Structural and Functional Response of a Mechanosensitive K2P K+ Channel to Asymmetric Membrane Tension

Wednesday, February 15, 2017 electronic coulomb blockade and resonant tunneling in quantum dots. It is a fundamental electrostatic phenomenon based on charge discreteness theory. We used the adaptive biasing method in the colvar module of NAMD to perform the free energy calculation. In the simulations, we found that the coulomb blockade energy is the main contributor as the pore size ranges from 0.6 to 1 nm, while for smaller pores the main contributor changes from ionic coulomb blockade to ion dehydration effects. Moreover, by incorporating both the transport barriers together, we introduced an analytical model to calculate the energy barriers and the current-voltage characteristic curve referring to the electronic coulomb blockade model and Poisson-Boltzmann-Nernst-Planck model. The discrete conductance of ionic transport was well predicted by our model. Therefore, we remark that our results could play an important role in charged pores including biological ion channels and synthetic nanopores. We also believe that the sub-1 nm pores can offer a new platform to explore novel physics in the research areas of nanoscale fluidics and biology. 2686-Pos Board B293 Glutamate Receptor Ion Channel Activation Mechanism Revealed by Cryo-EM Maps Xiongwu Wu, Bernard R. Brooks. NHLBI, National Institutes of Health, Bethesda, MD, USA. Ionotropic glutamate receptors (iGluRs) are cation channels that mediate signal transmission in the central nervous system by depolarizing the postdynaptic membrane in response to L-glutamate release from the presynaptic neuron. GluA2 is a subunit of AMPA, a major subtypes of iGluRs, which harbors a modular architecture composed of an amino-terminal domain (ATD), angonist-recognizing ligand-binding domain (LBD), a transmembrane domain (TMD) that forms the ion channel pore, and intracellular carboxyl-terminal domains. To understand the ion channel activation mechanism, many efforts have been dedicated to obtain the structures of GluA2 in various functional states. While ATD, LBD structures in the close and open states have been obtained through x-ray and cryo-EM, the TMD structure is only available in the close state because the open state TMD is difficult to capture with crystallography and its cryo-EM maps are low in resolution. The Low resolution cryo-EM maps of GluA2 in the close, open, and desensitized states have been obtained, which opens a window to investigate the structural mechanism of the ion channel activation. MapSGLD is a method to determine molecular structures from low resolution maps by utilizing structural information embedded in the force field. Through MapSGLD simulations with the low resolution maps of GluA2 in the close, open, and desensitized states, we determined the structures of GluA2 in the corresponding states. The structures show that LBD clamshell is open in the close state and closed in the open and desensitized states. The TMD ion pore is closed in the close and desensitized states but open in the open state. Comparing these structures we obtained an atomic detailed understanding of the ion channel activation mechanism. 2687-Pos Board B294 The C-Terminal Domain of Kv1.3 Interacts with KCNE4 to form Oligomeric Channels Sara R. Roig1, Laura Sole´2, Albert Vallejo-Gracia1, Daniel Sastre1, Antonio Serrano-Albarra´s1, Clara Serrano-Novillo1, Ramo´n Martı´nez-Ma´rmol3, Michael M. Tamkun2, Antonio Felipe1. 1 Biochemistry and Molecular Biomedicine, University of Barcelona, Barcelona, Spain, 2Department of Biomedical Sciences, Colorado State University, Fort Collins, CO, USA, 3Clem Jones Centre for Ageing Dementia Research, University of Queensland, Queensland, Australia. The voltage-gated potassium channel Kv1.3 plays crucial roles in the immune system. KCNE4, present in leukocytes, physically associates with Kv1.3 acting as a dominant negative subunit. KCNE4 inhibits Kþ currents and impairs membrane surface targeting. Although canonical KCNE-Kv7 interactions have been under intense investigation, the molecular determinants implicated in this essential Kv1.3/KCNE4 interaction remain unknown. Structural signatures between Kv7 and Kv1 channels display enormous differences. Our results indicate that the carboxy terminal of Kv1.3 is an essential and sufficient interacting domain for KCNE4. However, similar to what described for the Kv7.1 association, this motif apparently is not implicated in the modulation of the Kv1.3 activity. We also describe two independent and synergic mechanisms which potentiate the KCNE4-dependent intracellular retention of the Kv1.3/KCNE4 complex. First, KCNE4 association masks the YMVIEE motif at the C-terminal domain of the Kv1.3, which is crucial for the channel surface targeting; and second, we identify a basic endoplasmic reticulum retention signature in KCNE4. This element is transferred to the channelosome that further limits cell surface expression and triggers ER localization. Our results map specific molecular determinants which play a crucial role in the physiological function of Kv1.3 in leukocytes. Supported by MINECO, Spain (BFU2014-54928-R, BFU2015-70067-REDC and FEDER).

