The discovery of an association between mutant A-type lamins and disease dates back to 1999 when Bonne and co-workers uncovered mutant LMNA in an autosomal dominant form of Emery-Dreyfuss muscular dystrophy (EDMD) [1]. Additionally, Fatkin et al. reported a link between LMNA and dilated cardiomyopathy (DCM) [2]. A-type lamins are type V intermediate filament (IF) proteins located on the underside of the inner nuclear membrane [3, 4], where they polymerize to form a nuclear lamina meshwork interacting with B-type lamins. A-type lamins are also found in the nucleoplasm [5]. Like other type V IF proteins, lamins consist of a globular head and tail domain with an immunoglobulin-like fold, separated by a helical rod domain [6]. Lamins can broadly be categorized into A-type and B-type lamins. Although this review primarily focuses on A-type lamins, it is important to understand their differences. B-type lamins are encoded by LMNB1 and LMNB2 genes. Lamin A and lamin C are collectively referred to as A-type lamins and are isoforms resulting from alternative splicing and post-translational processing of the primary transcript of the LMNA gene [7]. Lamin A and C are identical until the 10th exon – while lamin A messenger ribonucleic acid (mRNA) contains 12 exons, lamin C lacks the last two exons [8]. Lamin A undergoes farnesylation (post-translational isoprenyl addition) at its CaaX motif and subsequent loss of farnesyl through endoproteolytic cleavages, whereas lamin C lacks a farnesylation site altogether [8]. The zinc metalloproteinase STE24 (ZMPSTE24), which is highly expressed in the heart, is responsible for lamin A maturation via proteolytic cleavage [9]. In contrast, farnesylation is maintained in B-type lamins [10]. The expression of A-type lamins is influenced by matrix stiffness [11], which directs cell differentiation towards either a stiff and contractile lineage or soft lineage, depending on A-type lamins content [12]. In contrast, B-type lamin expression patterns are not regulated in a stiffness-dependent manner [11]. See Fig. 1 for a graphical representation of the structure and expression pattern of lamins.

Graphical representation of the lamin isoforms. A Lamin A, B and C are isoforms resulting from alternative splicing and post-translational processing of the primary transcript of the LMNA and LMNB genes. B Bulk RNA-seq data of LMNA and LMNB in the cardiovascular system. C Single-cell RNA expression profiles of LMNA and LMNB in selected cell types within the left ventricle of the heart (source: Multi-Gene Single Cell Viewer from GTEx)

Mechanosensitive pathways in cardiolaminopathy constitute a complex and challenging medical puzzle. LMNA-related heart failure (HF) presents a broad spectrum of clinical manifestations, ranging from conduction problems, atrial and ventricular arrhythmias, and atrioventricular block to isolated cardiac dysfunction, often leading to devastating outcomes [13]. While treatment advancements have significantly improved the prognosis for most forms of HF, individuals afflicted by inherited forms of DCM, prominently featuring LMNA mutations, continue to face grim prospects. The continuous interaction between cells and their extracellular matrix (ECM) intricately shapes both the biomechanical and biochemical characteristics of the ECM. Simultaneously, this interaction profoundly impacts cellular functionality by activating signal transduction pathways responsible for governing gene and protein expression [14]. Here, we summarise the enigma of cardiolaminopathy, mainly focusing on the mechanosensitive pathways and their downstream effectors. Furthermore, we aim to shed light on the genetic and molecular complexity of cardiolaminopathy, highlighting the urgent necessity to uncover novel therapeutic targets. Unraveling and redefining the potential modifications in the pathways are essential mechanisms in the gene regulation of mechanotransduction. There is an increasing demand to speed up research from fundamental to preclinical studies to address the needs of individuals affected by this devastating condition.

To improve the accuracy and reliability of future research, it is essential that we take into account the inconsistency with which protein and pathway names were represented in previous studies. Our review aims to address discrepancies in existing research by providing a thorough alignment of protein and pathway names with correct gene nomenclature.

