Ion-channel function is determined by its gating movement. Yet, molecular dynamics and electrophysiological simulations were never combined to link molecular structure to function. We performed multiscale molecular dynamics and continuum electrostatics calculations to simulate a cardiac K ؉ channel (IKs) gating and its alteration by mutations that cause arrhythmias and sudden death. An all-atom model of the IKs ␣-subunit KCNQ1, based on the recent Kv1.2 structure, is used to calculate electrostatic energies during gating. Simulations are compared with experiments where varying degrees of positive charge-added via point mutationprogressively reduce current. Whole-cell simulations show that mutations cause action potential and ECG QT interval prolongation, consistent with clinical phenotypes. This framework allows integration of multiscale observations to study the molecular basis of excitation and its alteration by disease.cardiac arrhythmias ͉ long QT syndrome T he current carried by ion channels during the action potential (AP) in excitable tissues is determined by dynamic changes of the ion channel conformation during gating. Recently determined crystal structures of ion channel proteins (e.g., ref. 1) provide a structural basis for computer simulations of the ion-channel protein molecular dynamics (MD) during conformational changes. In parallel, recently acquired experimental data on ion-channel electrophysiological function have provided the basis for detailed Markov models of channel electrical function during gating (2, 3). These models can be used to link cellular consequences of genetic mutations in cardiac ion channels that alter channel kinetics to cardiac arrhythmias [see ref.4 for review]. Although consistent in reproducing the kinetics of channel opening and closing, the molecular conformations represented by the states of these Markov models are not identified.In this article, we develop a multiscale computational approach that combines molecular dynamics simulations and PoissonBoltzmann continuum electrostatic calculations with electrophysiological modeling for establishing structure-function relationships between ion channel movement during gating, its function as a carrier of transmembrane ionic current, and its role in the wholecell AP. Detailed understanding of these relationships is necessary for the development of effective genetic/molecular therapies and specific drug treatment of disease states that involve abnormal electrical function of excitable tissues. We present an example of a cardiac ion channel, the slow delayed rectifier I Ks , which participates in AP repolarization and its rate dependence (4). We consider mutations to its alpha subunit, KCNQ1, that lead to reduction of I Ks current and prolongation of the AP. The clinical phenotype associated with such mutations is the Long-QT syndrome type 1, LQT1; it presents as QT interval prolongation on the ECG and predisposes patients to cardiac arrhythmia and sudden death. Results and DiscussionSeveral charged residues that participate i...

Decker KF, Heijman J, Silva JR, Hund TJ, Rudy Y. Properties and ionic mechanisms of action potential adaptation, restitution, and accommodation in canine epicardium. Am J Physiol Heart Circ Physiol 296: H1017-H1026, 2009. First published January 23, 2009 doi:10.1152/ajpheart.01216.2008.-Computational models of cardiac myocytes are important tools for understanding ionic mechanisms of arrhythmia. This work presents a new model of the canine epicardial myocyte that reproduces a wide range of experimentally observed rate-dependent behaviors in cardiac cell and tissue, including action potential (AP) duration (APD) adaptation, restitution, and accommodation. Model behavior depends on updated formulations for the 4-aminopyridine-sensitive transient outward current (Ito1), the slow component of the delayed rectifier K ϩ current (IKs), the L-type Ca 2ϩ channel current (ICa,L), and the Na ϩ -K ϩ pump current (INaK) fit to data from canine ventricular myocytes. We found that Ito1 plays a limited role in potentiating peak ICa,L and sarcoplasmic reticulum Ca 2ϩ release for propagated APs but modulates the time course of APD restitution. IKs plays an important role in APD shortening at short diastolic intervals, despite a limited role in AP repolarization at longer cycle lengths. In addition, we found that ICa,L plays a critical role in APD accommodation and rate dependence of APD restitution. Ca 2ϩ entry via ICa,L at fast rate drives increased Na ϩ -Ca 2ϩ exchanger Ca 2ϩ extrusion and Na ϩ entry, which in turn increases Na ϩ extrusion via outward INaK. APD accommodation results from this increased outward INaK. Our simulation results provide valuable insight into the mechanistic basis of rate-dependent phenomena important for determining the heart's response to rapid and irregular pacing rates (e.g., arrhythmia). Accurate simulation of rate-dependent phenomena and increased understanding of their mechanistic basis will lead to more realistic multicellular simulations of arrhythmia and identification of molecular therapeutic targets. arrhythmia; cardiac electrophysiology; mathematical modeling; ion channels CARDIAC ARRHYTHMIAS and sudden death involve complex myocardial activation patterns, including unidirectional block, reentry, and fibrillation. To understand the relations and transitions between these patterns, the ionic determinants of the response of healthy and diseased cardiac myocytes to complex patterns of excitation must be understood. The single-cell response to such excitation patterns depends on the complex interaction between ionic currents, intracellular ion concentrations, and membrane voltage. Computational cell models provide critical tools for exploring these interactions, allowing the development and testing of hypotheses about underlying ionic mechanisms based on careful integration of available experimental data (37). The dog is a common animal model for studying cell electrophysiology in a range of disease states. Our group and others have developed detailed mathematical models of the canine action ...

