Cytosolic DNA sensing protein pathway is activated in human hearts with dilated cardiomyopathy

Abstract

Introduction: The genome is constantly exposed to numerous stressors, which induce DNA lesions, including double-stranded DNA breaks (DSBs). DSBs are the most dangerous, as they induce genomic instability. In response to DNA damage, the cell activates nuclear DNA damage response (DDR) and the cytosolic DNA sensing protein (CDSP) pathways, the latter upon release of the DSBs to the cytosol. The CDSP pathway activates NFκB and IRF3, which induce the expression of the pro-inflammatory genes. There is scant data on the activation of the CDSP pathway in human hearts with dilated cardiomyopathy (DCM).

Aim: We aimed to determine expression levels of selected components of the CDSP pathway in human hearts with DCM.

Methods: The DNA strand breaks were detected by the single-cell gel electrophoresis or the comet assay and expression of selected proteins by immunoblotting. Transcript levels were quantified in the RNA-Seq data.

Results: Single-cell gel electrophoresis showed an approximately 2-fold increase in the number of COMET cells in the DCM hearts. Immunoblotting showed increased levels of cyclic GMP-AMP synthase (CGAS), the canonical CDSP; TANK-binding kinase 1 (TBK1), an intermediary kinase in the pathway; and RELB, P52, and P50 components of the NFκB pathway in human heart samples from patients with DCM. Likewise, transcript levels of over 2 dozen genes involved in inflammatory responses were increased.

Conclusions: The findings provide the first set of evidence for the activation of the CDSP pathway in human hearts with DCM. The data in conjunction with the previous evidence of activation of the DDR pathway implicate the DSBs in the pathogenesis of human DCM.

Keywords

DNA damage / double-stranded DNA breaks / cardiomyopathy / heart failure

References

[1] López-Otín C,Partridge L,Kroemer G.Hallmarks of aging: an expanding universe.Cell2023;186:243-78

[2] Kronenberg A.Radiation-induced genomic instability.Int J Radiat Biol1994;66:603-9

[3] Aguilera A.Causes of genome instability.Annu Rev Genet2013;47:1-32

[4] Lindahl T.Instability and decay of the primary structure of DNA.Nature1993;362:709-15

[5] Vilenchik MM.Endogenous DNA double-strand breaks: production, fidelity of repair, and induction of cancer.Proc Natl Acad Sci U S A2003;100:12871-6 PMCID:PMC240711

[6] Cannan WJ.Mechanisms and consequences of double-strand dna break formation in chromatin.J Cell Physiol2016;231:3-14 PMCID:PMC4994891

[7] Chapman JR,Boulton SJ.Playing the end game: DNA double-strand break repair pathway choice.Mol Cell2012;47:497-510

[8] Haber JE.Deciphering the DNA damage response.Cell2015;162:1183-5

[9] Ablasser A.cGAS in action: expanding roles in immunity and inflammation.Science2019;363:eaat8657

[10] Motwani M,Fitzgerald KA.DNA sensing by the cGAS-STING pathway in health and disease.Nat Rev Genet2019;20:657-74

[11] Rouhi L,Zhou Q.Deletion of the Lmna gene in fibroblasts causes senescence-associated dilated cardiomyopathy by activating the double-stranded DNA damage response and induction of senescence-associated secretory phenotype.J Cardiovasc Aging2022;2:30 PMCID:PMC9311325

[12] Rouhi L,Chen SN.Haploinsufficiency of Tmem43 in cardiac myocytes activates the DNA damage response pathway leading to a late-onset senescence-associated pro-fibrotic cardiomyopathy.Cardiovasc Res2021;117:2377-94 PMCID:PMC8861264

[13] Chen SN,Karmouch J.DNA damage response/TP53 pathway Is activated and contributes to the pathogenesis of dilated cardiomyopathy associated with LMNA (Lamin A/C) mutations.Circ Res2019;124:856-73 PMCID:PMC6460911

[14] Marian AJ.Non-syndromic cardiac progeria in a patient with the rare pathogenic p.Asp300Asn variant in the LMNA gene.BMC Med Genet2017;18:116 PMCID:PMC5648416

[15] Cheedipudi SM,Marian AJ.Genetic ablation of the DNA damage response pathway attenuates lamin-associated dilated cardiomyopathy in mice.JACC Basic Transl Sci2022;7:1232-45 PMCID:PMC9831927

[16] Rouhi L,Cheedipudi SM.The EP300/TP53 pathway, a suppressor of the hippo and canonical WNT pathways, is activated in human hearts with arrhythmogenic cardiomyopathy in the absence of overt heart failure.Cardiovasc Res2022;118:1466-78 PMCID:PMC9074970

[17] Collins AR.The comet assay for DNA damage and repair: principles, applications, and limitations.Mol Biotechnol2004;26:249-61

[18] Cheedipudi SM,Coarfa C.Genomic reorganization of lamin-associated domains in cardiac myocytes is associated with differential gene expression and DNA methylation in human dilated cardiomyopathy.Circ Res2019;124:1198-213 PMCID:PMC6459729

[19] Thyagarajan B,Groth AC.Mammalian genomes contain active recombinase recognition sites.Gene2000;244:47-54

[20] Loonstra A,Beverloo HB.Growth inhibition and DNA damage induced by Cre recombinase in mammalian cells.Proc Natl Acad Sci U S A2001;98:9209-14 PMCID:PMC55399

[21] Litviňuková M,Maatz H.Cells of the adult human heart.Nature2020;588:466-72 PMCID:PMC7681775

[22] Cordero-Reyes AM,Hamilton DJ,Marian AJ.Molecular, cellular, and functional characterization of myocardial regions in hypertrophic cardiomyopathy.Circ Cardiovasc Imaging2012;5:419-22

[23] Hopfner KP.Molecular mechanisms and cellular functions of cGAS-STING signalling.Nat Rev Mol Cell Biol2020;21:501-21