Cellular uptake of vitamin B: Role and fate of TCblR/CD320, the transcobalamin receptor.

Cellular uptake of vitamin B (cobalamin, Cbl) is mediated by a cell surface receptor (TCblR/CD320) that binds transcobalamin (TC) saturated with Cbl. TC is secreted by the vascular endothelium, has a relatively short half-life, binds Cbl with high affinity and presents the vitamin to the receptor for cellular uptake. Here we show binding and internalization of the TC-Cbl complex along with its' receptor (TCblR) in several human cell lines.

Regulation of Na,K-ATPase alpha1 subunit gene transcription in response to low K(+): role of CRE/ATF- and GC box-binding proteins.

Na,K-ATPase expression is upregulated in mammalian cells as a consequence of persistent inhibition of Na,K-ATPase enzymatic activity by low external K(+). We previously demonstrated that exposure of neonatal rat cardiac myocytes to low K(+) increased Na,K-ATPase alpha1 subunit mRNA content and promoter activity. In this work, we utilized transient transfection studies with rat Na,K-ATPase alpha1 subunit 5'-flanking region deletion plasmids to identify DNA sequences required for low K(+)-mediated stimulation of alpha1 subunit promoter expression in cardiac myocytes.

Divergent signaling pathways mediate induction of Na,K-ATPase alpha1 and beta1 subunit gene transcription by low potassium.

Prolonged inhibition of Na,K-ATPase enzymatic activity by exposure of a variety of mammalian cells to low external K+ yields a subsequent adaptive up-regulation of Na,K-ATPase expression. The aim of this study was to examine the intracellular signal transduction system that is responsible for mediating increased Na,K-ATPase subunit gene expression in primary cultures of neonatal rat cardiac myocytes. In this work, we show long-term inhibition of Na,K-ATPase function with 0.

Angiotensin II regulation of the Na+ pump involves the phosphatidylinositol-3 kinase and p42/44 mitogen-activated protein kinase signaling pathways in vascular smooth muscle cells.

This investigation used primary cultured rat vascular smooth muscle cells to examine angiotensin II (Ang II) regulation of Na(+), K(+)-ATPase (Na(+) pump) activity, and Na(+) pump alpha(1)- and beta(1)-subunit gene transcription. This regulation was mediated through both phosphatidylinositol-3 kinase (PI3K) and p42/44 mitogen-activated protein kinase (p42/44(MAPK)) signaling pathways. Both acute (10 min) and prolonged (24 h) treatment with Ang II stimulated Na(+) pump activity.

Heart disease in diabetic patients.

Both type 1 and type 2 diabetic patients have an increased incidence of ischemic heart disease and congestive heart failure. Cardiovascular disease accounts for up to 80% of the excess mortality in patients with type 2 diabetes. The burden of cardiovascular disease is especially pronounced in diabetic women.

Regulation of Na/K-ATPase beta1-subunit gene expression by ouabain and other hypertrophic stimuli in neonatal rat cardiac myocytes.

Partial inhibition of Na/K-ATPase by ouabain causes hypertrophic growth and regulates several early and late response genes, including that of Na/K-ATPase alpha3 subunit, in cultured neonatal rat cardiac myocytes. The aim of this work was to determine whether ouabain and other hypertrophic stimuli affect Na/K-ATPase beta1 subunit gene expression. When myocytes were exposed to non-toxic concentrations of ouabain, ouabain increased beta1 subunit mRNA in a dose- and time-dependent manner.

Up-regulation of Na,K-ATPase beta 1 transcription by hyperoxia is mediated by SP1/SP3 binding.

The sodium pump, Na,K-ATPase, is an important protein for maintaining intracellular ion concentration, cellular volume, and ion transport and is regulated both transcriptionally and post-transcriptionally. We previously demonstrated that hyperoxia increased Na,K-ATPase beta(1) gene expression in Madin-Darby canine kidney (MDCK) cells. In this study, we identify a DNA element necessary for up-regulation of the Na,K-ATPase beta(1) transcription by hyperoxia and evaluate the nuclear proteins responsible for this up-regulation.

Regulation of Na,K-ATPase beta 1 subunit gene transcription by low external potassium in cardiac myocytes. Role of Sp1 AND Sp3.

