Dilated cardiomyopathy variant R14del increases phospholamban pentamer stability, blunting dynamic regulation of calcium.

The sarco(endo)plasmic reticulum Ca ATPase (SERCA) is a membrane transporter that creates and maintains intracellular Ca stores. In the heart, SERCA is regulated by an inhibitory interaction with the monomeric form of the transmembrane micropeptide phospholamban (PLB). PLB also forms avid homo-pentamers, and dynamic exchange of PLB between pentamers and SERCA is an important determinant of cardiac responsiveness to exercise.

Another-regulin regulates cardiomyocyte calcium handling via integration of neuroendocrine signaling with SERCA2a activity.

Calcium (Ca) dysregulation is a hallmark feature of cardiovascular disease. Intracellular Ca regulation is essential for proper heart function and is controlled by the sarco/endoplasmic reticulum Ca ATPase (SERCA2a). Another-regulin (ALN) is a newly discovered cardiomyocyte-expressed SERCA2a inhibitor, suggesting cardiomyocyte Ca-handling is more complex than previously appreciated.

Phosphorylation of phospholamban promotes SERCA2a activation by dwarf open reading frame (DWORF).

In cardiac myocytes, the type 2a sarco/endoplasmic reticulum Ca-ATPase (SERCA2a) plays a key role in intracellular Ca regulation. Due to its critical role in heart function, SERCA2a activity is tightly regulated by different mechanisms, including micropeptides. While phospholamban (PLB) is a well-known SERCA2a inhibitor, dwarf open reading frame (DWORF) is a recently identified SERCA2a activator.

Protein carbonylation causes sarcoplasmic reticulum Ca overload by increasing intracellular Na level in ventricular myocytes.

Diabetes is commonly associated with an elevated level of reactive carbonyl species due to alteration of glucose and fatty acid metabolism. These metabolic changes cause an abnormality in cardiac Ca regulation that can lead to cardiomyopathies. In this study, we explored how the reactive α-dicarbonyl methylglyoxal (MGO) affects Ca regulation in mouse ventricular myocytes.

Phospholamban inhibits the cardiac calcium pump by interrupting an allosteric activation pathway.

Phospholamban (PLB) is a transmembrane micropeptide that regulates the sarcoplasmic reticulum Ca-ATPase (SERCA) in cardiac muscle, but the physical mechanism of this regulation remains poorly understood. PLB reduces the Ca sensitivity of active SERCA, increasing the Ca concentration required for pump cycling. However, PLB does not decrease Ca binding to SERCA when ATP is absent, suggesting PLB does not inhibit SERCA Ca affinity.

Protein carbonylation causes sarcoplasmic reticulum Ca2+ overload by increasing intracellular Na+ level in ventricular myocytes.

Diabetes is commonly associated with an elevated level of reactive carbonyl species due to alteration of glucose and fatty acid metabolism. These metabolic changes cause an abnormality in cardiac Ca regulation that can lead to cardiomyopathies. In this study, we explored how the reactive α-dicarbonyl methylglyoxal (MGO) affects Ca regulation in mouse ventricular myocytes.

Mechanisms for cardiac calcium pump activation by its substrate and a synthetic allosteric modulator using fluorescence lifetime imaging.

The discovery of allosteric modulators is an emerging paradigm in drug discovery, and signal transduction is a subtle and dynamic process that is challenging to characterize. We developed a time-correlated single photon-counting imaging approach to investigate the structural mechanisms for small-molecule activation of the cardiac sarcoplasmic reticulum Ca-ATPase, a pharmacologically important pump that transports Ca at the expense of adenosine triphosphate (ATP) hydrolysis. We first tested whether the dissociation of sarcoplasmic reticulum Ca-ATPase from its regulatory protein phospholamban is required for small-molecule activation.

RyR2 Binding of an Antiarrhythmic Cyclic Depsipeptide Mapped Using Confocal Fluorescence Lifetime Detection of FRET.

Hyperactivity of cardiac sarcoplasmic reticulum (SR) ryanodine receptor (RyR2) Ca-release channels contributes to heart failure and arrhythmias. Reducing the RyR2 activity, particularly during cardiac relaxation (diastole), is a desirable therapeutic goal. We previously reported that the unnatural enantiomer () of an insect-RyR activator, verticilide, inhibits porcine and mouse RyR2 at diastolic (nanomolar) Ca and has in vivo efficacy against atrial and ventricular arrhythmia.