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2688-Pos Board B295 Structural and Functional Response of a Mechanosensitive K2P KD Channel to Asymmetric Membrane Tension Viwan Jarerattanachat, Michael V. Clausen, Prafulla Aryal, Elisabeth P. Carpenter, Mark S.P. Sansom, Stephen J. Tucker. University of Oxford, Oxford, United Kingdom. The mechanosensitive K2P Kþ channels play important roles in a range of diverse sensory processes, including touch, pain, and hearing. The ability of these channels to sense changes in pressure within the membrane is central to the process, but the structural and biophysical mechanisms underlying this remain unclear. Our previous studies have shown that the core transmembrane domains of the TREK-2 K2P channel are intrinsically sensitive to changes in bilayer tension and that the channel switches conformation from the ‘Down’ state to the ‘Up’ state in response to increasing levels of membrane stretch. However, the structure of the TREK-2 channel is highly asymmetric and increasing tension is expected to change the pressure profile equally in both the inner and outer leaflets of the bilayer. In this study we have examined the functional and structural effects of both negative and positive pressure on TREK-2 channels reconstituted into planar lipid bilayers, whilst their structural responses were investigated using MD simulations predicted to alter tension within only one leaflet of the bilayer. Our results demonstrate that TREK-2 exhibits a highly asymmetric response to changes in membrane tension, and we discuss the role that changes in the lateral pressure profile and local curvature of the membrane may play in this process. 2689-Pos Board B296 Fluctuation-Driven Transport in Bacterial Channels under Acidic Stress Marı´a L. Lo´pez, Marı´a Queralt-Martı´n, Vicente M. Aguilella, Antonio Alcaraz. Physics, University Jaume I, Castellon, Spain. Fluctuation-driven ion transport can be obtained in bacterial channels with the aid of different types of colored noise including biologically relevant Lorentzian one. Using the electrochemical rectification of the channel current as ratchet mechanism we observe transport of ions up their concentration gradient in conditions similar to that met in vivo, namely moderate pH gradients and asymmetrically charged lipid membranes. We find that depending on the direction of the concentration gradient the channel can pump either cations or anions from the diluted side to the concentrated one. We discuss the possible relevance of this phenomenon for the pH homeostasis of bacterial cells.

Ion Channels, Pharmacology, and Disease II 2690-Pos Board B297 Sodium Valproate Reverses Electrical Remodeling of Atrial Myocytes Isolated from CREM-IbDC-X Transgenic Mice C. Florentina Pluteanu, Beatrix Scholz, Jan S. Schulte, Frank U. M€uller. Inst. Pharmacology Toxicology, Westf€alische Wilhelms-Universit€at M€unster, M€unster, Germany. Patients that suffer from paroxysmal and chronic atrial fibrillation (AF) show increased levels of CREM-IbDC-X, a truncated form of cAMP response element modulator (CREM). In mice, cardiomyocyte directed over-expression of CREM-IbDC-X leads to atrial ectopy progressing to persistent AF. CREM represses gene transcription induced by cAMP response binding protein (CREB). Previously we showed that the treatment of mice with anticonvulsant sodium valproate (VPA), which also inhibits histone deacetylase (HDAC) classes I>II, prevented the atrial structural remodeling. The aim of this study was to investigate whether chronic VPA treatment also reverses electrical remodeling of atrial myocytes (AM) in CREM-IbDC-X mice. AM were isolated from 12-weeks old CREM-IbDC-X mice (TG) and wild type (WT) littermates, both treated for 7-weeks with VPA (0.4mM in the drinking water) or water (vehicle control). In TG mice, patch-clamp measurements showed that VPA treatment shortens action potentials (AP) by reducing AP duration (APD) at 50, 70 and 90% repolarization (7.350.6 vs 1151.2ms, 13.851.1 vs 20.552.2ms and 34.153 vs 4754.1ms, n=24 VPA-TG vs. n=37 untreated TG AM, p<0.05). In line with this observation total outward Kþ current densities were decreased by 36% in TG vs WT mice (p<0.05 vs WT) and were further increased by 38% only in VPA-TG mice (p<0.05 vs untreated TG). The inwardly rectifying IK1 currents were reduced by 35% in TG vs WT mice (p<0.05 vs WT) and increased by 47% in VPA-TG animals (p<0.05 vs untreated TG). In conclusion, VPA treatment reduced APD by increasing outward Kþ currents and may stabilize the resting membrane potential by increasing IK1 in atrial myocytes of CREM-IbDC-X mice. VPA has the potential to reverse atrial electrical remodeling, possibly via class I HDAC inhibition.