Three-Week-Old Rabbit Ventricular Cardiomyocytes as a Novel System to Study Cardiac Excitation and EC Coupling

Cardiac arrhythmias significantly contribute to cardiovascular morbidity and mortality. The rabbit heart serves as an accepted model system for studying cardiac cell excitation and arrhythmogenicity. Accordingly, primary cultures of adult rabbit ventricular cardiomyocytes serve as a preferable model to study molecular mechanisms of human cardiac excitation. However, the use of adult rabbit cardiomyocytes is often regarded as excessively costly. Therefore, we developed and characterized a novel low-cost rabbit cardiomyocyte model, namely, 3-week-old ventricular cardiomyocytes (3wRbCMs). Ventricular myocytes were isolated from whole ventricles of 3-week-old New Zealand White rabbits of both sexes by standard enzymatic techniques. Using wheat germ agglutinin, we found a clear T-tubule structure in acutely isolated 3wRbCMs. Cells were adenovirally infected (multiplicity of infection of 10) to express Green Fluorescent Protein (GFP) and cultured for 48 h. The cells showed action potential duration (APD90 = 253 ± 24 ms) and calcium transients similar to adult rabbit cardiomyocytes. Freshly isolated and 48-h-old-cultured cells expressed critical ion channel proteins: calcium voltage-gated channel subunit alpha1 C (Cavα1c), sodium voltage-gated channel alpha subunit 5 (Nav1.5), potassium voltage-gated channel subfamily D member 3 (Kv4.3), and subfamily A member 4 (Kv1.4), and also subfamily H member 2 (RERG. Kv11.1), KvLQT1 (K7.1) protein and inward-rectifier potassium channel (Kir2.1). The cells displayed an appropriate electrophysiological phenotype, including fast sodium current (INa), transient outward potassium current (Ito), L-type calcium channel peak current (ICa,L), rapid and slow components of the delayed rectifier potassium current (IKr and IKs), and inward rectifier (IK1). Although expression of the channel proteins and some currents decreased during the 48 h of culturing, we conclude that 3wRbCMs are a new, low-cost alternative to the adult-rabbit-cardiomyocytes system, which allows the investigation of molecular mechanisms of cardiac excitation on morphological, biochemical, genetic, physiological, and biophysical levels.

Chlorine-binding structures: role and organization in different proteins

The review focuses on chloride-binding structures in the proteins of bacteria, plants, viruses and animals. The structure and amino acid composition of the chloride-binding site and its role in the functioning of structural, regulatory, transport, receptor, channel proteins, transcription factors and enzymes are considered. Data on the important role of chloride-binding structures and chloride anions in the polymerization of fibrin are presented.

Ion Channels in Epithelial Dynamics and Morphogenesis

Mechanosensitive ion channels mediate the neuronal sensation of mechanical signals such as sound, touch, and pain. Recent studies point to a function of these channel proteins in cell types and tissues in addition to the nervous system, such as epithelia, where they have been little studied, and their role has remained elusive. Dynamic epithelia are intrinsically exposed to mechanical forces. A response to pull and push is assumed to constitute an essential part of morphogenetic movements of epithelial tissues, for example. Mechano-gated channels may participate in sensing and responding to such forces. In this review, focusing on Drosophila, we highlight recent results that will guide further investigations concerned with the mechanistic role of these ion channels in epithelial cells.

Introduction to the Theme on Membrane Channels

This volume of the Annual Review of Biochemistry contains three reviews on membrane channel proteins: the first by Szczot et al., titled The Form and Function of PIEZO2; the second by Ruprecht & Kunji, titled Structural Mechanism of Transport of Mitochondrial Carriers; and the third by McIlwain et al., titled Membrane Exporters of Fluoride Ion. These reviews provide nice illustrations of just how far evolution has been able to play with the basic helix-bundle architecture of integral membrane proteins to produce membrane channels and transporters of widely different functions.

Genome-Wide Analysis of Potassium Channel Genes in Rice: Expression of the OsAKT and OsKAT Genes under Salt Stress

Potassium (K+), as a vital element, is involved in regulating important cellular processes such as enzyme activity, cell turgor, and nutrient movement in plant cells, which affects plant growth and production. Potassium channels are involved in the transport and release of potassium in plant cells. In the current study, three OsKAT genes and two OsAKT genes, along with 11 nonredundant putative potassium channel genes in the rice genome, were characterized based on their physiochemical properties, protein structure, evolution, duplication, in silico gene expression, and protein–protein interactions. In addition, the expression patterns of OsAKTs and OsKATs were studied in root and shoot tissues under salt stress using real-time PCR in three rice cultivars. K+ channel genes were found to have diverse functions and structures, and OsKATs showed high genetic divergence from other K+ channel genes. Furthermore, the Ka/Ks ratios of duplicated gene pairs from the K+ channel gene family in rice suggested that these genes underwent purifying selection. Among the studied K+ channel proteins, OsKAT1 and OsAKT1 were identified as proteins with high potential N-glycosylation and phosphorylation sites, and LEU, VAL, SER, PRO, HIS, GLY, LYS, TYR, CYC, and ARG amino acids were predicted as the binding residues in the ligand-binding sites of K+ channel proteins. Regarding the coexpression network and KEGG ontology results, several metabolic pathways, including sugar metabolism, purine metabolism, carbon metabolism, glycerophospholipid metabolism, monoterpenoid biosynthesis, and folate biosynthesis, were recognized in the coexpression network of K+ channel proteins. Based on the available RNA-seq data, the K+ channel genes showed differential expression levels in rice tissues in response to biotic and abiotic stresses. In addition, the real-time PCR results revealed that OsAKTs and OsKATs are induced by salt stress in root and shoot tissues of rice cultivars, and OsKAT1 was identified as a key gene involved in the rice response to salt stress. In the present study, we found that the repression of OsAKTs, OsKAT2, and OsKAT2 in roots was related to salinity tolerance in rice. Our findings provide valuable insights for further structural and functional assays of K+ channel genes in rice.

Transport Proteins Enabling Plant Photorespiratory Metabolism

Photorespiration (PR) is a metabolic repair pathway that acts in oxygenic photosynthetic organisms to degrade a toxic product of oxygen fixation generated by the enzyme ribulose 1,5-bisphosphate carboxylase/oxygenase. Within the metabolic pathway, energy is consumed and carbon dioxide released. Consequently, PR is seen as a wasteful process making it a promising target for engineering to enhance plant productivity. Transport and channel proteins connect the organelles accomplishing the PR pathway—chloroplast, peroxisome, and mitochondrion—and thus enable efficient flux of PR metabolites. Although the pathway and the enzymes catalyzing the biochemical reactions have been the focus of research for the last several decades, the knowledge about transport proteins involved in PR is still limited. This review presents a timely state of knowledge with regard to metabolite channeling in PR and the participating proteins. The significance of transporters for implementation of synthetic bypasses to PR is highlighted. As an excursion, the physiological contribution of transport proteins that are involved in C4 metabolism is discussed.