Studying lipid flip-flop in asymmetric liposomes using 1H NMR and TR-SANS.

The specific spatial and temporal distribution of lipids in membranes play a crucial role in determining the biochemical and biophysical properties of the system. In nature, the asymmetric distribution of lipids is a dynamic process with ATP-dependent lipid transporters maintaining asymmetry, and passive transbilayer diffusion, that is, flip-flop, counteracting it. In this chapter, two probe-free techniques, 1H NMR and time-resolved small angle neutron scattering, are described in detail as methods of investigating lipid flip-flop rates in synthetic liposomes that have been generated with an asymmetric bilayer composition.

Modeling asymmetric cell membranes at all-atom resolution.

Molecular dynamics (MD) simulations are a useful tool when studying the properties of membranes as they allow for a molecular view of lipid interactions with proteins, nucleic acids, or small molecules. While model membranes are usually symmetric in their lipid composition between leaflets and include a small number of lipid components, physiological membranes are highly complex and vary in the level of asymmetry. Simulation studies have shown that changes in leaflet asymmetry can alter the properties of a membrane. It is therefore necessary to carefully build asymmetric membranes to accurately simulate membranes. This chapter carefully describes the different methods for building asymmetric membranes and the advantages/disadvantages of each method. The simplest methods involve building a membrane with either an equal number of lipids per leaflet or an equal initial surface area (SA) estimated by the area per lipid. More detailed methods include combining two symmetric membranes of equal SA or altering an asymmetric membrane and adjusting the number of lipids after equilibration to minimize an observable such as differential stress (0-DS). More complex methods that require specific simulation software are also briefly described. The challenges and assumptions are listed for each method which should help guide the researcher to choose the best method for their unique MD simulation of an asymmetric membrane.

Predicting lipid sorting in curved membranes.

Most biological membranes are curved, and both lipids and proteins play a role in generating curvature. For any given membrane shape and composition, it is not trivial to determine whether lipids are laterally distributed in a homogeneous or inhomogeneous way, and whether the inter-leaflet distribution is symmetric or not. Here we present a simple computational tool that allows to predict the preference of any lipid type for membranes with positive vs. negative curvature, for any given value of curvature. The tool is based on molecular dynamics simulations of tubular membranes with hydrophilic pores. The pores allow spontaneous, barrierless flip-flop of most lipids, while also preventing differences in pressure between the inner and outer water compartments and minimizing membrane asymmetric stresses. Specifically, we provide scripts to build and analyze the simulations. We test the tool by performing simulations on simple binary lipid mixtures, and we show that, as expected, lipids with negative intrinsic curvature distribute to the tubule inner leaflet, the more so when the radius of the tubular membrane is small. Compared to other existing computational methods, relying on membrane buckles and tethers, our method is based on spontaneous inter-leaflet transport of lipids, and therefore allows to explore lipid distribution in asymmetric membranes. The method can easily be adapted to work with any molecular dynamics code and any force field.

Disruption of Transmembrane Phosphatidylserine Asymmetry by HIV-1 Incorporated SERINC5 Is Not Responsible for Virus Restriction.

Host restriction factor SERINC5 (SER5) incorporates into the HIV-1 membrane and inhibits infectivity by a poorly understood mechanism. Recently, SER5 was found to exhibit scramblase-like activity leading to the externalization of phosphatidylserine (PS) on the viral surface, which has been proposed to be responsible for SER5's antiviral activity. This and other reports that document modulation of HIV-1 infectivity by viral lipid composition prompted us to investigate the role of PS in regulating SER5-mediated HIV-1 restriction. First, we show that the level of SER5 incorporation into virions correlates with an increase in PS levels in the outer leaflet of the viral membrane. We developed an assay to estimate the PS distribution across the viral membrane and found that SER5, but not SER2, which lacks antiviral activity, abrogates PS asymmetry by externalizing this lipid. Second, SER5 incorporation diminished the infectivity of pseudoviruses produced from cells lacking a flippase subunit CDC50a and, therefore, exhibited a higher baseline level of surface-accessible PS. Finally, exogenous manipulation of the viral PS levels utilizing methyl-alpha-cyclodextrin revealed a lack of correlation between external PS and virion infectivity. Taken together, our study implies that the increased PS exposure to SER5-containing virions itself is not directly linked to HIV-1 restriction.

