MYL2

MYL2 encodes the ventricular regulatory myosin light chain (RLC), a core subunit of the cardiac sarcomeric myosin hexamer that binds the myosin heavy chain neck region and controls cross-bridge cycling kinetics, calcium sensitivity of force, and the equilibrium between the energy-conserving super-relaxed (SRX) and force-generating disordered-relaxed (DRX) states of myosin (PMID:26074085, PMID:35177471). Phosphorylation of MYL2 at Ser15, catalyzed by MYLK3 downstream of GATA4, is essential for normal cardiac contractile function and displays an epicardial-to-endocardial gradient; its loss leads to sarcomere disarray and cardiac dysfunction (PMID:26074085, PMID:41330421, PMID:31101927). Pathogenic mutations produce distinct cardiomyopathy phenotypes: HCM-associated variants (e.g., D166V, R58Q, IVS6-1) disrupt the SRX state, increase calcium sensitivity, and cause cellular hypertrophy, whereas the DCM-associated D94A mutation favors SRX, reduces calcium sensitivity, and repositions cross-bridges away from actin (PMID:35177471, PMID:25825243, PMID:29463717, PMID:30796699). Recessive loss-of-function MYL2 mutations cause infantile cardioskeletal myopathy through protein degradation or mislocalization and loss of normal sarcomeric incorporation (PMID:23365102, PMID:32453731).

1. 2013 Medium

   Establishing that recessive MYL2 mutations cause human disease resolved whether RLC loss-of-function is tolerated: biallelic mutations disrupting the C-terminal EF-hand domain cause cardioskeletal myopathy with loss of normal RLC protein expression and myofibrillar disorganization.

   Evidence Linkage analysis, exome sequencing, and immunohistochemistry of patient skeletal muscle

   PMID:23365102

   Open questions at the time

   • No in vitro reconstitution of mutant RLC function
   • Mechanism of protein loss (degradation vs. misfolding) not determined
   • Skeletal muscle involvement mechanism unclear
2. 2015 High

   Defining the functional consequences of MYL2 phosphorylation at Ser15 established that RLC phosphorylation directly regulates cross-bridge cycling kinetics, calcium-dependent contraction, and cardiac torsion, with a specific transmural gradient across the ventricular wall.

   Evidence Genetic mouse models with phosphorylation-site mutations, computational models, and in vitro phosphorylation assays

   PMID:26074085

   Open questions at the time

   • Kinase(s) responsible for the spatial phosphorylation gradient not fully defined in vivo
   • Whether phosphorylation acts through SRX/DRX transitions was not yet tested
3. 2015 High

   Characterizing the DCM-associated D94A mutation in vitro revealed that a disease-causing RLC variant reduces α-helical content, impairs RLC–MHC binding and incorporation, and paradoxically increases actin-activated ATPase activity, establishing that RLC structural integrity controls myosin enzymatic function.

   Evidence Recombinant RLC expression, circular dichroism, ATPase assays, and reconstitution into RLC-depleted porcine cardiac myosin

   PMID:25825243

   Open questions at the time

   • In vitro ATPase increase contradicted later in vivo findings
   • Cross-bridge structural positioning not assessed
4. 2016 High

   Analysis of the HCM-associated IVS6-1 splice site mutation demonstrated that it impairs RLC–MHC binding, slows ATP-induced acto-myosin dissociation, and increases calcium sensitivity of force, defining a shared functional signature among HCM-causing RLC mutations.

   Evidence Reconstitution of recombinant IVS6-1 RLC into depleted porcine cardiac preparations with ATPase, stopped-flow kinetics, and skinned muscle force assays

   PMID:27378946

   Open questions at the time

   • No in vivo validation of this splice variant
   • Whether this variant alters SRX/DRX balance was unknown
5. 2018 High

   Transgenic D94A mice resolved a discrepancy between in vitro and in vivo ATPase effects and showed that DCM-D94A decreases calcium sensitivity and repositions cross-bridges toward the thick-filament backbone, directly linking RLC mutation to ventricular dilation and reduced ejection fraction in vivo.