Dilated cardiomyopathy variant R14del increases phospholamban pentamer stability, blunting dynamic regulation of cardiac calcium handling.

The sarco(endo)plasmic reticulum Ca ATPase (SERCA) is a membrane transporter that creates and maintains intracellular Ca stores. In the heart, SERCA is regulated by an inhibitory interaction with the monomeric form of the transmembrane micropeptide phospholamban (PLB). PLB also forms avid homo-pentamers, and dynamic exchange of PLB between pentamers and the regulatory complex with SERCA is an important determinant of cardiac responsiveness to exercise.

Micropeptide hetero-oligomerization adds complexity to the calcium pump regulatory network.

The sarco(endo)plasmic reticulum calcium ATPase (SERCA) is an ion transporter that creates and maintains intracellular calcium stores. SERCA is inhibited or stimulated by several membrane micropeptides including another-regulin, dwarf open reading frame, endoregulin, phospholamban (PLB), and sarcolipin. We previously showed that these micropeptides assemble into homo-oligomeric complexes with varying affinity.

Regulation of the regulator: Endopeptidase cleavage controls the expression of a micropeptide that regulates SERCA.

Micropeptides regulate cellular calcium handling by modulating the function of the calcium transporter SERCA. In a recent Nature Communications paper [4] authors Schiemann et al. describe regulation of an invertebrate SERCA-active micropeptide, sarcolamban, by an endopeptidase called neprilysin 4 (NEP4).

Primitive Phospholamban- and Sarcolipin-like Peptides Inhibit the Sarcoplasmic Reticulum Calcium Pump SERCA.

Intracellular calcium signaling is essential for all kingdoms of life. An important part of this process is the sarco-endoplasmic reticulum Ca-ATPase (SERCA), which maintains the low cytosolic calcium levels required for intracellular calcium homeostasis. In higher organisms, SERCA is regulated by a series of tissue-specific transmembrane subunits such as phospholamban in cardiac muscles and sarcolipin in skeletal muscles.

Inhibitory and stimulatory micropeptides preferentially bind to different conformations of the cardiac calcium pump.

The ATP-dependent ion pump sarco/endoplasmic reticulum Ca-ATPase (SERCA) sequesters Ca in the endoplasmic reticulum to establish a reservoir for cell signaling. Because of its central importance in physiology, the activity of this transporter is tightly controlled via direct interactions with tissue-specific regulatory micropeptides that tune SERCA function to match changing physiological conditions. In the heart, the micropeptide phospholamban (PLB) inhibits SERCA, while dwarf open reading frame (DWORF) stimulates SERCA.

Fluorescence lifetime imaging microscopy reveals sodium pump dimers in live cells.

The sodium-potassium ATPase (Na/K-ATPase, NKA) establishes ion gradients that facilitate many physiological functions including action potentials and secondary transport processes. NKA comprises a catalytic subunit (alpha) that interacts closely with an essential subunit (beta) and regulatory transmembrane micropeptides called FXYD proteins. In the heart, a key modulatory partner is the FXYD protein phospholemman (PLM, FXYD1), but the stoichiometry of the alpha-beta-PLM regulatory complex is unknown.

Presenilin 1 is a direct regulator of the cardiac sarco/endoplasmic reticulum calcium pump.

The gamma secretase catalytic subunit presenilin 1 (PS1) is expressed in the endoplasmic reticulum (ER) of neurons, where it regulates Ca signaling. PS1 is also expressed in heart, but its role in regulation of cardiac Ca transport remains unknown. Since the type 2 sarco/endoplasmic reticulum Ca ATPase (SERCA2a) plays a central role in cardiac Ca homeostasis, we studied whether PS1 regulates the cardiac SERCA2a function.

The Ile191Val is a partial loss-of-function variant of the TAS1R2 sweet-taste receptor and is associated with reduced glucose excursions in humans.

Objective: Sweet taste receptors (STR) are expressed in the gut and other extra-oral tissues, suggesting that STR-mediated nutrient sensing may contribute to human physiology beyond taste. A common variant (Ile191Val) in the TAS1R2 gene of STR is associated with nutritional and metabolic outcomes independent of changes in taste perception. It is unclear whether this polymorphism directly alters STR function and how it may contribute to metabolic regulation.

