Regulation Of Cation Transport Research Articles

Regulatory role of some protein kinases in signal transduction pathways in heart health and disease

Various protein kinases including protein kinase A (PKA), Ca2+-calmodulin kinase (CaMK), phosphoinositide 3-kinase (PI3K), protein kinase C (PKC) and mitogen-activated protein kinase (MAPK: ERK1/2, p38-MAPK and JNK) are integral part of different signal transduction pathways, which are known to regulate cardiac structure, function and metabolism. In addition, these signal transducing proteins are involved in the regulation of cation transport, cellular growth, gene expression, apoptosis and fibrosis by modifying the function of different target sites of subcellular organelles in the myocardium. However, the information regarding these signal transducing molecules is scattered and mechanisms of their involvement in diverse regulatory processes are poorly understood. While PKA, CaMK, PI3K and PKC are activated by different hormones and mechanical stimuli, MAPKs are activated by growth factors and some cellular stresses such as oxidative stress, inflammation and Ca2+-overload. Each type of these protein kinases is expressed in the form of two or more isozymes showing different biochemical characteristics and distinct biological functions. It has been demonstrated that all specific isoforms of these kinases produce both beneficial and detrimental effects on the heart, which are dependent upon the intensity and duration of stimulus for their activation. While PKA, PKC and CaMK are mainly involved in augmenting cardiac function as well as inducing cardiac hypertrophy and arrhythmias, PI3K is mainly involved in maintaining b-adrenoceptor function and inducing inflammation as well as arrhythmias. On the other hand, ERK1/2 mainly participate in the genesis of cardiac hypertrophy and cytoprotection whereas p38-MAPK and JNK are primarily involved in cardiac dysfunction, apoptosis and fibrosis. Since the activities of most protein kinases are increased under prolonged pathological conditions, a wide variety of their inhibitors have been shown to produce beneficial effects. However, extensive research needs to be carried out to understand the pathophysiology of different isoforms of each protein kinase as well as for the development of their isoform-specific inhibitors.

Comparative RNA sequencing for deciphering nodes of multiple abiotic stress tolerance in lentil (Lens culinaris Medikus)

Lentil is a highly proteinaceous crop whose productivity is severely hampered by various abiotic stresses. Transcriptome analysis unveils important metabolic pathways, regulatory genes and can help in identification of genomic resources which can be utilized for ameliorating abiotic stress tolerance. Therefore, the present study was undertaken to dissect transcriptomes of contrasting lentil genotypes under different stresses; drought, heat, salinity and alkalinity. Differential gene expression analysis and annotation were conducted by de novo transcriptomics and their functional correlation under diverse stresses was established using gene ontology analyses. Major differentially expressed genes (DEGs) which were found to be up-regulated under drought and heat stresses were involved in organic acid synthesis and translational regulation. Similarly, DEGs which were co-upregulated under salinity and alkalinity stresses belonged to abscisic acid (ABA) signaling, epigenetic regulation, sucrose transport, vesicular trafficking and regulation of cation transport at the root symplast-xylem interface. Data generated in this study can be used to elucidate converging tolerance pathways under multiple abiotic stresses. The information can be utilized in molecular breeding and gene pyramiding programmes of lentil and related species.

Cation transport goes with the flow

sodium reabsorption and potassium secretion in the cortical collecting duct are tightly regulated by luminal fluid flow ([9][1], [11][2]). Raising perfusion rates in isolated tubules exerts two forces on the apical surface of the epithelium, fluid shear stress and circumferential stretch. Flores et

Regulation of cation transport in the distal nephron by mechanical forces

Thiazide and loop diuretics induce renal K(+) secretion, often leading to renal K(+) wasting and hypokalemia. This phenomenon has been proposed to reflect an increase in delivery to and reabsorption of Na(+) by the distal nephron, with a resultant increase in the driving force for passive K(+) efflux across the apical membrane. Recent studies suggest that cellular mechanisms that lead to enhanced rates of Na(+) reabsorption as well as K(+) secretion in response to increases tubular flow rates are more complex. Increases in tubular flow rates directly enhance the activity of apical membrane Na(+) channels and indirectly activate a class of K(+) channels, referred to as maxi-K, that are functionally inactive under low flow states. This review addresses the role of biomechanical forces, generated by variations in urinary flow rate and tubular fluid volume, in the regulation of transepithelial Na(+) and K(+) transport in the distal nephron. The question of why the distal nephron has evolved to include a component of flow-dependent K(+) secretion is also addressed.

