our results suggest that a decreased Lamin A positive ratio induces nuclear enlargement in adenomas, which thereby induce promotion to carcinomas. Furthermore, Lamin A expression can be a reliable biomarker for distinguishing between IPMC and IPMA.

Dp71 Point Mutations Induce Protein Aggregation, Loss of Nuclear Lamina Integrity and Impaired Braf35 and Ibraf Function in Neuronal Cells.

Dystrophin Dp71 is the most abundant product of the Duchenne muscular dystrophy gene in the nervous system, and mutations impairing its function have been associated with the neurodevelopmental symptoms present in a third of DMD patients. Dp71 is required for the clustering of neurotransmitter receptors and the neuronal differentiation of cultured cells; nonetheless, its precise role in neuronal cells remains to be poorly understood. In this study, we analyzed the effect of two pathogenic DMD gene point mutations on the Dp71 function in neurons. We engineered C272Y and E299del mutations to express GFP-tagged Dp71 protein variants in N1E-115 and SH-SY5Y neuronal cells. Unexpectedly, the ectopic expression of Dp71 mutants resulted in protein aggregation, which may be mechanistically caused by the effect of the mutations on Dp71 structure, as predicted by protein modeling and molecular dynamics simulations. Interestingly, Dp71 mutant variants acquired a dominant negative function that, in turn, dramatically impaired the distribution of different Dp71 protein partners, including ß-dystroglycan, nuclear lamins A/C and B1, the high-mobility group (HMG)-containing protein (BRAF35) and the BRAF35-family-member inhibitor of BRAF35 (iBRAF). Further analysis of Dp71 mutants provided evidence showing a role for Dp71 in modulating both heterochromatin marker H3K9me2 organization and the neuronal genes' expression, via its interaction with iBRAF and BRAF5.

The Paradox of Nuclear Lamins in Pathologies: Apparently Controversial Roles Explained by Tissue-Specific Mechanobiology.

The nuclear lamina is a complex meshwork of intermediate filaments (lamins) that is located beneath the inner nuclear membrane and the surrounding nucleoplasm. The lamins exert both structural and functional roles in the nucleus and, by interacting with several nuclear proteins, are involved in a wide range of nuclear and cellular activities. Due their pivotal roles in basic cellular processes, lamin gene mutations, or modulations in lamin expression, are often associated with pathological conditions, ranging from rare genetic diseases, such as laminopathies, to cancer. Although a substantial amount of literature describes the effects that are mediated by the deregulation of nuclear lamins, some apparently controversial results have been reported, which may appear to conflict with each other. In this context, we herein provide our explanation of such "controversy", which, in our opinion, derives from the tissue-specific expression of nuclear lamins and their close correlation with mechanotransduction processes, which could be very different, or even opposite, depending on the specific mechanical conditions that should not be compared (a tissue vs. another tissue, in vivo studies vs. cell cultures on glass/plastic supports, etc.). Moreover, we have stressed the relevance of considering and reproducing the "mechano-environment" in in vitro experimentation. Indeed, when primary cells that are collected from patients or donors are maintained in a culture, the mechanical signals deriving from canonical experimental procedures of cell culturing could alter the lamin expression, thereby profoundly modifying the assessed cell type, in some cases even too much, compared to the cell of origin.

CTCF supports preferentially short lamina-associated domains.

More than one third of the mammalian genome is in a close association with the nuclear lamina, thus these genomic regions were termed lamina-associated domains (LADs). This association is fundamental for many aspects of chromatin biology including transcription, replication, and DNA damage repair. LADs association with the nuclear envelope is thought to be dependent on two major mechanisms: The first mechanism is the interaction between nuclear membrane proteins such as LBR with heterochromatin modifications that are enriched in LADs chromatin. The second mechanism is based on proteins that bind the borders of the LADs and support the association of the LADs with the nuclear envelope. Two factors were suggested to support the second mechanism: CCCTC-binding factor (CTCF) and YY1 based on their enriched binding to LADs borders. However, this mechanism has not been proven yet at a whole genome level. Here, to test if CTCF supports the LADs landscape, we generated melanoma cells with a partial loss of function (pLoF) of CTCF by the CRISPR-Cas9 system and determined the LADs landscape by lamin B ChIP-seq analysis. We found that under regular growth conditions, CTCF pLoF led to modest changes in the LADs landscape that included an increase in the signal of 2% of the LADs and a decrease in the signal of 8% of the LADs. However, CTCF importance for the LADs landscape was much higher upon induction of a chromatin stress. We induced chromatin stress by inhibiting RNA polymerase II, an intervention that is known to alter chromatin compaction and supercoiling. Notably, only in CTCF pLoF cells, the chromatin stress led to the dissociation of 7% of the LADs from the lamina. The CTCF-dependent LADs had almost three times shorter median length than the non-affected LADs, were enriched in CTCF binding at their borders, and were higher in their facultative-status (cell-type specific). Thus, it appears that CTCF is a key factor in facilitating the association of short facultative LADs with the nuclear lamina upon chromatin stress.