Expression of Na,K-ATPase activity is up-regulated in cells incubated for extended intervals in the presence of low external K(+). Our previous data showed that exposure of cardiac myocytes to low K(+) increased the steady-state abundance of Na,K-ATPase beta1 subunit mRNA. In the present study we determined that incubation of primary cultures of neonatal rat cardiac myocytes with low K(+) augmented Na,K-ATPase beta1 gene expression at a transcriptional level and that this effect required extracellular Ca(2+).

Selective induction of Na,K-ATPase alpha3 subunit mRNA abundance in cardiac myocytes by retinoic acid.

Retinoic acid (RA) is a high affinity ligand for a nuclear receptor which regulates transcription in target cells. Specific effects of RA on cardiac development and myocardial cell hypertrophy have been demonstrated; however, little information exists concerning RA-mediated regulation of cardiac genes. This study was initiated to investigate whether RA regulates Na,K-ATPase subunit gene expression in primary cultures of neonatal rat cardiac myocytes.

Characterization of a negative thyroid hormone response element in the rat sodium, potassium-adenosine triphosphatase alpha3 gene promoter.

The thyroid hormone L-T3 elicits either a stimulatory or an inhibitory effect on expression of the Na,K-adenosine triphosphatase alpha3-subunit gene in primary cultures of neonatal rat cardiac myocytes. The present study was undertaken to characterize a negative thyroid hormone response element present within the rat Na,K-adenosine triphosphatase alpha3-subunit gene proximal promoter. Transient transfection assays indicated that the DNA-binding domain of thyroid hormone receptor was essential for mediating repression of alpha3 gene transcription by thyroid hormone.

Alterations in cardiac gene expression during ventricular remodeling following experimental myocardial infarction.

Following myocardial Infarction (MI) the heart undergoes a process of remodeling characterized by considerable hypertrophy of the non-infarcted myocardium. We have recently characterized the molecular basis of key electrophysiologic alterations that may provide insight into the arrhythmogenecity of post-MI remodeled hypertrophied myocardium. To further characterize other key alterations in the pattern of cardiac gene expression in a time-dependent manner, we have measured mRNA and immunoreactive protein levels of selective cardiac genes in the remodeled hypertrophied left-ventricular (LV) myocardium of rats, 3 and 21 days after left-coronary ligation and compared the results with sham-operated rats.

Regulation of Na-K-ATPase gene expression by hyperoxia in MDCK cells.

Na-K-ATPase plays a central role in a variety of physiological processes, including ion transport and regulation of cell volume. Our previous data showed that hyperoxia increased the expression of Na-K-ATPase alpha 1 and beta 1 mRNA in lung type II cells. We similarly show that hyperoxia (> or = 95% O2 for 24-48 h) increased steady-state mRNA levels in both Na-K-ATPase subunits in Madin-Darby canine kidney (MDCK) cells.

Negative regulation of the rat Na-K-ATPase alpha 3-subunit gene promoter by thyroid hormone.

Na-K-ATPase alpha 3-subunit mRNA levels are both positively and negatively controlled by thyroid hormone [3,5,3'triiodothyronine (T3)] in primary cultures of neonatal rat cardiac myocytes. In this study, transient transfection analysis indicated that two regions of the rat alpha 3 gene between nucleotides -116 and -6 and -6 and +80 conferred T3-mediated inhibition of reporter gene expression. Electrophoretic mobility shift assays showed specific binding of T3 receptor monomers and T3 receptor-retinoid X receptor heterodimers at each alpha 3 gene negative T3-response region.

Low external K+ regulates Na,K-ATPase alpha 1 and beta 1 gene expression in rat cardiac myocytes.

Incubation of mammalian cells in medium containing low K+ is often associated with an increase in Na,K-ATPase activity and content. In the present studies, we investigated the effect of low K+ on the regulation of Na,K-ATPase alpha 1 and beta 1 gene expression in primary cultures of neonatal rat cardiac myocytes. Northern blot hybridization analysis indicated that exposure of cardiocytes to a medium containing 0.

Thyroid hormone regulation of Na,K-ATPase alpha 2 gene expression in cardiac myocytes.

Thyroid hormone (T3) stimulates Na,K-ATPase activity and alpha and beta subunit mRNA abundances in myocardial cells in vivo and in vitro. In this study, we used transient transfection and nuclear run-on assays to determine whether T3 regulates the transcription rate of the Na,K-ATPase alpha 2 subunit gene. Primary cultures of neonatal rat cardiac myocytes were incubated with 100 nM T3 for 1, 3, and 6 d, and alpha 2 mRNA levels were measured by Northern blot hybridization analysis.