Mild spherocytosis and altered red cell ion transport in protein 4. 2-null mice.

Protein 4.2 is a major component of the red blood cell (RBC) membrane skeleton. We used targeted mutagenesis in embryonic stem (ES) cells to elucidate protein 4.2 functions in vivo. Protein 4. 2-null (4.2(-/-)) mice have mild hereditary spherocytosis (HS). Scanning electron microscopy and ektacytometry confirm loss of membrane surface in 4.2(-/-) RBCs. The membrane skeleton architecture is intact, and the spectrin and ankyrin content of 4. 2(-/-) RBCs are normal. Band 3 and band 3-mediated anion transport are decreased. Protein 4.2(-/-) RBCs show altered cation content (increased K+/decreased Na+)resulting in dehydration. The passive Na+ permeability and the activities of the Na-K-2Cl and K-Cl cotransporters, the Na/H exchanger, and the Gardos channel in 4. 2(-/-) RBCs are significantly increased. Protein 4.2(-/-) RBCs demonstrate an abnormal regulation of cation transport by cell volume. Cell shrinkage induces a greater activation of Na/H exchange and Na-K-2Cl cotransport in 4.2(-/-) RBCs compared with controls. The increased passive Na+ permeability of 4.2(-/-) RBCs is also dependent on cell shrinkage. We conclude that protein 4.2 is important in the maintenance of normal surface area in RBCs and for normal RBC cation transport.

Vph6 mutants of Saccharomyces cerevisiae require calcineurin for growth and are defective in vacuolar H(+)-ATPase assembly.

We have characterized a Saccharomyces cerevisiae mutant strain that is hypersensitive to cyclosporin A (CsA) and FK506, immunosuppressants that inhibit calcineurin, a serine-threonine-specific phosphatase (PP2B). A single nuclear mutation, designated cev1 for calcineurin essential for viability, is responsible for the CsA-FK506-sensitive phenotype. The peptidyl-prolyl cis-trans isomerases cyclophilin A and FKBP12, respectively, mediate CsA and FK506 toxicity in the cev1 mutant strain. We demonstrate that cev1 is an allele of the VPH6 gene and that vph6 mutant strains fail to assemble the vacuolar H(+)-ATPase (V-ATPase). The VPH6 gene was mapped on chromosome VIII and is predicted to encode a 181-amino acid (21 kD) protein with no identity to other known proteins. We find that calcineurin is essential for viability in many mutant strains with defects in V-ATPase function or vacuolar acidification. In addition, we find that calcineurin modulates extracellular acidification in response to glucose, which we propose occurs via calcineurin regulation of the plasma membrane H(+)-ATPase PMA1. Taken together, our findings suggest calcineurin plays a general role in the regulation of cation transport and homeostasis.

Regulation of cation transport in Saccharomyces cerevisiae by the salt tolerance gene HAL3.

Dynamic regulation of ion transport is essential for homeostasis as cells confront changes in their environment. The gene HAL3 encodes a novel component of this regulatory circuit in the yeast Saccharomyces cerevisiae. Overexpression of HAL3 improves growth of wild-type cells exposed to toxic concentrations of sodium and lithium and suppresses the salt sensitivity conferred by mutation of the calcium-dependent protein phosphatase calcineurin. Null mutants of HAL3 display salt sensitivity. The sequence of HAL3 gives little clue to its function. However, alterations in intracellular cation concentrations associated with changes in HAL3 expression suggest that HAL3 activity may directly increase cytoplasmic K+ and decrease Na+ and Li+. Cation efflux in S. cerevisiae is mediated by the P-type ATPase encoded by the ENA1/PMR24 gene, a putative plasma membrane Na+ pump whose expression is salt induced. Acting in concert with calcineurin, HAL3 is necessary for full activation of ENA1 expression. This functional complementarity is also reflected in the participation of both proteins in recovery from alpha-factor-induced growth arrest. Recently, HAL3 was isolated as a gene (named SIS2) which when overexpressed partially relieves loss of transcription of G1 cyclins in mutants lacking the protein phosphatase Sit4p. Therefore, HAL3 influences cell cycle control and ion homeostasis, acting in parallel to the protein phosphatases Sit4p and calcineurin.