Gaussian curvature dilutes the nuclear lamina, favoring nuclear rupture, especially at high strain rate.

Nuclear rupture has long been associated with deficits or defects in lamins, with recent results also indicating a role for actomyosin stress, but key physical determinants of rupture remain unclear. Here, lamin-B filaments stably interact with the nuclear membrane at sites of low Gaussian curvature yet dilute at high curvature to favor rupture, whereas lamin-A depletion requires high strain-rates. Live-cell imaging of lamin-B1 gene-edited cancer cells is complemented by fixed-cell imaging of rupture in: iPS-derived progeria patients cells, cells within beating chick embryo hearts, and cancer cells with multi-site rupture after migration through small pores. Data fit a model of stiff filaments that detach from a curved surface.Rupture is modestly suppressed by inhibiting myosin-II and by hypotonic stress, which slow the strain-rates. Lamin-A dilution and rupture probability indeed increase above a threshold rate of nuclear pulling. Curvature-sensing mechanisms of proteins at plasma membranes, including Piezo1, might thus apply at nuclear membranes.Summary statement: High nuclear curvature drives lamina dilution and nuclear envelope rupture even when myosin stress is inhibited. Stiff filaments generally dilute from sites of high Gaussian curvature, providing mathematical fits of experiments.

Nuclear Cytoskeleton in Virus Infection.

The nuclear lamina is the main component of the nuclear cytoskeleton that maintains the integrity of the nucleus. However, it represents a natural barrier for viruses replicating in the cell nucleus. The lamina blocks viruses from being trafficked to the nucleus for replication, but it also impedes the nuclear egress of the progeny of viral particles. Thus, viruses have evolved mechanisms to overcome this obstacle. Large viruses induce the assembly of multiprotein complexes that are anchored to the inner nuclear membrane. Important components of these complexes are the viral and cellular kinases phosphorylating the lamina and promoting its disaggregation, therefore allowing virus egress. Small viruses also use cellular kinases to induce lamina phosphorylation and the subsequent disruption in order to facilitate the import of viral particles during the early stages of infection or during their nuclear egress. Another component of the nuclear cytoskeleton, nuclear actin, is exploited by viruses for the intranuclear movement of their particles from the replication sites to the nuclear periphery. This study focuses on exploitation of the nuclear cytoskeleton by viruses, although this is just the beginning for many viruses, and promises to reveal the mechanisms and dynamic of physiological and pathological processes in the nucleus.

Shear stress regulates the migration of suspended breast cancer cells by nuclear lamina protein A/C and large tumor suppressor through yes-associated protein.

Breast cancer is life threatening among women because its migration by hematogenous metastasis, where, besides biochemical cues, breast circulating tumor cells (CTCs) expose to suspension state and shear stress. However, the combined effects of these mechanical factors on CTCs migration were unclear. Here, suspension state and shear stress were loaded to breast tumor cells (BTCs) to mimic two mechanical cues in the mechanical environment of breast CTCs and the mechanobiological mechanism of suspension state and shear stress regulating the migration of (BTCs) was investigated. The migration and nuclear lamina protein A/C (Lamin A/C) accumulation were enhanced in MDA-MB-231 and SK-BR-3 BTCs exposed to shear stress though lower than that of suspended cells with different yes-associated protein (YAP) subcellular localization. Knockdown of LMNA downregulated and upregulated YAP targets in suspended BTCs and BTCs exposed to shear stress, respectively, which inhibited MDA-MB-231 BTCs migration in vitro and in vivo. Large tumor suppressor (LATS) responded to suspension state and shear stress, knockdown of which decreased the migration of MDA-MB-231 BTCs. These findings uncover the mechanobiological mechanism that suspension state and shear stress antagonistically promote BTCs migration by Lamin A/C and LATS through YAP and the potential for targeting YAP in CTCs prognosis. Shear stress regulates suspended breast cancer cells migration by Lamin A/C and LATS through YAP.

Aberrant nuclear lamina contributes to the malignancy of human gliomas.