Na,K-ATPase in several tissues of the rat: tissue-specific expression of subunit mRNAs and enzyme activity.

The relative contents of Na,K-ATPase subunit mRNAs in rat renal cortex, ventricular myocardium, skeletal muscle (hind limb), liver and brain (cerebrum) were measured. Expressed per unit DNA, mRNA alpha 1 content was approximately 2-fold greater in the kidney and brain as compared to either heart, skeletal muscle or liver. The hierarchy of mRNA alpha 2 expression was brain > skeletal muscle > heart, whereas mRNA alpha 3 was restricted to brain.

Regulation of Na,K-ATPase beta 1 mRNA content by thyroid hormone in neonatal rat cardiac myocytes.

Incubation of primary cultures of neonatal rat cardiac myocytes in the presence of 100 nM triiodothyronine (T3) resulted in a three- to fivefold increase in the content of Na,K-ATPase beta 1 subunit mRNA which was maximal at 1 d of exposure to hormone. To investigate the mechanism by which T3 stimulates the abundance of beta 1 mRNA, transient transfection experiments were conducted with a chimeric gene containing a portion of the 5' end of the rat beta 1 gene linked to a luciferase reporter gene. We found no effect of T3 on chimeric gene activity either in the absence or presence of cotransfected T3 receptor.

Characterization of the 5' flanking region of the rat Na+/K(+)-ATPase beta 1 subunit gene.

We have isolated the 5' end of the rat Na+/K(+)-ATPase beta 1 subunit gene. A genomic fragment containing 817 bp of the 5' flanking sequence, exon 1 and 479 bp of intron 1 was sequenced. The 5' flanking region contains a potential TATA box and several putative CAAT and GC boxes.

Thyroidal enhancement of rat myocardial Na,K-ATPase: preferential expression of alpha 2 activity and mRNA abundance.

In hypothyroid rat myocardium, the low-ouabain-sensitivity Na,K-ATPase activity had a KI = 10(-4) M and accounted for approximately 95% of the enzyme activity, while the high-ouabain-sensitivity activity contributed approximately 5% to the total activity, with a KI = 3 x 10(-7) M. mRNA alpha 1 was 7.2- and 5.

Thyroid hormone induction of Na(+)-K(+)-ATPase and its mRNAs in a rat liver cell line.

The expression of mRNAs encoding the alpha- and beta-subunits of Na(+)-K(+)-ATPase (Na(+)-K+ pump) was examined in a rat liver cell line, Clone 9, in various thyroidal states. Northern blot analysis of total RNA isolated from cells incubated in hypothyroid serum-containing medium revealed the expression of mRNAs encoding Na(+)-K(+)-ATPase alpha 1-(mRNA alpha 1) and beta- (mRNA beta) subunits; mRNAs encoding the alpha 2- and alpha 3-subunits were undetectable. There was a discrepancy in the abundance of mRNA alpha 1 relative to mRNA beta such that mRNA alpha 1 exceeded the sum of the multiple mRNA beta bands by approximately 35-fold.

Thyroidal regulation of rat renal and hepatic Na,K-ATPase gene expression.

Na,K-ATPase activity, Na,K-ATPase alpha- and beta-subunit mRNA abundance (mRNA alpha and mRNA beta), and gene transcription rates were determined in kidney cortex and liver of hypothyroid and triiodothyronine (T3)-treated rats. In hypothyroid rats, Na,K-ATPase activity (expressed per unit of DNA) was 3.6-fold greater in kidney cortex than liver, and the abundance of mRNA alpha and mRNA beta in kidney cortex exceeded that of liver by 2.

Increased abundance of Na+-K+-ATPase mRNAs in response to low external K+.

Exposure of ARL 15 cells, an established line from adult rat liver, to external K+ concentrations less than 1 mM for 24 h increases Na+-K+ pump abundance (Na+-K+-ATPase) (J. Gen. Physiol.

Kinetic analysis of Na,K-activated adenosine triphosphatase induced by low external K+ in a rat liver cell line.

Exposure of ARL 15 cells to medium containing reduced concentrations of K+ (0.65 mM) elicited a 50-100% increase in Na,K-ATPase activity. The inhibition by ouabain of both the basal and the induced enzyme conformed to a single-site model (KI = 1 x 10(-4) M).