Regulation of iodine-glucose flip-flop in SW1736 anaplastic thyroid cancer cell line.

AIMS AND BACKGROUND: The alternative manner of iodide and glucose uptake found in different types of thyroid cancer, referred to flip-flop. ATC cells indicate low iodide uptake and high glucose uptake, which lack the morphology and genetic characteristics of well-differentiated tumors and become increasingly invasive. Importance placed on the discovery of innovative multi-targeted medicines to suppress the dysregulated signaling in cancer. In this research, we aimed to clarify molecular mechanism of Rutin as a phytomedicine on anaplastic thyroid cancer cell line based on iodide and glucose uptake. MATERIAL METHODS: The MTT test was employed to test cell viability. Iodide uptake assay was performed using a spectrophotometric assay to determine iodide uptake in SW1736 cells based on Sandell-Kolthoff reaction. For glucose uptake detection, ''GOD-PAP'' enzymatic colorimetric assay was applied to measure the direct glucose levels inside of the cells. Determination of NIS, GLUT1 and 3 mRNA expression in SW1736 cells was performed by qRT-PCR. Determination of NIS, GLUT1 and 3 protein levels in SW1736 cells was performed by western blotting. RESULTS: According to our results, Rutin inhibited the viability of SW1736 cells in a time- and dose-dependent manner. Quantitative Real-time RT-PCR analysis exposed that NIS mRNA levels were increased in Rutin treated group compared to the control group. Accordingly, western blot showed high expression of NIS protein and low expression of GLUT 1 and 3 in Rutin treated SW1736 cell line. Rutin increased iodide uptake and decreased glucose uptake in thyroid cancer cell line SW1736 compared to control group. CONCLUSION: Multiple mechanisms point to Rutin's role as a major stimulator of iodide uptake and inhibitor of glucose uptake, including effects at the mRNA and protein levels for both NIS and GLUTs, respectively. Here in, we described the flip-flop phenomenon as a possible therapeutic target for ATC. Moreover, Rutin is first documented here as a NIS expression inducer capable of restoring cell differentiation in SW1736 cell line. It also be concluded that GLUTs as metabolic targets can be blocked specifically by Rutin for thyroid cancer prevention and treatment.

Exploring Flip Flop memories and beyond: training Recurrent Neural Networks with key insights.

Training neural networks to perform different tasks is relevant across various disciplines. In particular, Recurrent Neural Networks (RNNs) are of great interest in Computational Neuroscience. Open-source frameworks dedicated to Machine Learning, such as Tensorflow and Keras have produced significant changes in the development of technologies that we currently use. This work contributes by comprehensively investigating and describing the application of RNNs for temporal processing through a study of a 3-bit Flip Flop memory implementation. We delve into the entire modeling process, encompassing equations, task parametrization, and software development. The obtained networks are meticulously analyzed to elucidate dynamics, aided by an array of visualization and analysis tools. Moreover, the provided code is versatile enough to facilitate the modeling of diverse tasks and systems. Furthermore, we present how memory states can be efficiently stored in the vertices of a cube in the dimensionally reduced space, supplementing previous results with a distinct approach.

Insertases scramble lipids: Molecular simulations of MTCH2.

Scramblases play a pivotal role in facilitating bidirectional lipid transport across cell membranes, thereby influencing lipid metabolism, membrane homeostasis, and cellular signaling. MTCH2, a mitochondrial outer membrane protein insertase, has a membrane-spanning hydrophilic groove resembling those that form the lipid transit pathway in known scramblases. Employing both coarse-grained and atomistic molecular dynamics simulations, we show that MTCH2 significantly reduces the free energy barrier for lipid movement along the groove and therefore can indeed function as a scramblase. Notably, the scrambling rate of MTCH2 in silico is similar to that of voltage-dependent anion channel (VDAC), a recently discovered scramblase of the outer mitochondrial membrane, suggesting a potential complementary physiological role for these mitochondrial proteins. Finally, our findings suggest that other insertases which possess a hydrophilic path across the membrane like MTCH2, can also function as scramblases.

