Get started
Work

Lamin: guardian against DNA damage by transcription stress

Year: 2024
Type: editorial
Abstract: This editorial refers to ‘DNA double-stranded breaks, a hallmark of aging, defined at the nucleotide resolution, are increased and associated with transcription in the cardiac myocytes in LMNA-cardiomyopathy’, by B. Cathcart et al., https://doi.org/10.1093/cvr/cvae063. The human genome spans 3.2 billion base pairs, and more than 20 000 genes govern various cellular processes and functions. A linear DNA is wrapped around histones to form nucleosomes that are further compacted into chromatin fibres. Chromatin fibres undergo a series of additional compression and condensation to form various three-dimensional structural units such as chromatin loops, topologically associated domains, and chromosome territory. Based on the gene activity, chromatin fibres can be grouped into two compartments: A and B compartments. A compartments generally contain transcriptionally active euchromatin and are found inside the nucleus. On the other hand, B compartments contain inactive heterochromatin and are located at the nuclear periphery.1 At the nuclear periphery, the inner side of nuclear envelop is coated by nuclear lamina that contains a complex of protein and nuclear lamins, members of type V intermediate filament, which play important roles in nuclear processes and mechanical properties of nuclei. There are two types of lamins, A type and B type. A-type lamins include lamins A/C and are transcribed from a single gene called LMNA, whereas B-type lamins include lamins B1 and B2 that are transcribed from two separate genes, LMNB1 and LMNB2. Cells lacking lamins or expressing mutant forms of lamins fail to establish a proper nuclear lamina, resulting in laminopathies that is characterized by nuclear distortion, aberrant gene expression, and impaired mechanotransduction. Laminopathies are associated with various diseases such as the premature aging syndrome, including Hutchinson–Gilford progeria syndrome, restrictive dermopathy, lipodystrophy, and muscular dystrophy. Notably, mutations in LMNA lead to arrhythmogenic and/or dilated cardiomyopathy (DCM) in patients or animal models of LMNA cardiomyopathy.2 In this issue of Cardiovascular Research, Cathcart et al.3 revealed the genome-wide distribution of double stranded breaks (DSBs) in cardiomyocytes for the first time with END-sequencing (END-seq) and provided novel insights into the role of nuclear lamina in regulating baseline DSBs by transcription. END-seq is a sensitive method for detecting DSBs at nucleotide resolution by ligating a sequencing adaptor to the ends of DSBs enabling the subsequent next generation sequencing to map DSB distribution at the genome-wide level.4 Cathcart et al.3 performed END-seq on 1 million cardiomyocytes from a well-established LMNA-DCM model (Myh6-Cre:LmnaF/F)5,6 and compared them with wild-type (WT) mice. END-seq was conducted on 3-week-old mice, close to the end of their 4-week lifespan. Here, END-seq revealed a relatively modest increase in total DSBs, approximately three times higher compared to WT (2.2% vs. 0.8%). Given severe cardiac phenotypes typically observed at 3-week-old animals in earlier studies,5,6 this change may seem mild yet the spatial distribution of DSBs remains intriguing (Figure 1). Excessive DSB distribution in the genome due to transcription stress. Chromatin fibres are organized into distinct territories that can be largely classified into A and B compartments. A compartments typically reside in the nuclear interior and are composed of euchromatin with high level of gene expression. B compartments typically reside in the nuclear periphery and are composed of heterochromatin with low level of gene expression. Transcription has been known to cause genomic instability due to torsional stress by unwinding double stranded DNA, inducing transient DNA damage by TOP2. In addition, transcription results in formation of non-B-DNA structures such as G-quadruplex and Z-DNA in non-transcribed DNA strand, and excessive presence of non-B-DNA during transcription has been reported to be associated with genomic instability. The study by Cathcart et al.3 demonstrated that cardiomyocytes from LMNA-DCM mice suffered from excessive DSBs at baseline due to transcriptional stress. Combining END-seq and CUT&RUN assay, they found that juxta-positioned LADs in LMNA-DCM mice samples had high levels of DSBs, suggesting a significant role for nuclear laminins in regulating transcriptional stress. DSBs, double stranded breaks; LADs, laminin associated domains; LMNA, laminin A; RNA polymerase II, Pol II; TOP2, topoisomerase 2; TSS, transcription start site. Cathcart et al.3 also performed in-depth profiling of DSB distribution by classifying the gene and the intergenic regions. By comparing RNA sequencing (RNA-seq) data from the earlier study,5 DSBs were preferentially localized at the highly expressed gene regions sites, especially at the transcription start site. A motif analysis of the DSB sites identified 26 transcription factors, including ETS, ETV4, GATA4, and NFATC3, which are related to cardiac development and cardiac function. Additionally, the authors classified DSB sites by distinguishing canonical double helical B-DNA and non-B-DNA structures. Intriguingly, DSB sites are preferentially located near the non-B-DNA structures like Z-DNA. Given that non-B-DNA structures are known to be associated with transcriptional stress,7 Cathcart’s team further investigated the relationship between the nuclear lamina and transcription stress. Transcription stress refers to the physical torsional stress imposed by the unwinding of double stranded DNA, transient exposure of non-transcribed strand to the extrinsic DNA damages, and secondary non-B-DNA structures, which collectively accumulate DSBs at R-loops. Cathcart et al.3 conducted a CUT&RUN assay, a chip-seq equivalent technique with lower background, against LMNA antibody to identify ‘lamina-associated domains’ (LADs) which accounted for 30–40% of the genome.1 The authors delineated constitutive LAD (cLAD) regions to reveal a significant reduction in DSB distribution compared to non-LAD regions. Conversely, in cardiomyocytes from LMNA-DCM mice, juxtaposed LAD regions or originally cLAD regions in WT samples exhibited excessive DSBs, suggesting that nuclear lamina can protect LADs from DSBs induced by transcriptional stress. Taken together, the current study3 not only confirms the conventional understanding that genes in LADs are repressed in in vivo cardiomyocytes but it also provides new insights into how aberrant transcription activations increases the burden of DSBs, which are the underlying cause of heart failure due to laminopathies. This is consistent with previous results showing that targeting subsequent pathway (cGAS-STING, TP53) or epigenetic regulator of DSBs can successfully reverse the progression of heart failure by LMNA deficiency.5,6 Particularly, BRD4 inhibition to target transcription stress may be further investigated as a novel approach as it has been demonstrated effective against non-LMNA heart failure.8 One important question raised by this study is the source of excessive DSBs. Is it due to accumulation of R-loops? Or lack of topoisomerase activity? The current study is limited in that it provides only observational evidence of DSBs accumulation at the active transcription site. Future follow-up investigations should focus on identifying the causality of DSBs accumulation through mechanistic studies. In addition, this study was performed in LMNA depletion models. To expedite the significance of LADs and the burden of DSBs by transcription stress in LMNA-DCM, future efforts could be performed in human models with clinically relevant mutations such as induced pluripotent stem cell derived cardiomyocytes.9,10 Overall, this study represents the first exploration of DSB sites in in vivo heart samples that found a site-specific localization of DSBs in actively transcribed regions and LADs. This publication was supported by grants from the National Institutes of Health R01 HL113006, R01 HL130020, and R01 HL141371 (J.C.W.). All other authors report no relationships relevant to the contents of this paper to disclose. No new data were generated or analysed in support of this research.
DOI: 10.1093/cvr/cvae122
Language: English
Open Access status: green
OpenAlex ID: W4399780809
Cites: 10
Cited by: 2
Related to: 10