scholarly journals Impaired lamin localization to the nuclear envelope is responsible for nuclear damage in LMNA mutant iPSC-derived cardiomyocytes

2021 ◽  
Author(s):  
Melanie Maurer ◽  
Shriya Perati ◽  
Lindsey E. Johnson ◽  
Anthony M. Gacita ◽  
Shuping Lai ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Intracellular Signaling ◽  
Mechanical Stability ◽  
Induced Pluripotent Stem Cell ◽  
Nuclear Lamina ◽  
Lamin A ◽  
Nuclear Abnormalities ◽  
Healthy Control ◽  
Induced Pluripotent ◽  
Cellular Dysfunction

The LMNA gene encodes the nuclear envelope proteins Lamins A and C, which comprise a major part of the nuclear lamina, provide mechanical support to the nucleus, and participate in diverse intracellular signaling. LMNA mutations give rise to a collection of diseases called laminopathies, including dilated cardiomyopathy (LMNA-DCM) and muscular dystrophies. Although nuclear deformities are a hallmark of LMNA-DCM, the role of nuclear abnormalities in the pathogenesis of LMNA-DCM remains incompletely understood. Using induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) from LMNA mutant patients and healthy controls, we show that LMNA mutant iPSC-CM nuclei have altered shape or increased size compared to healthy control iPSC-CM nuclei. The LMNA mutation exhibiting the most severe nuclear deformities, R249Q, additionally caused reduced nuclear stiffness and increased nuclear fragility. Importantly, for all cell lines, the degree of nuclear abnormalities corresponded to the degree of Lamin A/C and Lamin B1 mislocalization from the nuclear envelope. The mislocalization was likely due to altered assembly of Lamin A/C. Collectively, these results point to the importance of correct lamin assembly at the nuclear envelope in providing mechanical stability to the nucleus and illustrate that defects in nuclear lamina organization can contribute to the nuclear and cellular dysfunction in LMNA-DCM.

scholarly journals Chromatin compartment dynamics in a haploinsufficient model of cardiac laminopathy

2019 ◽  
Vol 218 (9) ◽  
pp. 2919-2944 ◽  
Author(s):  
Alessandro Bertero ◽  
Paul A. Fields ◽  
Alec S.T. Smith ◽  
Andrea Leonard ◽  
Kevin Beussman ◽  
...  
Keyword(s):  
Gene Expression ◽  
Large Scale ◽  
Induced Pluripotent Stem Cell ◽  
Nuclear Lamina ◽  
Lamin A ◽  
Chromosome Conformation ◽  
Nuclear Lamins ◽  
Genome Wide ◽  
Induced Pluripotent ◽  
Gene Expression Alterations

Mutations in A-type nuclear lamins cause dilated cardiomyopathy, which is postulated to result from dysregulated gene expression due to changes in chromatin organization into active and inactive compartments. To test this, we performed genome-wide chromosome conformation analyses in human induced pluripotent stem cell–derived cardiomyocytes (hiPSC-CMs) with a haploinsufficient mutation for lamin A/C. Compared with gene-corrected cells, mutant hiPSC-CMs have marked electrophysiological and contractile alterations, with modest gene expression changes. While large-scale changes in chromosomal topology are evident, differences in chromatin compartmentalization are limited to a few hotspots that escape segregation to the nuclear lamina and inactivation during cardiogenesis. These regions exhibit up-regulation of multiple noncardiac genes including CACNA1A, encoding for neuronal P/Q-type calcium channels. Pharmacological inhibition of the resulting current partially mitigates the electrical alterations. However, chromatin compartment changes do not explain most gene expression alterations in mutant hiPSC-CMs. Thus, global errors in chromosomal compartmentation are not the primary pathogenic mechanism in heart failure due to lamin A/C haploinsufficiency.

scholarly journals Compromised Barrier Function in Human Induced Pluripotent Stem-Cell-Derived Retinal Pigment Epithelial Cells from Type 2 Diabetic Patients

2019 ◽  
Vol 20 (15) ◽  
pp. 3773 ◽  
Author(s):  
Mostafa Kiamehr ◽  
Alexa Klettner ◽  
Elisabeth Richert ◽  
Ali Koskela ◽  
Arto Koistinen ◽  
...  
Keyword(s):  
Pigment Epithelium ◽  
Retinal Pigment ◽  
Induced Pluripotent Stem Cell ◽  
Diabetic Patients ◽  
Rpe Cells ◽  
Normal Glucose ◽  
Healthy Control ◽  
Induced Pluripotent ◽  
Type 2 Diabetic