Data on quantum dot cellular automata based flip flops for designing serial-in-serial-out shift register.

There has been remarkable research carried out on Nano-electronics where Quantum dot Cellular automata emerge as the forthcoming paradigm in computing. The QCA-based circuits are used in the computational Nano hardware to present computations at ultra-high speed. A systematic approach has been utilized to design the Serial in Serial out Shift (SISO) Register using JK flip flop (JK-FF) and D flip flop (D-FF). These flip flops were initially designed with lower complexity which is the dominant factor for designing any complex sequential circuit. The QCA based designs have been validated and subjected to simulation using the QCA Designer tool ver. 2.0.3.

Stochastic Adder Circuits with Improved Entropy Output.

Random pulse computing (RPC), the third paradigm along with digital and quantum computing, draws inspiration from biology, particularly the functioning of neurons. Here, we study information processing in random pulse computing circuits intended for the summation of numbers. Based on the information-theoretic merits of entropy budget and relative Kolmogorov-Sinai entropy, we investigate the prior art and propose new circuits: three deterministic adders with significantly improved output entropy and one exact nondeterministic adder that requires much less additional entropy than the previous art. All circuits are realized and tested experimentally, using quantum entropy sources and reconfigurable logic devices. Not only the proposed circuits yield a precise mathematical result and have output entropy near maximum, which satisfies the need for building a programmable random pulse computer, but also they provide affordable hardware options for generating additional entropy.

Photonic Crystal Flip-Flops: Recent Developments in All Optical Memory Components.

This paper reviews recent advancements in all-optical memory components, particularly focusing on various types of all-optical flip-flops (FFs) based on photonic crystal (PC) structures proposed in recent years. PCs, with their unique optical properties and engineered structures, including photonic bandgap control, enhanced light-matter interaction, and compact size, make them especially suitable for optical FFs. The study explores three key materials, silicon, chalcogenide glass, and gallium arsenide, known for their high refractive index contrast, compact size, hybrid integration capability, and easy fabrication processes. Furthermore, these materials exhibit excellent compatibility with different technologies like CMOS and fiber optics, enhancing their versatility in various applications. The structures proposed in the research leverage mechanisms such as waveguides, ring resonators, scattering rods, coupling rods, edge rods, switches, resonant cavities, and multi-mode interference. The paper delves into crucial properties and parameters of all-optical FFs, including response time, contrast ratio, and operating wavelength. Optical FFs possess significant advantages, such as high speed, low power consumption, and potential for integration, making them a promising technology for advancing optical computing and optical memory systems.

SEU Hardened D Flip-Flop Design with Low Area Overhead.

D flip-flop (DFF) is the basic unit of sequential logic in digital circuits. However, because of an internal cross-coupled inverter pair, it can easily appear as a single event upset (SEU) when hit by high-energy particles, resulting in the error in the value stored in the flip-flop. On this basis, a new structure D flip-flop is proposed in this paper. This flip-flop uses an asymmetric scheme in which the master-slave latch adopts different hardening structures. By sacrificing circuit speed in exchange for stronger SEU fortification capability, the SEU threshold of this structure is improved by 10 times compared to traditional D flip-flops. It has also been compared with Dual Interlocked Storage Elements (DICEs), and it saves the area cost of six transistors compared to the DICE structure. Under the same operating conditions, the average power consumption and peak power consumption are, respectively, 9.8% and 18.8% lower than those of the DICE circuit, making it suitable for soft radiation environments where high circuit speed is not a critical requirement.

Quantum Random Access Memory for Dummies.

Quantum Random Access Memory (QRAM) has the potential to revolutionize the area of quantum computing. QRAM uses quantum computing principles to store and modify quantum or classical data efficiently, greatly accelerating a wide range of computer processes. Despite its importance, there is a lack of comprehensive surveys that cover the entire spectrum of QRAM architectures. We fill this gap by providing a comprehensive review of QRAM, emphasizing its significance and viability in existing noisy quantum computers. By drawing comparisons with conventional RAM for ease of understanding, this survey clarifies the fundamental ideas and actions of QRAM. QRAM provides an exponential time advantage compared to its classical counterpart by reading and writing all data at once, which is achieved owing to storage of data in a superposition of states. Overall, we compare six different QRAM technologies in terms of their structure and workings, circuit width and depth, unique qualities, practical implementation, and drawbacks. In general, with the exception of trainable machine learning-based QRAMs, we observe that QRAM has exponential depth/width requirements in terms of the number of qubits/qudits and that most QRAM implementations are practical for superconducting and trapped-ion qubit systems.