Dwarf open reading frame (DWORF) is a direct activator of the sarcoplasmic reticulum calcium pump SERCA.

The sarco-plasmic reticulum calcium pump (SERCA) plays a critical role in the contraction-relaxation cycle of muscle. In cardiac muscle, SERCA is regulated by the inhibitor phospholamban. A new regulator, dwarf open reading frame (DWORF), has been reported to displace phospholamban from SERCA.

FXYD proteins and sodium pump regulatory mechanisms.

The sodium/potassium-ATPase (NKA) is the enzyme that establishes gradients of sodium and potassium across the plasma membrane. NKA activity is tightly regulated for different physiological contexts through interactions with single-span transmembrane peptides, the FXYD proteins. This diverse family of regulators has in common a domain containing a Phe-X-Tyr-Asp (FXYD) motif, two conserved glycines, and one serine residue.

Dynamics-Driven Allostery Underlies Ca-Mediated Release of SERCA Inhibition by Phospholamban.

Sarcoplasmic reticulum (SR) Ca-ATPase (SERCA) and phospholamban (PLB) are essential for intracellular Ca transport in myocytes. Ca-dependent activation of SERCA-PLB provides a control function that regulates cytosolic and SR Ca levels. Although experimental and computational studies alone have led to a greater insight into SERCA-PLB regulation, the structural mechanisms for Ca binding reversing inhibition of the complex remain poorly understood.

Dimerization of SERCA2a Enhances Transport Rate and Improves Energetic Efficiency in Living Cells.

The type 2a sarco/endoplasmic reticulum (ER) Ca-ATPase (SERCA2a) plays a key role in intracellular Ca regulation in the heart. We have previously shown evidence of stable homodimers of SERCA2a in heterologous cells and cardiomyocytes. However, the functional significance of the pump dimerization remains unclear.

Protein docking and steered molecular dynamics suggest alternative phospholamban-binding sites on the SERCA calcium transporter.

The transport activity of the sarco(endo)plasmic reticulum calcium ATPase (SERCA) in cardiac myocytes is modulated by an inhibitory interaction with a transmembrane peptide, phospholamban (PLB). Previous biochemical studies have revealed that PLB interacts with a specific inhibitory site on SERCA, and low-resolution structural evidence suggests that PLB interacts with distinct alternative sites on SERCA. High-resolution details of the structural determinants of SERCA regulation have been elusive because of the dynamic nature of the regulatory complex.

Newly Discovered Micropeptide Regulators of SERCA Form Oligomers but Bind to the Pump as Monomers.

The recently-discovered single-span transmembrane proteins endoregulin (ELN), dwarf open reading frame (DWORF), myoregulin (MLN), and another-regulin (ALN) are reported to bind to the SERCA calcium pump in a manner similar to that of known regulators of SERCA activity, phospholamban (PLB) and sarcolipin (SLN). To determine how micropeptide assembly into oligomers affects the availability of the micropeptide to bind to SERCA in a regulatory complex, we used co-immunoprecipitation and fluorescence resonance energy transfer (FRET) to quantify micropeptide oligomerization and SERCA-binding. Micropeptides formed avid homo-oligomers with high-order stoichiometry (n > 2 protomers per homo-oligomer), but it was the monomeric form of all micropeptides that interacted with SERCA.

Intrinsically disordered HAX-1 regulates Ca cycling by interacting with lipid membranes and the phospholamban cytoplasmic region.

Hematopoietic-substrate-1 associated protein X-1 (HAX-1) is a 279 amino acid protein expressed ubiquitously. In cardiac muscle, HAX-1 was found to modulate the sarcoendoplasmic reticulum calcium ATPase (SERCA) by shifting its apparent Ca affinity (pCa). It has been hypothesized that HAX-1 binds phospholamban (PLN), enhancing its inhibitory function on SERCA.

Novel approach for quantification of endoplasmic reticulum Ca transport.

The type 2a sarco-/endoplasmic reticulum Ca-ATPase (SERCA2a) plays a key role in Ca regulation in the heart. However, available techniques to study SERCA function are either cell destructive or lack sensitivity. The goal of this study was to develop an approach to selectively measure SERCA2a function in the cellular environment.