In diabetic patients, high blood glucose induces alterations in retinal function and can lead to visual impairment due to diabetic retinopathy. In immortalized retinal pigment epithelial (RPE) cultures, high glucose concentrations are shown to lead to impairment in epithelial barrier properties. For the first time, the induced pluripotent stem-cell-derived retinal pigment epithelium (hiPSC-RPE) cell lines derived from type 2 diabetics and healthy control patients were utilized to assess the effects of glucose concentration on the cellular functionality. We show that both type 2 diabetic and healthy control hiPSC-RPE lines differentiate and mature well, both in high and normal glucose concentrations, express RPE specific genes, secrete pigment epithelium derived factor, and form a polarized cell layer. Here, type 2 diabetic hiPSC-RPE cells had a decreased barrier function compared to controls. Added insulin increased the epithelial cell layer tightness in normal glucose concentrations, and the effect was more evident in type 2 diabetics than in healthy control hiPSC-RPE cells. In addition, the preliminary functionality assessments showed that type 2 diabetic hiPSC-RPE cells had attenuated autophagy detected via ubiquitin-binding protein p62/Sequestosome-1 (p62/SQSTM1) accumulation, and lowered pro- matrix metalloproteinase 2 (proMMP2) as well as increased pro-MMP9 secretion. These results suggest that the cellular ability to tolerate stress is possibly decreased in type 2 diabetic RPE cells.

scholarly journals Physiological and pathological roles of LRRK2 in the nuclear envelope integrity

2019 ◽  
Vol 28 (23) ◽  
pp. 3982-3996 ◽  
Author(s):  
Vered Shani ◽  
Hazem Safory ◽  
Raymonde Szargel ◽  
Ninghan Wang ◽  
Tsipora Cohen ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Cortical Neurons ◽  
Nuclear Lamina ◽  
Lamin A ◽  
Loss Of Function ◽  
Function Mechanism ◽  
Disease Mutations ◽  
Lrrk2 G2019s ◽  
Sporadic Parkinson’S Disease ◽  
Seven In Absentia

Abstract Mutations in LRRK2 cause autosomal dominant and sporadic Parkinson’s disease, but the mechanisms involved in LRRK2 toxicity in PD are yet to be fully understood. We found that LRRK2 translocates to the nucleus by binding to seven in absentia homolog (SIAH-1), and in the nucleus it directly interacts with lamin A/C, independent of its kinase activity. LRRK2 knockdown caused nuclear lamina abnormalities and nuclear disruption. LRRK2 disease mutations mostly abolish the interaction with lamin A/C and, similar to LRRK2 knockdown, cause disorganization of lamin A/C and leakage of nuclear proteins. Dopaminergic neurons of LRRK2 G2019S transgenic and LRRK2 −/− mice display decreased circularity of the nuclear lamina and leakage of the nuclear protein 53BP1 to the cytosol. Dopaminergic nigral and cortical neurons of both LRRK2 G2019S and idiopathic PD patients exhibit abnormalities of the nuclear lamina. Our data indicate that LRRK2 plays an essential role in maintaining nuclear envelope integrity. Disruption of this function by disease mutations suggests a novel phosphorylation-independent loss-of-function mechanism that may synergize with other neurotoxic effects caused by LRRK2 mutations.

Prelamin A-mediated nuclear envelope dynamics in normal and laminopathic cells

2011 ◽  
Vol 39 (6) ◽  
pp. 1698-1704 ◽  
Author(s):  
Giovanna Lattanzi
Keyword(s):  
Nuclear Envelope ◽  
Accelerated Aging ◽  
Nuclear Lamina ◽  
Major Constituent ◽  
Lamin A ◽  
Post Translational Modifications ◽  
Prelamin A ◽  
Chromatin Dynamics ◽  
Chromatin Arrangement ◽  
And Function