SARS-CoV-2 fusion peptide sculpting of a membrane with insertion of charged and polar groups.

The fusion peptide of SARS-CoV-2 spike is essential for infection. How this charged and hydrophobic domain occupies and affects membranes needs clarification. Its depth in zwitterionic, bilayered micelles at pH 5 (resembling late endosomes) was measured by paramagnetic NMR relaxation enhancements used to bias molecular dynamics simulations. Asp830 inserted deeply, along with Lys825 or Lys835. Protonation of Asp830 appeared to enhance agreement of simulated and NMR-measured depths. While the fusion peptide occupied a leaflet of the DMPC bilayer, the opposite leaflet invaginated with influx of water and choline head groups in around Asp830 and bilayer-inserted polar side chains. NMR-detected hydrogen exchange found corroborating hydration of the backbone of Thr827-Phe833 inserted deeply in bicelles. Pinching of the membrane at the inserted charge and the intramembrane hydration of polar groups agree with theory. Formation of corridors of hydrated, inward-turned head groups was accompanied by flip-flop of head groups. Potential roles of the defects are discussed.

Modeling homeostatic and circadian modulation of human pain sensitivity.

Introduction: Mathematical modeling has played a significant role in understanding how homeostatic sleep pressure and the circadian rhythm interact to influence sleep-wake behavior. Pain sensitivity is also affected by these processes, and recent experimental results have measured the circadian and homeostatic components of the 24 h rhythm of thermal pain sensitivity in humans. To analyze how rhythms in pain sensitivity are affected by disruptions in sleep behavior and shifts in circadian rhythms, we introduce a dynamic mathematical model for circadian and homeostatic regulation of sleep-wake states and pain intensity. Methods: The model consists of a biophysically based, sleep-wake regulation network model coupled to data-driven functions for the circadian and homeostatic modulation of pain sensitivity. This coupled sleep-wake-pain sensitivity model is validated by comparison to thermal pain intensities in adult humans measured across a 34 h sleep deprivation protocol. Results: We use the model to predict dysregulation of pain sensitivity rhythms across different scenarios of sleep deprivation and circadian rhythm shifts, including entrainment to new environmental light and activity timing as occurs with jet lag and chronic sleep restriction. Model results show that increases in pain sensitivity occur under conditions of increased homeostatic sleep drive with nonlinear modulation by the circadian rhythm, leading to unexpected decreased pain sensitivity in some scenarios. Discussion: This model provides a useful tool for pain management by predicting alterations in pain sensitivity due to varying or disrupted sleep schedules.

A dopamine receptor D2 genetic polymorphism associated with transition to mental disorders in a cohort of individuals with at-risk mental state for psychosis.

OBJECTIVES: To test the association of 45 single nucleotide polymorphisms (SNPs) with transition to psychiatric disorders in a cohort of individuals at ultrahigh risk (UHR) mental state for psychosis. METHODS: Through general population screening, 88 non-help-seeking UHR subjects and 130 healthy control individuals were genotyped for 45 SNPs related to psychosis. They were followed for a mean of 2.5 years, and conversion to psychotic and to general psychiatric disorders was assessed. Genotype frequencies between controls, converters, and non-converters were analyzed. RESULTS: There were no differences in sociodemographics between controls and UHR. Also, UHR converters and non-converters had no differences in their baseline symptoms scores. The dopamine receptor D2 gene (DRD2) SNP rs6277 was significantly more common among UHR who transitioned to psychosis (p < 0.001) and to UHR who transitioned to any psychiatric disorders (p = 0.001) when compared to UHR who did not transition. The rs6277 T allele was related to psychiatric morbidity in a dose-response fashion, being significantly more frequent in UHR converters than UHR non-converters and control subjects (p = 0.003). CONCLUSION: Our findings suggest that rs6277 could potentially constitute a genetic marker of transition to psychiatric disorders in subjects with at-risk mental states, warranting further investigation in larger samples.

