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Nesprin-2-dependent ERK1/2 compartmentalisation regulates the DNA damage response in vascular smooth muscle cell ageing.

ABSTRACT: Prelamin A accumulation and persistent DNA damage response (DDR) are hallmarks of vascular smooth muscle cell (VSMC) ageing and dysfunction. Although prelamin A is proposed to interfere with DNA repair, our understanding of the crosstalk between prelamin A and the repair process remains limited. The extracellular signal-regulated kinases 1 and 2 (ERK1/2) have emerged as key players in the DDR and are known to enhance ataxia telangiectasia-mutated protein (ATM) activity at DNA lesions, and in this study, we identified a novel relationship between prelamin A accumulation and ERK1/2 nuclear compartmentalisation during VSMC ageing. We show both prelamin A accumulation and increased DNA damage occur concomitantly, before VSMC replicative senescence, and induce the localisation of ERK1/2 to promyelocytic leukaemia protein nuclear bodies (PML NBs) at the sites of DNA damage via nesprin-2 and lamin A interactions. Importantly, VSMCs treated with DNA damaging agents also displayed prelamin A accumulation and ERK compartmentalisation at PML NBs, suggesting that prelamin A and nesprin-2 are novel components of the DDR. In support of this, disruption of ERK compartmentalisation at PML NBs, by either depletion of nesprin-2 or lamins A/C, resulted in the loss of ATM from DNA lesions. However, ATM signalling and DNA repair remained intact after lamins A/C depletion, whereas nesprin-2 disruption ablated downstream Chk2 activation and induced genomic instability. We conclude that lamins A/C and PML act as scaffolds to organise DNA-repair foci and compartmentalise nesprin-2/ERK signalling. However, nesprin-2/ERK signalling fidelity, but not their compartmentalisation at PML NBs, is essential for efficient DDR in VSMCs.

SUBMITTER: Warren DT

PROVIDER: S-EPMC4532777 | biostudies-literature | 2015 Sep

REPOSITORIES: biostudies-literature

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Nesprin-2-dependent ERK1/2 compartmentalisation regulates the DNA damage response in vascular smooth muscle cell ageing.

Warren D T DT Tajsic T T Porter L J LJ Minaisah R M RM Cobb A A Jacob A A Rajgor D D Zhang Q P QP Shanahan C M CM

Cell death and differentiation 20150306 9

Prelamin A accumulation and persistent DNA damage response (DDR) are hallmarks of vascular smooth muscle cell (VSMC) ageing and dysfunction. Although prelamin A is proposed to interfere with DNA repair, our understanding of the crosstalk between prelamin A and the repair process remains limited. The extracellular signal-regulated kinases 1 and 2 (ERK1/2) have emerged as key players in the DDR and are known to enhance ataxia telangiectasia-mutated protein (ATM) activity at DNA lesions, and in thi ...[more]

PMID: 25744025

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Project description:Persistently unrepaired DNA damage has been identified as a causative factor for vascular ageing. We have previously shown that a defect in the function or expression of the DNA repair endonuclease ERCC1 (excision repair cross complement 1) in mice leads to accelerated, nonatherosclerotic ageing of the vascular system from as early as 8 weeks after birth. Removal of ERCC1 from endothelial alone partly explains this ageing, as shown in endothelial-specific Ercc1 knockout mice. In this study, we determined vascular ageing due to DNA damage in vascular smooth muscle cells, as achieved by smooth muscle-selective genetic removal of ERCC1 DNA repair in mice (SMC-KO: SM22αCre+ Ercc1fl/-). Vascular ageing features in SMC-KO and their wild-type littermates (WT: SM22αCre+ Ercc1fl/+) were examined at the age of 14 weeks and 25 weeks. Both SMC-KO and WT mice were normotensive. Compared to WT, SMC-KO showed a reduced heart rate, fractional shortening, and cardiac output. SMC-KO showed progressive features of nonatherosclerotic vascular ageing as they aged from 14 to 25 weeks. Decreased subcutaneous microvascular dilatation and increased carotid artery stiffness were observed. Vasodilator responses measured in aortic rings in organ baths showed decreased endothelium-dependent and endothelium-independent responses, mostly due to decreased NO-cGMP signaling. NADPH oxidase 2 and phosphodiesterase 1 inhibition improved dilations. SMC-KO mice showed elevated levels of various cytokines that indicate a balance shift in pro- and anti-inflammatory pathways. In conclusion, SMC-KO mice showed a progressive vascular ageing phenotype in resistant and conduit arteries that is associated with cardiac remodeling and contractile dysfunction. The changes induced by DNA damage might be limited to VSMC but eventually affect EC-mediated responses. The fact that NADPH oxidase 2 as wells as phosphodiesterase 1 inhibition restores vasodilation suggests that both decreased NO bioavailability and cGMP degradation play a role in local vascular smooth muscle cell ageing induced by DNA damage.

Project description:ObjectiveWe examined the role of syndecan-1 in modulating the phenotype of vascular smooth muscle cells in the context of endogenous inflammatory factors and altered microenvironments that occur in disease or injury-induced vascular remodeling.Methods and resultsVascular smooth muscle cells (vSMCs) display a continuum of phenotypes that can be altered during vascular remodeling. While the syndecans have emerged as powerful and complex regulators of cell function, their role in controlling vSMC phenotype is unknown. Here, we isolated vSMCs from wild type (WT) and syndecan-1 knockout (S1KO) mice. Gene expression and western blotting studies indicated decreased levels of α-smooth muscle actin (α-SMA), calponin, and other vSMC-specific differentiation markers in S1KO relative to WT cells. The spread area of the S1KO cells was found to be greater than WT cells, with a corresponding increase in focal adhesion formation, Src phosphorylation, and alterations in actin cytoskeletal arrangement. In addition, S1KO led to increased S6RP phosphorylation and decreased AKT and PKC-α phosphorylation. To examine whether these changes were present in vivo, isolated aortae from aged WT and S1KO mice were stained for calponin. Consistent with our in-vitro findings, the WT mice aortae stained higher for calponin relative to S1KO. When exposed to the inflammatory cytokine TNF-α, WT vSMCs had an 80% reduction in syndecan-1 expression. Further, with TNF-α, S1KO vSMCs produced increased pro-inflammatory cytokines relative to WT. Finally, inhibition of interactions between syndecan-1 and integrins αvβ3 and αvβ5 using the inhibitory peptide synstatin appeared to have similar effects on vSMCs as knocking out syndecan-1, with decreased expression of vSMC differentiation markers and increased expression of inflammatory cytokines, receptors, and osteopontin.ConclusionsTaken together, our results support that syndecan-1 promotes vSMC differentiation and quiescence. Thus, the presence of syndecan-1 would have a protective effect against vSMC dedifferentiation and this activity is linked to interactions with integrins αvβ3 and αvβ5.

Dataset Information

Nesprin-2-dependent ERK1/2 compartmentalisation regulates the DNA damage response in vascular smooth muscle cell ageing.

Publications

Nesprin-2-dependent ERK1/2 compartmentalisation regulates the DNA damage response in vascular smooth muscle cell ageing.

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