Prelamin A is the precursor protein of lamin A, a major constituent of the nuclear lamina in higher eukaryotes. Increasing attention to prelamin A processing and function has been given after the discovery, from 2002 to 2004, of diseases caused by prelamin A accumulation. These diseases, belonging to the group of laminopathies and mostly featuring LMNA mutations, are characterized, at the clinical level, by different degrees of accelerated aging, and adipose tissue, skin and bone abnormalities. The outcome of studies conducted in the last few years consists of three major findings. First, prelamin A is processed at different rates under physiological conditions depending on the differentiation state of the cell. This means that, for instance, in muscle cells, prelamin A itself plays a biological role, besides production of mature lamin A. Secondly, prelamin A post-translational modifications give rise to different processing intermediates, which elicit different effects in the nucleus, mostly by modification of the chromatin arrangement. Thirdly, there is a threshold of toxicity, especially of the farnesylated form of prelamin A, whose accumulation is obviously linked to cell and organism senescence. The present review is focused on prelamin A-mediated nuclear envelope modifications that are upstream of chromatin dynamics and gene expression mechanisms regulated by the lamin A precursor.

scholarly journals Keratins couple with the nuclear lamina and regulate proliferation in colonic epithelial cells

2020 ◽  
Author(s):  
Carl-Gustaf A. Stenvall ◽  
Joel H. Nyström ◽  
Ciarán Butler-Hallissey ◽  
Stephen A. Adam ◽  
Roland Foisner ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Nuclear Structure ◽  
Mechanical Stability ◽  
Nuclear Lamina ◽  
Lamin A ◽  
Intestinal Epithelial ◽  
Linc Complex ◽  
Colonic Inflammation ◽  
Associated Proteins

AbstractKeratin intermediate filaments (IFs) convey mechanical stability and protection against stress to epithelial cells, and may participate in nuclear structure and organization. Keratins are important for colon health as observed in keratin 8 knockout (K8−/−) mice, which exhibit colonic inflammation and epithelial hyperproliferation. Here, using a full body and two intestinal epithelial-specific K8−/− knockout mouse models, we determine if cytoplasmic keratins affect the nuclear structure and lamina in epithelial colonocytes. K8−/− colonocytes in vivo and in organoid cultures exhibit significantly decreased levels of the major lamins A/C, B1 and B2 in a colon-specific and cell-intrinsic manner independent of major changes in colonic inflammation or microbiota. Downregulation of K8 by siRNA in Caco-2 cells similarly decreases lamin A levels, which recover after re-expression of K8. K8 loss is associated with reduced plectin, LINC complex proteins and lamin-associated proteins, indicating a dysfunctional keratin-nuclear lamina coupling. Immunoprecipitation identifies complexes of colonocyte keratins with the LINC protein SUN2 and lamin A. Hyperphosphorylation of the lamin A-associated cell cycle regulator pRb in K8−/− colonocytes together with increased nuclear localization of the mechanosensor YAP provide a molecular mechanism for the hyperproliferation phenotype. These findings identify a novel, colonocyte-specific role for K8 in nuclear function.

scholarly journals Chromatin compartment dynamics in a haploinsufficient model of cardiac laminopathy

2019 ◽  
Author(s):  
Alessandro Bertero ◽  
Paul A. Fields ◽  
Alec S. T. Smith ◽  
Andrea Leonard ◽  
Kevin Beussman ◽  
...  
Keyword(s):  
Gene Expression ◽  
Large Scale ◽  
Induced Pluripotent Stem Cell ◽  
Lamin A ◽  
Chromosome Conformation ◽  
Human Cardiomyocytes ◽  
Nuclear Lamins ◽  
Genome Wide ◽  
Induced Pluripotent ◽  
Gene Expression Alterations

AbstractPathogenic mutations in A-type nuclear lamins cause dilated cardiomyopathy, which is postulated to result from dysregulated gene expression due to changes in chromatin organization into active and inactive compartments. To test this, we performed genome-wide chromosome conformation analyses (Hi-C) in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) with a haploinsufficient mutation for lamin A/C. Compared to gene-corrected cells, mutant hiPSC-CMs have marked electrophysiological and contractile alterations, with modest gene expression changes. While large-scale changes in chromosomal topology are evident, differences in chromatin compartmentalization are limited to a few hotspots that escape inactivation during cardiogenesis. These regions exhibit upregulation of multiple non-cardiac genes including CACNA1A, encoding for neuronal P/Q-type calcium channels. Pharmacological inhibition of the resulting current partially mitigates the electrical alterations. On the other hand, A/B compartment changes do not explain most gene expression alterations in mutant hiPSC-CMs. We conclude that global errors in chromosomal compartmentation are not the primary pathogenic mechanism in heart failure due to lamin A/C haploinsufficiency.SummaryBertero et al. observe that lamin A/C haploinsufficiency in human cardiomyocytes markedly alters electrophysiology, contractility, gene expression, and chromosomal topology. Contrary to expectations, however, changes in chromatin compartments involve just few regions, and most dysregulated genes lie outside these hotspots.Condensed titleGenomic effects of lamin A/C haploinsufficiency