Differential regulation of cation transport of vascular smooth muscle cells in a high glucose concentration milieu

To gain new insights into the pathogenesis of diabetic angiopathy, the influence of high glucose concentration on cation transport of vascular smooth muscle cell (VSMC) membrane was investigated by measuring Na, K and Ca transport in serially passaged cultured VSMC. (1) NaK pump activity, described as ouabain sensitive 86Rb uptake, and NaK cotransport, described as bumetanide sensitive 86Rb washout of VSMC, grown in high glucose concentration medium (460 mg/dl), was lower than that grown in normal glucose concentration medium (100 mg/dl). A smaller 5- N,N-hexamethylene amiloride (HMA) sensitive 22Na uptake (NaH antiport) in high glucose concentration medium accounted for this difference. (2) 45Ca uptake was also smaller in VSMC cultured in high glucose concentration medium. However, the washout rate constant for 45Ca was comparable between high and normal glucose cultured VSMC. (3) Both intracellular concentration of Na and cytosolic free Ca concentration ([Ca] i) of high glucose cultured VSMC were greater than normal glucose cultured VSMC. (4) Intracellular water volume based on the equilibrium distribution of 3- O-methyl-[ 14C]glucose was not different between normal and high glucose cultured VSMC. It is concluded that VSMC grown in high glucose concentration milieu manifests a decreased NaK, and Ca transport in conjunction with an increase in intracellular concentration of Na and [Ca] i. These results suggest that high glucose, per se, may alter membrane permeability to cations, possibly leading to changes in VSMC contractility and/or proliferation. This abnormality seen in the diabetic state may closely link to the pathogenesis of diabetic angiopathy, thus as a result risking hypertenion and vascular disease.

3 The membrane defect in hereditary stomatocytosis

Hereditary stomatocytosis and allied conditions represent a series of diseases in which abnormal movements of univalent cations across the plasma membrane play an important part in cellular disease. The primary problem lies not in the active transporters but in the basal permeability of the membrane, which is always increased, and the extent of the increase correlates with the cellular dysfunction. A number of structural abnormalities have been described in these membranes, but the most consistent and convincing is the deficiency of a hitherto uncharacterized integral membrane protein of molecular weight 31 kDa in the severe, 'overhydrated' form of the disease. The true function of this protein remains enigmatic, but its deficiency in this condition indicates that it may have a role in the regulation of cation transport.

Calcium-dependent regulation of cation transport in cultured human nonpigmented ciliary epithelial cells.

We performed 86Rb flux studies to examine Na-K-adenosinetriphosphatase (ATPase), Na-K-2Cl cotransporter, and potassium channel activity in an established cell line derived from human nonpigmented ciliary epithelium (ODM2). The elevation of intracellular calcium by A23187 (3 microM) or thapsigargin (200 nM) increased both ouabain-sensitive potassium (86Rb) uptake (Na-K-ATPase mediated) and ouabain-insensitive potassium (86Rb) uptake. The ouabain-insensitive component could be inhibited substantially by bumetanide (0.1 mM), suggesting the involvement of a Na-K-2Cl cotransporter. The increase of potassium (86Rb) uptake caused by thapsigargin could be prevented by the intracellular calcium buffer 1,2-bis(2-amino-phenoxy)ethane N,N,N',N'-tetraacetic acetoxymethyl ester (BAPTA/AM); in BAPTA/AM-treated cells, the thapsigargin stimulation of the bumetanide-sensitive portion of 86Rb uptake was abolished. After A23187 (5 microM), the 86Rb efflux rate was significantly increased; the increase could be blocked partially by quinidine (0.1 mM) and partially by bumetanide, suggesting that potassium channels and the Na-K-2Cl cotransporter contribute to the effect. We propose that the cell potassium loss after activation of quinidine-sensitive potassium channels is involved in the calcium-induced activation of Na-K-ATPase because 0.1 mM quinidine and 100 mM external potassium both markedly inhibited the A23187-induced increases of the ouabain-sensitive component of potassium (86Rb) uptake. Calcium-induced stimulation of the Na-K-2Cl cotransporter may not be linked to channel activation.

Regulation of cation transport by low doses of glucocorticoids in in vivo adrenalectomized rat colon.