Glioma is the most common type of tumor in the central nervous system, accounting for around 80% of all malignant brain tumors. Previous studies showed a significant association between nuclear morphology and the malignant progress of gliomas. By virtue of integrated proteomics and genomics analyses as well as experimental validations, we identify three nuclear lamin genes (LMNA, LMNB1, and LMNB2) that are significantly upregulated in glioma tissues compared with normal brain tissues. We show that elevated expressions of LMNB1, LMNB2, and LMNA in glioma cells are highly associated with the rapid progression of the disease and the knockdown of LMNB1, LMNB2, and LMNA dramatically suppresses glioma progression in both in vitro and in vivo mouse models. Moreover, the repression of glioma cell growth by lamin knockdown is mediated by the pRb-mediated G1-S inhibition. On the contrary, overexpression of lamins in normal human astrocytes dramatically induced nuclear morphological aberrations and accelerated cell growth. Together, our multi-omics-based analysis has revealed a previously unrecognized role of lamin genes in gliomagenesis, providing a strong support for the key link between aberrant tumor nuclear shape and the survival of glioma patients. Based on these findings, lamins are proposed to be potential oncogene targets for therapeutic treatments of brain tumors.

Regulatory role of cathepsin L in induction of nuclear laminopathy in Alzheimer's disease.

Experimental and clinical therapies in the field of Alzheimer's disease (AD) have focused on elimination of extracellular amyloid beta aggregates or prevention of cytoplasmic neuronal fibrillary tangles formation, yet these approaches have been generally ineffective. Interruption of nuclear lamina integrity, or laminopathy, is a newly identified concept in AD pathophysiology. Unraveling the molecular players in the induction of nuclear lamina damage may lead to identification of new therapies. Here, using 3xTg and APP/PS1 mouse models of AD, and in vitro model of amyloid beta42 (Aß42) toxicity in primary neuronal cultures and SH-SY5Y neuroblastoma cells, we have uncovered a key role for cathepsin L in the induction of nuclear lamina damage. The applicability of our findings to AD pathophysiology was validated in brain autopsy samples from patients. We report that upregulation of cathepsin L is an important process in the induction of nuclear lamina damage, shown by lamin B1 cleavage, and is associated with epigenetic modifications in AD pathophysiology. More importantly, pharmacological targeting and genetic knock out of cathepsin L mitigated Aß42 induced lamin B1 degradation and downstream structural and molecular changes. Affirming these findings, overexpression of cathepsin L alone was sufficient to induce lamin B1 cleavage. The proteolytic activity of cathepsin L on lamin B1 was confirmed using mass spectrometry. Our research identifies cathepsin L as a newly identified lamin B1 protease and mediator of laminopathy observed in AD. These results uncover a new aspect in the pathophysiology of AD that can be pharmacologically prevented, raising hope for potential therapeutic interventions.

Lamin A as a Determinant of Mechanical Properties of the Cell Nucleus in Health and Disease.

One of the main factors associated with worse prognosis in oncology is metastasis, which is based on the ability of tumor cells to migrate from the primary source and to form secondary tumors. The search for new strategies to control migration of metastatic cells is one of the urgent issues in biomedicine. One of the strategies to stop spread of cancer cells could be regulation of the nuclear elasticity. Nucleus, as the biggest and stiffest cellular compartment, determines mechanical properties of the cell as a whole, and, hence, could prevent cell migration through the three-dimensional extracellular matrix. Nuclear rigidity is maintained by the nuclear lamina, two-dimensional network of intermediate filaments in the inner nuclear membrane (INM). Here we present the most significant factors defining nucleus rigidity, discuss the role of nuclear envelope composition in the cell migration, as well consider possible approaches to control lamina composition in order to change plasticity of the cell nucleus and ability of the tumor cells to metastasize.

Characterization of subcellular localization of eukaryotic clamp loader/unloader and its regulatory mechanism.

Proliferating cell nuclear antigen (PCNA) plays a critical role as a processivity clamp for eukaryotic DNA polymerases and a binding platform for many DNA replication and repair proteins. The enzymatic activities of PCNA loading and unloading have been studied extensively in vitro. However, the subcellular locations of PCNA loaders, replication complex C (RFC) and CTF18-RFC-like-complex (RLC), and PCNA unloader ATAD5-RLC remain elusive, and the role of their subunits RFC2-5 is unknown. Here we used protein fractionation to determine the subcellular localization of RFC and RLCs and affinity purification to find molecular requirements for the newly defined location. All RFC/RLC proteins were detected in the nuclease-resistant pellet fraction. RFC1 and ATAD5 were not detected in the non-ionic detergent-soluble and nuclease-susceptible chromatin fractions, independent of cell cycle or exogenous DNA damage. We found that small RFC proteins contribute to maintaining protein levels of the RFC/RLCs. RFC1, ATAD5, and RFC4 co-immunoprecipitated with lamina-associated polypeptide 2 (LAP2) α which regulates intranuclear lamin A/C. LAP2α knockout consistently reduced detection of RFC/RLCs in the pellet fraction, while marginally affecting total protein levels. Our findings strongly suggest that PCNA-mediated DNA transaction occurs through regulatory machinery associated with nuclear structures, such as the nuclear matrix.