Given the location of the nuclear lamina, it is hardly surprising that A-type lamins connect the nuclear interior to the cytoskeleton. A-type lamins are part of a linker between the cytoskeleton and nucleoplasm complex (LINC), which spans the nuclear envelope (NE) [15]. The key elements in this complex are SUN domain proteins at the inner nuclear membrane that extend into the perinuclear space, where they link to and localize nesprins (SYNE1 and SYNE2) at the outer nuclear membrane [15]. On the cytoplasmic side, nesprins link to cytoskeletal elements as they can bind actin directly [16] and indirectly connect to microtubules and IFs [17]. On the nucleoplasmic side, the SUN domain protein SUN1 has a high affinity for lamin A [18]. Lamins can bind chromatin directly. A-type lamins are involved in many biological processes, such as nuclear patterning, and regulation of large chromatin domains called lamina-associated domains (LADs) [19]. LADs are localized at gene-poor heterochromatic regions, and genes localized at LADs are often repressed [20]. A-type lamins also interact with chromatin-binding proteins, such as the LAP2-emerin-MAN1-containing (LEM) domain in emerin (EMD). The LEM domain binds to the chromatin protein BAF (BANF1), which undergoes a conformational change during self-assembly of the emerin N-terminal region [21,22,23]. On the cytoplasmic side, nesprins link to cytoskeletal elements as they can bind actin directly [16] and indirectly connect to microtubules and IFs [17]. In the myocardium and skeletal muscle, nesprins also connect to the Z-discs of the contractile sarcomeres [16, 21].

More than a decade ago, it was determined that the LINC complex is not only a physical connection but also a mechanosensor, as it was discovered that the sun-nesprin interaction is critical for cellular mechanotransduction and cellular tension [24]. Thus, the LINC complex is part of the mechanotransduction pathway and transduces mechanical forces to the nucleus. Signals from the environment are transmitted to ECM molecules that enter the cell through integrins and are transmitted to actin fibers via focal adhesion molecules and further into the nucleus via LINC molecules [25]. The perinuclear actin cap, which forms around the nucleus in a LINC-mediated process, is much more sensitive to mechanical stresses than stress fibers. Therefore, rapid mechanosensitive transduction is required between the environment and the nucleus to protect and properly exert a mechanoresponse [26].

The increased expression of stiff matrices suggests that A-type lamins have an important role in the mechanotransduction of extracellular mechanical signals into appropriate cellular responses [27]. Other roles for lamins include maintenance of NE stiffness and integrity [28]. B-type lamins are associated with nuclear elasticity in this role, while A-type lamins are responsible for viscous characteristics [11, 29]. Lamins also regulate gene expression through binding (hetero)chromatin at lamin-associated domains and interacting with various transcription factors [5]. Indeed, many of lamin's functions are performed indirectly through interactions with proteins [30, 31]. Therefore, abnormal lamins could lead to severe interruptions to biological and mechanosensitive processes.

Given that the nucleus functions as a mechanosensor through its connection to the cytoskeleton and, consequently, to the ECM via the LINC complex [30], cardiolaminopathy results in substantial disruption of nuclear mechanobiological processes. The degree of these disruptions is directly associated with the overall clinical severity of the phenotype [32]. Abnormal lamina contributes to an abnormal LINC complex, an imbalance between extracellular and intracellular tension, and, as a consequence, impaired mechanotransduction [33]. Uncoupling of the nucleus from the cytoplasm results in the inability to properly sense and respond to matrix stiffness: it was shown that regardless of matrix stiffness, LMNA pathogenic variant-carrying striated muscle cells behaved as if they were on a rigid matrix by expressing more stress fibers, vinculin (VCL), and focal adhesion kinases than wild-type cells [34]. In three Lmna mouse models and muscle biopsies from individuals with LMNA-related muscular dystrophy, the mutations destabilized the nucleus, causing temporary nuclear envelope ruptures in muscle cells. This led to DNA damage, activation of DNA damage responses, and reduced cell viability [35]. Furthermore, it was demonstrated that mutant cells fail to adapt to mechanical stress, as evidenced by unchanged focal adhesions and damage to the actin filaments in the cytoskeleton. The protein reflecting this abnormal response compared to wild-type (WT) myoblasts is Yes-associated protein (YAP/YAP1) [34].