Alternative Splicing of the Flip/Flop Cassette and TARP Auxiliary Subunits Engage in a Privileged Relationship That Fine-Tunes AMPA Receptor Gating.

Alternative splicing of AMPA-type glutamate receptors (AMPARs) and allosteric modulation by auxiliary subunits, such as transmembrane AMPAR regulatory proteins (TARPs), are two important mechanisms that regulate the time course of glutamatergic neurotransmission. Prior work has shown that alternative splicing of the flip/flop cassette profoundly regulates TARP γ2 modulation, where flip receptor gating exhibits robust sensitivity to TARPs while flop isoforms are relatively insensitive to TARP modulation. Whether this splice variant-specific regulation extends to other auxiliary subunit families, such as cornichons (CNIHs), GSG1L, or CKAMPs, remains unknown. Here, we demonstrate that CNIH-3 modulation is unaffected by AMPAR alternative splicing due to inherent differences in how CNIH-3 and TARP γ2 modify channel gating. CNIH-3 slows receptor deactivation from the outset of current decay, consistent with structural evidence showing its point of contact at the level of the pore. In contrast, TARP γ2 acts via the KGK site of the ligand-binding domain (LBD) to slow the onset of desensitization. Although GSG1L and CKAMP44 primarily slow recovery from desensitization, their effects on channel gating are unaffected by alternative splicing, further underlining that structural events leading to the onset and recovery from desensitization are separable. Together, this work establishes that alternative splicing and TARP auxiliary subunits form a unique partnership that governs fast glutamatergic signaling at central synapses. Since proteomic studies suggest that all native AMPARs co-assemble with at least two TARPs, allosteric coupling between the flip/flop cassette and TARPs may represent a common design element in all AMPAR complexes of the mammalian brain.SIGNIFICANCE STATEMENT All fast excitatory neurotransmission in the mammalian brain is mediated by AMPA-type glutamate receptors (AMPARs). The time course of AMPAR gating can be regulated by two distinct mechanisms: alternative splicing of the flip/flop cassette and association with auxiliary subunits. Although these regulatory mechanisms have been well studied individually, it is not clear whether alternative splicing impacts auxiliary protein modulation of AMPARs. Here, we compare the four main families of AMPAR auxiliary subunits, transmembrane AMPAR regulatory proteins (TARPs; γ2), cornichons (CNIH-3), GSG1L and CKAMPs (CKAMP44), and find a privileged relationship between TARPs and the flip/flop cassette that is not shared by others. The flop cassette acts as a master switch to override TARP action, and this coupling represents a way to fine-tune AMPAR signaling.

Single-molecule Sequencing of an Animal Mitochondrial Genome Reveals Chloroplast-like Architecture and Repeat-mediated Recombination.

Recent advances in long-read sequencing technology have allowed for single-molecule sequencing of entire mitochondrial genomes, opening the door for direct investigation of the mitochondrial genome architecture and recombination. We used PacBio sequencing to reassemble mitochondrial genomes from two species of New Zealand freshwater snails, Potamopyrgus antipodarum and Potamopyrgus estuarinus. These assemblies revealed a ∼1.7 kb structure within the mitochondrial genomes of both species that was previously undetected by an assembly of short reads and likely corresponding to a large noncoding region commonly present in the mitochondrial genomes. The overall architecture of these Potamopyrgus mitochondrial genomes is reminiscent of the chloroplast genomes of land plants, harboring a large single-copy (LSC) region and a small single-copy (SSC) region separated by a pair of inverted repeats (IRa and IRb). Individual sequencing reads that spanned across the Potamopyrgus IRa-SSC-IRb structure revealed the occurrence of a "flip-flop" recombination. We also detected evidence for two distinct IR haplotypes and recombination between them in wild-caught P. estuarinus, as well as extensive intermolecular recombination between single-nucleotide polymorphisms in the LSC region. The chloroplast-like architecture and repeat-mediated mitochondrial recombination we describe here raise fundamental questions regarding the origins and commonness of inverted repeats in cytoplasmic genomes and their role in mitochondrial genome evolution.