Mapping the micro-proteome of the nuclear lamina and lamina-associated domains.

The nuclear lamina is a proteinaceous network of filaments that provide both structural and gene regulatory functions by tethering proteins and large domains of DNA, the so-called lamina-associated domains (LADs), to the periphery of the nucleus. LADs are a large fraction of the mammalian genome that are repressed, in part, by their association to the nuclear periphery. The genesis and maintenance of LADs is poorly understood as are the proteins that participate in these functions. In an effort to identify proteins that reside at the nuclear periphery and potentially interact with LADs, we have taken a two-pronged approach. First, we have undertaken an interactome analysis of the inner nuclear membrane bound LAP2ß to further characterize the nuclear lamina proteome. To accomplish this, we have leveraged the BioID system, which previously has been successfully used to characterize the nuclear lamina proteome. Second, we have established a system to identify proteins that bind to LADs by developing a chromatin-directed BioID system. We combined the BioID system with the m6A-tracer system which binds to LADs in live cells to identify both LAD proximal and nuclear lamina proteins. In combining these datasets, we have further characterized the protein network at the nuclear lamina, identified putative LAD proximal proteins and found several proteins that appear to interface with both micro-proteomes. Importantly, several proteins essential for LAD function, including heterochromatin regulating proteins related to H3K9 methylation, were identified in this study.

Lamin B1 acetylation slows the G1 to S cell cycle transition through inhibition of DNA repair.

The integrity and regulation of the nuclear lamina is essential for nuclear organization and chromatin stability, with its dysregulation being linked to laminopathy diseases and cancer. Although numerous posttranslational modifications have been identified on lamins, few have been ascribed a regulatory function. Here, we establish that lamin B1 (LMNB1) acetylation at K134 is a molecular toggle that controls nuclear periphery stability, cell cycle progression, and DNA repair. LMNB1 acetylation prevents lamina disruption during herpesvirus type 1 (HSV-1) infection, thereby inhibiting virus production. We also demonstrate the broad impact of this site on laminar processes in uninfected cells. LMNB1 acetylation negatively regulates canonical nonhomologous end joining by impairing the recruitment of 53BP1 to damaged DNA. This defect causes a delay in DNA damage resolution and a persistent activation of the G1/S checkpoint. Altogether, we reveal LMNB1 acetylation as a mechanism for controlling DNA repair pathway choice and stabilizing the nuclear periphery.

Akt1-associated actomyosin remodelling is required for nuclear lamina dispersal and nuclear shrinkage in epidermal terminal differentiation.

Keratinocyte cornification and epidermal barrier formation are tightly controlled processes, which require complete degradation of intracellular organelles, including removal of keratinocyte nuclei. Keratinocyte nuclear destruction requires Akt1-dependent phosphorylation and degradation of the nuclear lamina protein, Lamin A/C, essential for nuclear integrity. However, the molecular mechanisms that result in complete nuclear removal and their regulation are not well defined. Post-confluent cultures of rat epidermal keratinocytes (REKs) undergo spontaneous and complete differentiation, allowing visualisation and perturbation of the differentiation process in vitro. We demonstrate that there is dispersal of phosphorylated Lamin A/C to structures throughout the cytoplasm in differentiating keratinocytes. We show that the dispersal of phosphorylated Lamin A/C is Akt1-dependent and these structures are specific for the removal of Lamin A/C from the nuclear lamina; nuclear contents and Lamin B were not present in these structures. Immunoprecipitation identified a group of functionally related Akt1 target proteins involved in Lamin A/C dispersal, including actin, which forms cytoskeletal microfilaments, Arp3, required for actin filament nucleation, and Myh9, a component of myosin IIa, a molecular motor that can translocate along actin filaments. Disruption of actin filament polymerisation, nucleation or myosin IIa activity prevented formation and dispersal of cytoplasmic Lamin A/C structures. Live imaging of keratinocytes expressing fluorescently tagged nuclear proteins showed a nuclear volume reduction step taking less than 40 min precedes final nuclear destruction. Preventing Akt1-dependent Lamin A/C phosphorylation and disrupting cytoskeletal Akt1-associated proteins prevented nuclear volume reduction. We propose keratinocyte nuclear destruction and differentiation requires myosin II activity and the actin cytoskeleton for two intermediate processes: Lamin A/C dispersal and rapid nuclear volume reduction.