WW domain containing transcription regulator 1 (WWTR1), previously also known as transcriptional coactivator with PDZ-binding motif (TAZ, note: it should not be confused with phospholipid-lysophospholipid transacylase TAFFAZIN, previously also known as TAZ), binds to YAP1. Together, they are the effectors of an important pathway implicated in mechanotransduction, namely the Hippo pathway [36]. YAP1 and WWTR1 (YAP1/WWTR1) are considered both sensors and mediators of mechanical cues originating from the cell environment [37]. First identified in Drosophila, the core (also referred to as the cassette) of the Hippo pathway, composed of Hippo (Hpo), Salvador (Sav), Warts (Wts), and Mob-as-tumour-suppressor proteins, as well as the effector Yorkie (Yki) [38], is highly conserved in mammals [39]. The mammalian pathway consists of an enzymatic cascade of two main kinase families, Hpo orthologs macrophage stimulating 1 (MST1) and serine/threonine kinase 3 (STK3, previously known as MST2) and Wts orthologs large tumor suppressor kinase 1/2 (LATS1/2) [40], the activation of which results in the inactivation of YAP1/WWTR1 [41]. MST1/STK3 can be activated by autophosphorylation or via phosphorylation by TAO kinases (TAOK1/2/3) [42]. When active, MST1/STK3, along with the adaptor protein salvador family WW domain-containing protein 1 (SAV1, Sav ortholog), phosphorylates MOB kinase activators MOB1A/1B (Mats orthologs) [36], which bind to LATS1/2, phosphorylating the latter [43]. Phosphorylated LATS1/2 can phosphorylate YAP1/WWTR1 (Yki ortholog) [41]. The phosphorylation of YAP1 by LATS1/2 occurs at Ser127 residue, and phosphorylated YAP1 is subsequently inactivated and sequestered in the cytoplasm through the binding to 14–3-3 proteins and, thereby, inhibiting its function by conducting the complex towards proteasomal degradation [42]. In contrast, if the Hippo cassette is inactive, YAP1/WWTR1 remains unphosphorylated and is able to translocate into the nucleus. YAP1/WWTR1 lacks deoxyribonucleic acid (DNA) sequence-specificity. Therefore, to stimulate the transcription of target genes, they must bind to sequence-specific transcription factors, the most common of which is the TEAD group (containing genes TEAD1/2/3/4, Scalloped ortholog) [36]. See Fig. 2 for more details on the function and expression of the Hippo pathway genes.

A schematic overview of the Hippo signaling pathway. A The laminopathic cell experiences stress, and activate the canonical Hippo pathway, resulting in the YAP1-TEAD complex to drive the transcription of proliferative genes. During homeostasis, residue-phosphorylation generates a 14–3-3-protein binding site, causing cytoplasmic sequestration and marking them for proteasomal degradation, ultimately inhibiting YAP and WWTR1 (previously known as TAZ) activity and partial degradation of YAP1. Note that the 14–3-3 protein family is encoded by six genes (YWHAB, YWHAE, YWHAG, YWHAH, YWHAQ, YWHAZ, and SFN) B Single-cell RNA expression profiles of genes encoding crucial proteins of the Hippo pathway in selected myocardial cell types (source: Multi-Gene Single Cell Viewer from GTEx). While the displayed genes are considered to be involved in the Hippo pathway and laminopathy, only some of them show sufficient RNA expression in cardiac cells (e.g., see the difference between paralogues MST1 and STK3) and might have different involvement in cardiolaminopathy

Targets of YAP1 are involved in mitogenic activity necessary for cell proliferation, survival, and tissue growth [44]. In adult myocardium, overexpression of unphosphorylated YAP1 leads to cardiomyocyte (CM) proliferation, thickening of ventricular walls, and improvement in cardiac function, effectively inducing a fetal cell state in adult cells [45], corresponding to the postnatal expansion of the cardiac regeneration window [46]. Additionally, the Hippo pathway is a convergence point of cellular signaling interacting with multiple major pathways, including Wnt/β-catenin, insulin-like growth factor, phosphoinositide 3-kinase mediated phosphorylation of AKT1 (RACα serine/threonine-protein kinase or PI3K-RACα/Akt), and mTOR (MTOR) signaling [47].