Distinct roles of graphene and graphene oxide nanosheets in regulating phospholipid flip-flop.

Two-dimensional (2D) nanomaterials, such as graphene nanosheets (GNs) and graphene oxide nanosheets (GOs), could adhere onto or insert into a biological membrane, leading to a change in membrane properties and biological activities. Consequently, GN and GO become potential candidates for mediating interleaflet phospholipid transfer. In this work, molecular dynamics (MD) simulations were employed to investigate the effects of GN and GO on lipid flip-flop behavior and the underlying molecular mechanisms. Of great interest is that GN and GO work in opposite directions. The inserted GN can induce the formation of an ordered nanodomain, which dramatically elevates the free energy barrier of flipping phospholipids from one leaflet to the other, thus leading to a decreased lipid flip-flop rate. In contrast, the embedded GO can catalyze the transport of phospholipids between membrane leaflets by facilitating the formation of water pores. These results suggest that GN may work as an inhibitor of the interleaflet lipid translocation, while GO may play the role of scramblases. These findings are expected to expand promising biomedical applications of 2D nanomaterials.

A dopamine receptor D2 genetic polymorphism associated with transition to mental disorders in a cohort of individuals with at-risk mental state for psychosis

Objectives: To test the association of 45 single nucleotide polymorphisms (SNPs) with transition to psychiatric disorders in a cohort of individuals at ultrahigh risk (UHR) mental state for psychosis. Methods: Through general population screening, 88 non-help-seeking UHR subjects and 130 healthy control individuals were genotyped for 45 SNPs related to psychosis. They were followed for a mean of 2.5 years, and conversion to psychotic and to general psychiatric disorders was assessed. Genotype frequencies between controls, converters, and non-converters were analyzed. Results: There were no differences in sociodemographics between controls and UHR. Also, UHR converters and non-converters had no differences in their baseline symptoms scores. The dopamine receptor D2 gene (DRD2) SNP rs6277 was significantly more common among UHR who transitioned to psychosis (p < 0.001) and to UHR who transitioned to any psychiatric disorders (p = 0.001) when compared to UHR who did not transition. The rs6277 T allele was related to psychiatric morbidity in a dose-response fashion, being significantly more frequent in UHR converters than UHR non-converters and control subjects (p = 0.003). Conclusion: Our findings suggest that rs6277 could potentially constitute a genetic marker of transition to psychiatric disorders in subjects with at-risk mental states, warranting further investigation in larger samples.

BIAS-3D: Brain inspired attentional search model fashioned after what and where/how pathways for target search in 3D environment.

We propose a brain inspired attentional search model for target search in a 3D environment, which has two separate channels-one for the object classification, analogous to the "what" pathway in the human visual system, and the other for prediction of the next location of the camera, analogous to the "where" pathway. To evaluate the proposed model, we generated 3D Cluttered Cube datasets that consist of an image on one vertical face, and clutter or background images on the other faces. The camera goes around each cube on a circular orbit and determines the identity of the image pasted on the face. The images pasted on the cube faces were drawn from: MNIST handwriting digit, QuickDraw, and RGB MNIST handwriting digit datasets. The attentional input of three concentric cropped windows resembling the high-resolution central fovea and low-resolution periphery of the retina, flows through a Classifier Network and a Camera Motion Network. The Classifier Network classifies the current view into one of the target classes or the clutter. The Camera Motion Network predicts the camera's next position on the orbit (varying the azimuthal angle or "θ"). Here the camera performs one of three actions: move right, move left, or do not move. The Camera-Position Network adds the camera's current position (θ) into the higher features level of the Classifier Network and the Camera Motion Network. The Camera Motion Network is trained using Q-learning where the reward is 1 if the classifier network gives the correct classification, otherwise 0. Total loss is computed by adding the mean square loss of temporal difference and cross entropy loss. Then the model is trained end-to-end by backpropagating the total loss using Adam optimizer. Results on two grayscale image datasets and one RGB image dataset show that the proposed model is successfully able to discover the desired search pattern to find the target face on the cube, and also classify the target face accurately.