A cellular model of Brugada syndrome with SCN10A variants using human-induced pluripotent stem cell-derived cardiomyocytes

EP Europace ◽  
2019 ◽  
Vol 21 (9) ◽  
pp. 1410-1421 ◽  
Author(s):  
Ibrahim El-Battrawy ◽  
Sebastian Albers ◽  
Lukas Cyganek ◽  
Zhihan Zhao ◽  
Huan Lan ◽  
...  
Keyword(s):  
Stem Cell ◽  
Sodium Channel ◽  
Brugada Syndrome ◽  
Pluripotent Stem Cell ◽  
Induced Pluripotent Stem Cell ◽  
Dermal Fibroblasts ◽  
Cellular Model ◽  
Channel Current ◽  
Healthy Control ◽  
Induced Pluripotent

Abstract Aims Brugada syndrome (BrS) is associated with a pronounced risk to develop sudden cardiac death (SCD). Up to 21% of patients are related to mutations in SCN5A. Studies identified SCN10A as a contributor of BrS. However, the investigation of the human cellular phenotype of BrS in the presence of SCN10A mutations remains lacking. The objective of this study was to establish a cellular model of BrS in presence of SCN10A mutations using human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). Methods and results Dermal fibroblasts obtained from a BrS patient suffering from SCD harbouring the SCN10A double variants (c.3803G>A and c.3749G>A) and three independent healthy control subjects were reprogrammed to hiPSCs. Human-induced pluripotent stem cells were differentiated into cardiomyocytes (hiPSC-CMs).The hiPSC-CMs from the BrS patient showed a significantly reduced peak sodium channel current (INa) and a significantly reduced ATX II (sea anemone toxin, an enhancer of late INa) sensitive as well as A-887826 (a blocker of SCN10A channel) sensitive late sodium channel current (INa) when compared with the healthy control hiPSC-CMs, indicating loss-of-function of sodium channels. Consistent with reduced INa the action potential amplitude and upstroke velocity (Vmax) were significantly reduced, which may contribute to arrhythmogenesis of BrS. Moreover, Ajmaline effects on action potentials were stronger in BrS-hiPSC-CMs than in healthy control cells. This is in agreement with the higher susceptibility of patients to sodium channel blocking drugs in unmasking BrS. Conclusion Patient-specific hiPSC-CMs are able to recapitulate single-cell phenotype features of BrS with SCN10A mutations and may provide novel opportunities to further elucidate the cellular disease mechanism.

scholarly journals Deformation of the nucleus by TGFβ1 via the remodeling of nuclear envelope and histone isoforms

2022 ◽  
Vol 15 (1) ◽  
Author(s):  
Ya-Hui Chi ◽  
Wan-Ping Wang ◽  
Ming-Chun Hung ◽  
Gunn-Guang Liou ◽  
Jing-Ya Wang ◽  
...  
Keyword(s):  
Nuclear Envelope ◽  
Cell Lines ◽  
Nuclear Periphery ◽  
Nuclear Lamina ◽  
Lamin A ◽  
Nuclear Deformation ◽  
Smad Signaling ◽  
Lamin B1 ◽  
Compositional Changes ◽  
Nucleosomal Structure

AbstractThe cause of nuclear shape abnormalities which are often seen in pre-neoplastic and malignant tissues is not clear. In this study we report that deformation of the nucleus can be induced by TGFβ1 stimulation in several cell lines including Huh7. In our results, the upregulated histone H3.3 expression downstream of SMAD signaling contributed to TGFβ1-induced nuclear deformation, a process of which requires incorporation of the nuclear envelope (NE) proteins lamin B1 and SUN1. During this process, the NE constitutively ruptured and reformed. Contrast to lamin B1 which was relatively stationary around the nucleus, the upregulated lamin A was highly mobile, clustering at the nuclear periphery and reintegrating into the nucleoplasm. The chromatin regions that lost NE coverage formed a supra-nucleosomal structure characterized by elevated histone H3K27me3 and histone H1, the formation of which depended on the presence of lamin A. These results provide evidence that shape of the nucleus can be modulated through TGFβ1-induced compositional changes in the chromatin and nuclear lamina.

Sign in / Sign up

Export Citation Format

Share Document