The androgen receptor (AR) is a ligand-inducible transcription factor that mediates androgen action in target tissues. Upon ligand binding, the AR binds to thousands of genomic loci and activates a cell-type specific gene program. Prostate cancer growth and progression depend on androgen-induced AR signaling. Treatment of advanced prostate cancer through medical or surgical castration leads to initial response and durable remission, but resistance inevitably develops. In castration-resistant prostate cancer (CRPC), AR activity remains critical for tumor growth despite androgen deprivation. Although previous studies have focused on ligand-dependent AR signaling, in this study we explore AR function under the androgen-deprived conditions characteristic of CRPC. Our data demonstrate that AR persistently occupies a distinct set of genomic loci after androgen deprivation in CRPC. These androgen-independent AR occupied regions have constitutively open chromatin structures that lack the canonical androgen response element and are independent of FoxA1, a transcription factor involved in ligand-dependent AR targeting. Many AR binding events occur at proximal promoters, which can act as enhancers to augment transcriptional activities of other promoters through DNA looping. We further show that androgen-independent AR binding directs a gene expression program in CRPC, which is necessary for the growth of CRPC after androgen withdrawal.

Ca 2+ /calmodulin-dependent protein kinase II is a multifunctional serine/threonine kinase with diverse cardiac roles including regulation of excitation contraction, transcription, and apoptosis. Dynamic regulation of CaMKII activity occurs in cardiac disease and is linked to specific disease phenotypes through its effects on ion channels, transporters, transcription and cell death pathways. Recent mathematical models of the cardiomyocyte have incorporated limited elements of CaMKII signaling to advance our understanding of how CaMKII regulates cardiac contractility and excitability. Given the importance of CaMKII in cardiac disease, it is imperative that computer models evolve to capture the dynamic range of CaMKII activity. In this study, using mathematical modeling combined with biochemical and imaging techniques, we test the hypothesis that CaMKII signaling in the canine infarct border zone (BZ) contributes to impaired calcium homeostasis and electrical remodeling. We report that the level of CaMKII autophosphorylation is significantly increased in the BZ region. Computer simulations using an updated mathematical model of CaMKII signaling reproduce abnormal Ca 2+ transients and action potentials characteristic of the BZ. Our simulations show that CaMKII hyperactivity contributes to abnormal Ca 2+ homeostasis and reduced action potential upstroke velocity due to effects on I Na gating kinetics. In conclusion, we present a new mathematical tool for studying effects of CaMKII signaling on cardiac excitability and Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. NIH Public Access

Methylation of the CpG-rich region (CpG island) overlapping a gene’s promoter is a generally accepted mechanism for silencing expression. While recent technological advances have enabled measurement of DNA methylation and expression changes genome-wide, only modest correlations between differential methylation at gene promoters and expression have been found. We hypothesize that stronger associations are not observed because existing analysis methods oversimplify their representation of the data and do not capture the diversity of existing methylation patterns. Recently, other patterns such as CpG island shore methylation and long partially hypomethylated domains have also been linked with gene silencing. Here, we detail a new approach for discovering differential methylation patterns associated with expression change using genome-wide high-resolution methylation data: we represent differential methylation as an interpolated curve, or signature, and then identify groups of genes with similarly shaped signatures and corresponding expression changes. Our technique uncovers a diverse set of patterns that are conserved across embryonic stem cell and cancer data sets. Overall, we find strong associations between these methylation patterns and expression. We further show that an extension of our method also outperforms other approaches by generating a longer list of genes with higher quality associations between differential methylation and expression.