A dose response curve for glucocorticoid-induced proximal and distal colonic cation transport in vivo was established in adrenalectomized rats. All doses (0.5-50 nmol/100 g body wt) stimulated sodium absorption. Distal sodium absorption did not saturate at dexamethasone levels that saturate the glucocorticoid receptor but also bind to greater than 35% of aldosterone receptors. Saturation of the pure glucocorticoid response occurred in both segments with RU26988, a synthetic glucocorticoid that does not occupy aldosterone receptors. Maximum velocities for pure glucocorticoid-induced sodium absorption were 15 and 16 mu eq/min per g dry tissue, and Michaelis constants (Km) were 4.2 and 4.6 X 10(-9) mol/liter for proximal and distal colon. Kms are similar to the dissociation constant for the colonic glucocorticoid receptor and too low for significant aldosterone receptor occupancy. Dexamethasone increased sodium absorption significantly within 30 min of injection, suggesting the response is not dependent on new protein synthesis. Similar time and dose responses in proximal and distal colon suggest glucocorticoids stimulate the same pathway in both segments.

The effect of X-irradiation on cation transport in rabbit lens

The effect of X-irradiation on the active transport and diffusion of 86 Rb was investigated at various times following exposure to a single dose of 2000 rad. In addition the levels of ascorbic acid, glutathione (GSH) and superoxide dismutase (SOD) in lens and ascorbic acid and H 2 O 2 in aqueous humor were studied as a function of time after irradiation. A decrease in active transport of Rb and an increase in permeability was first observed after 4 weeks following X-ray; after 7 weeks, the active transport decreased by 80% and permeability increased by 44 times the control value. Despite the dramatic changes in cation transport and permeability, the hydration of the lens remained unaffected until the development of maturre cataracts, which appeared 8–9 weeks following exposure to X-ray. In contrast to the delayed effect of X-ray on hydration and cation transport, there was a gradual decrease in the level of GSH. The values of ascorbate in aqueous humor were significantly lower in the X-rayed eyes than the contralateral controls at all time periods after irradiation, while a lower concentration of H 2 O 2 in the aqueous of X-rayed eyes was first observed 4 weeks after exposure. The activity of SOD in the lens was unaffected at any time after X-irradiation. The decrease in the level of GSH in capsule and epithelium was less pronounced than in the decapsulated or whole lens The possible role of GSH in the epithelium in the regulation of cation transport and hydration is discussed.

Effect of exogenous ATP on sodium transport in mammalian red cells.

The effect of exogenous adenosine triphosphate (ATP) and other nucleotides on the transport of Na in various mammalian red cells has been studied. While they have no effect on the transport of Na in human and cat red cells, in dog red cells adenosine and its mono-, di- and triphosphorylated forms were found to increase Na-influx. Of these, ATP has the most striking effect, causing a more than 8-fold increase at a concentration of 0.6 mM and exerting this effect at a dose range of 10(-5) to 10(-3) M. The effect of ATP is rapid (less than 5 minutes) and can be reversed by washing or the addition of calcium or magnesium. In contrast to the adenosine series other phosphorylated nucleotides (GTP, CTP, UDP, GDP and cAMP) have no effect. The well known volume dependent Na-transport in these cells is reversed in the presence of 0.6 mM ATP. It is suggested that ATP acts on passive cation movements either by chelation of membrane charge or by a direct interaction with membrane proteins and may be involved in the volume regulation of cation transport in the dog erythrocyte.

The effect of salt adaptation on the permeability and cation selectivity of the goldfish intestinal epithelium.

1. The short-term uptake of Na by the goldfish mucosa was compared using both inulin and choline as markers of extracellular space. The results were virtually identical, the distribution of both choline and inulin increasing rapidly to measure a space at 1 min which then remained nearly constant during a following 4 min incubation.2. Using inulin as space marker, the uptake of various alkali metal cations was determined from a 1 min contact with the mucosa. The relative rates of uptake were Tl > K > Rb > Cs > Na > Li, with a low selectivity ratio, the range of permeabilities being no greater than 5.3. The selectivity sequence was the same in both salt and fresh-water adapted fish. Of the alkali metal cations tested, only Na showed a significantly decreased uptake on adaptation to salt.4. Isolated intestinal preparations from salt-adapted fish showed a reduced short-circuit current compared with fish adapted to fresh water, the values being 12.3 +/- 1.2 and 35.7 +/- 1.5 muA cm(-2) respectively. In both cases the short-circuit current was equivalent to the net transport of Na measured isotopically.5. In Krebs-Henseleit medium, the measured tissue resistance was approximately 100 Omega cm(-2) for both salt and fresh-water adapted fish.6. It is concluded that regulation of cation transport in goldfish intestinal epithelium is specific for Na and mediated primarily through cellular rather than extracellular pathways.