Structural and Mechanical Aberrations of the Nuclear Lamina in Disease.

The nuclear lamins are the major components of the nuclear lamina in the nuclear envelope. Lamins are involved in numerous functions, including a role in providing structural support to the cell and the mechanosensing of the cell. Mutations in the genes encoding for lamins lead to the rare diseases termed laminopathies. However, not only laminopathies show alterations in the nuclear lamina. Deregulation of lamin expression is reported in multiple cancers and several viral infections lead to a disrupted nuclear lamina. The structural and mechanical effects of alterations in the nuclear lamina can partly explain the phenotypes seen in disease, such as muscular weakness in certain laminopathies and transmigration of cancer cells. However, a lot of answers to questions about the relation between changes in the nuclear lamina and disease development remain elusive. Here, we review the current understandings of the contribution of the nuclear lamina in the structural support and mechanosensing of healthy and diseased cells.

TOPORS, a tumor suppressor protein, contributes to the maintenance of higher-order chromatin architecture.

In the nucleus, chromosomes are hierarchically folded into active (A) and inactive (B) compartments composed of topologically associating domains (TADs). Genomic regions interact with nuclear lamina, termed lamina-associated domains (LADs). However, the molecular mechanisms underlying these 3D chromatin architectures remain incompletely understood. Here, we investigated the role of a potential tumor suppressor, TOP1 Binding Arginine/Serine Rich Protein (TOPORS), in genome organization. In mouse hepatocytes, chromatin interactions between A and B compartments increase and compartmentalization strength is reduced significantly upon Topors knockdown. Correspondingly, strength of TAD boundaries located at A/B borders is weakened. In the absence of TOPORS, chromatin-lamina interactions decrease and the coverage of LADs reduces from 53.31% to 46.52%. Interestingly, these changes in 3D genome are associated with PML nuclear bodies and PML-associated domains (PADs). Moreover, chromatin accessibility is altered predominantly at intergenic regions upon Topors knockdown, including a subset of enhancers. These alterations of chromatin are concordant with transcriptome changes, which are associated with carcinogenesis. Collectively, our findings demonstrate that TOPORS functions as a regulator in chromatin structure, providing novel insight into the architectural roles of tumor suppressors in higher-order genome organization.

Actin assembly ruptures the nuclear envelope by prying the lamina away from nuclear pores and nuclear membranes in starfish oocytes.

The nucleus of oocytes (germinal vesicle) is unusually large and its nuclear envelope (NE) is densely packed with nuclear pore complexes (NPCs) that are stockpiled for embryonic development. We showed that breakdown of this specialized NE is mediated by an Arp2/3-nucleated F-actin 'shell' in starfish oocytes, in contrast to microtubule-driven tearing in mammalian fibroblasts. Here, we address the mechanism of F-actin-driven NE rupture by correlated live-cell, super-resolution and electron microscopy. We show that actin is nucleated within the lamina, sprouting filopodia-like spikes towards the nuclear membranes. These F-actin spikes protrude pore-free nuclear membranes, whereas the adjoining stretches of membrane accumulate NPCs that are associated with the still-intact lamina. Packed NPCs sort into a distinct membrane network, while breaks appear in ER-like, pore-free regions. We reveal a new function for actin-mediated membrane shaping in nuclear rupture that is likely to have implications in other contexts, such as nuclear rupture observed in cancer cells.

Lamin A/C promotes DNA base excision repair.

The A-type lamins (lamin A/C), encoded by the LMNA gene, are important structural components of the nuclear lamina. LMNA mutations lead to degenerative disorders known as laminopathies, including the premature aging disease Hutchinson-Gilford progeria syndrome. In addition, altered lamin A/C expression is found in various cancers. Reports indicate that lamin A/C plays a role in DNA double strand break repair, but a role in DNA base excision repair (BER) has not been described. We provide evidence for reduced BER efficiency in lamin A/C-depleted cells (Lmna null MEFs and lamin A/C-knockdown U2OS). The mechanism involves impairment of the APE1 and POLß BER activities, partly effectuated by associated reduction in poly-ADP-ribose chain formation. Also, Lmna null MEFs displayed reduced expression of several core BER enzymes (PARP1, LIG3 and POLß). Absence of Lmna led to accumulation of 8-oxoguanine (8-oxoG) lesions, and to an increased frequency of substitution mutations induced by chronic oxidative stress including GC>TA transversions (a fingerprint of 8-oxoG:A mismatches). Collectively, our results provide novel insights into the functional interplay between the nuclear lamina and cellular defenses against oxidative DNA damage, with implications for cancer and aging.