   Evidence Transgenic mouse echocardiography, invasive hemodynamics, skinned fiber force-pCa, and small-angle X-ray diffraction

   PMID:29463717

   Open questions at the time

   • SRX/DRX equilibrium not directly measured
   • How cross-bridge repositioning mechanistically causes dilation not resolved
6. 2019 High

   AAV9-mediated delivery of phosphomimetic S15D-RLC rescued cardiac function in HCM-D166V mice, demonstrating that Ser15 phosphorylation is not merely correlative but causally required for normal contraction and is a viable therapeutic target.

   Evidence AAV9 gene therapy in D166V transgenic mice with echocardiography, PV loops, and skinned papillary muscle mechanics

   PMID:31101927

   Open questions at the time

   • Long-term effects and potential immunogenicity not assessed
   • Whether S15D also corrects SRX/DRX imbalance was not tested
7. 2019 Medium

   iPSC-derived cardiomyocytes from R58Q patients established that HCM-causing RLC mutations perturb cellular calcium handling (reduced peak transients, decreased L-type Ca²⁺ current) in addition to sarcomere mechanics, broadening the pathomechanism beyond cross-bridge cycling.

   Evidence Patient iPSC-CM model with cell size measurements, calcium imaging, and patch-clamp electrophysiology

   PMID:30796699

   Open questions at the time

   • Single patient line; isogenic controls not described
   • Whether calcium channel effects are primary or secondary to sarcomere dysfunction is unclear
8. 2020 Medium

   Demonstrating that a recessive frameshift MYL2 variant is degraded via the proteasome while HCM missense variants are stably expressed but mislocalized clarified that distinct protein quality control fates underlie recessive vs. dominant MYL2 disease mechanisms.

   Evidence In vitro overexpression with proteasome inhibitor rescue; Drosophila Mlc2 knockdown rescue experiments

   PMID:32453731

   Open questions at the time

   • Proteasome degradation shown by inhibitor rescue, not by direct ubiquitination assay
   • Drosophila Mlc2 is not identical to human MYL2
9. 2022 High

   Simultaneous X-ray diffraction and force measurements in D166V and D94A mouse hearts unified the SRX/DRX framework for RLC mutations: HCM-D166V destabilizes SRX (shifting myosin toward DRX/actin), while DCM-D94A stabilizes SRX, providing an energetic basis for opposing cardiomyopathy phenotypes.

   Evidence Small-angle X-ray diffraction with isometric force in skinned papillary muscles; mant-ATP SRX/DRX assays

   PMID:35177471

   Open questions at the time

   • Whether phosphorylation state modulates the SRX/DRX effect of each mutation was not tested
   • Structural basis at atomic resolution was lacking
10. 2025 Medium

    Identification of the GATA4→MYLK3→MYL2 phosphorylation axis as the mechanism of osimertinib-induced cardiotoxicity placed MYL2 phosphorylation regulation in a defined transcriptional signaling pathway and demonstrated reversibility upon drug withdrawal or myosin activator treatment.

    Evidence snRNA-seq, iPSC-CM assays, mouse TAC model, and pharmacological intervention with omecamtiv mecarbil

    PMID:41330421

    Open questions at the time

    • Published year listed as 2026 suggesting early online; independent replication needed
    • Whether GATA4-MYLK3 is the dominant pathway for MYL2 phosphorylation in physiological conditions is not established

• A complete atomic-resolution understanding of how individual RLC mutations alter the interacting-heads motif (IHM) and thick-filament quaternary structure in the human heart remains unresolved, as does the question of whether SRX/DRX rebalancing is a universal therapeutic target across all MYL2 cardiomyopathy variants.
• No high-resolution human cardiac thick-filament structure with mutant RLC
• Therapeutic SRX/DRX modulation not tested for recessive loss-of-function variants
• In vivo role of MYL2 in NLRP3 inflammasome regulation requires independent confirmation