Next to regulation via the phosphorylation of the Hippo pathway, non-canonical regulation of YAP1 activity has been described [37]. YAP1/WWTR1 activation and translocation into the nucleus can be induced through mechanical forces coming from the cell environment [48], illustrating YAP1/WWTR1 mechanosensing abilities. ECM stiffness is an important factor in dictating YAP1/WWTR1 subcellular shuttling. On soft matrices, YAP1 activity is inhibited due to nuclear exclusion, while stiff matrices have enhanced YAP1 activity and cell spreading [37]. The stiffness-dependence of YAP1 has been demonstrated using engineered fluorescent reporter genes coupled with YAP1 target promoters [49]. This study confirmed that soft matrices favor cytoplasmic localization and inactivation of YAP1, while on stiff substrates, nuclear aggregation and fluorescent reporter expression indicative of YAP1 activation were observed. Notably, rather than the ECM itself, cytoskeletal elements are paramount in regulating YAP1 localization [50]. Using a nanopillar measurement method, it was revealed that YAP1 translocation into the nucleus is guided by force-induced displacements in the perinuclear region dominated by the actin cap [51].

The aforementioned YAP1-induced cell spreading coincides with Ras homolog family member A (RHOA/Rho) activation, which regulates stress fibers, actin bundles, and actomyosin structures [41, 51]. Inhibition experiments showed that RHOA activation and cytoskeletal tension, which can be characterized by stress fibers, are necessary for YAP1 nuclear activity. Further, RHOA stimulates the nuclear import of TAZ [36]. Therefore, it has been shown that non-canonical regulation of YAP1/WWTR1 is involved in the translation of mechanical cues by YAP1/WWTR1 rather than the Hippo pathway. Indeed, RHOA inhibition reinforces the “soft state” response from the cell, resulting in YAP1/WWTR1 inactivation [52]. Moreover, RHOA inhibition can activate LATS, revealing the involvement of mechanical cues in the activity of the canonical Hippo pathway [52]. The precise role of the Hippo pathway connected to mechanotransduction is being revealed slowly. The inhibitory role of the Ras-related GTPase RAP2 (RAP2A) in inhibiting YAP1/WWTR1 nuclear entry was studied, revealing that ECM stiffness communicates via focal adhesions to downstream RAP2A, which in turn activates LATS via intermediates [53].

The aberrant localization of YAP1 is a feature of LMNA mutants and is associated with faulty mechanotransduction. However, it remains unknown how the Hippo pathway regulates cardiac remodeling in different pathologies [34, 54]. Upregulation of YAP1 expression and its increased nuclear levels have been reported in failing human hearts [54]. YAP1 knockout mediates the development of DCM, while hypertrophic cardiomyopathy (HCM) involves a deficiency in Hippo signaling, the upregulation of YAP1 transcript and protein levels, as well as the downregulation of its Ser127 phosphorylation [46, 54]. This sequence of events leads to increased transcription levels of target genes, including myosin heavy chain 7 (MYH7) and troponin T2 cardiac type (TNNT2) [55]. YAP1 inactivation correlates with decreased proliferation of CMs and cardiac hypoplasia [44]. Interestingly, it has been suggested that LATS can regulate hypertrophic cues independently of YAP1, suggesting that YAP1 is more important for CM survival and regulation than hypertrophic growth [44]. Thus, the involvement of Hippo and YAP1 in cardiomyopathies is unclear and likely mutation- and tissue-specific, warranting further investigation. Some indirect clues have been gathered through the Lmna H222P/H222P murine model, as it has been shown that treatment with an angiotensin-converting enzyme (ACE) inhibitor improves fractional shortening [