Recent studies have ascribed many non-pumping functions to the Na/K-ATPase. We show here that graded knockdown of cellular Na/K-ATPase alpha1 subunit produces a parallel decrease in both caveolin-1 and cholesterol in light fractions of LLC-PK1 cell lysates. This observation is further substantiated by imaging analyses, showing redistribution of cholesterol from the plasma membrane to intracellular compartments in the knockdown cells. Moreover, this regulation is confirmed in alpha1(+/-) mouse liver. Functionally, the knockdown-induced redistribution appears to affect the cholesterol sensing in the endoplasmic reticulum, because it activates the sterol regulatory element-binding protein pathway and increases expression of hydroxymethylglutaryl-CoA reductase and low density lipoprotein receptor in the liver. Consistently, we detect a modest increase in hepatic cholesterol as well as a reduction in the plasma cholesterol. Mechanistically, alpha1(+/-) livers show increases in cellular Src and ERK activity and redistribution of caveolin-1. Although activation of Src is not required in Na/K-ATPase-mediated regulation of cholesterol distribution, the interaction between the Na/K-ATPase and caveolin-1 is important for this regulation. Taken together, our new findings demonstrate a novel function of the Na/K-ATPase in control of the plasma membrane cholesterol distribution. Moreover, the data also suggest that the plasma membrane Na/K-ATPase-caveolin-1 interaction may represent an important sensing mechanism by which the cells regulate the sterol regulatory element-binding protein pathway.
"In the present report we claim an essential and novel role for CypB in the maturation and trafficking of Na/Kα1-ATPase pump through the secretory pathway. A series of important studies have demonstrated non-pumping functions for the Na/K-ATPase that include regulation of oncogenic transformation , epithelial to mesenchymal cell transition , tight junction formation  and control of the plasma membrane cholesterol distribution . The novel interaction found between CypB and Na/K-β1 and the importance of CypB as a Na/K-ATPase functional modulator should be very relevant to further understand the role of CypB in renal and extrarenal pathophysiology. "
[Show abstract][Hide abstract] ABSTRACT: Cyclophilins (Cyps), the intracellular receptors for Cyclosporine A (CsA), are responsible for peptidyl-prolyl cis-trans isomerisation and for chaperoning several membrane proteins. Those functions are inhibited upon CsA binding. Albeit its great benefits as immunosuppressant, the use of CsA has been limited by undesirable nephrotoxic effects, including sodium retention, hypertension, hyperkalemia, interstial fibrosis and progressive renal failure in transplant recipients. In this report, we focused on the identification of novel CypB-interacting proteins to understand the role of CypB in kidney function and, in turn, to gain further insight into the molecular mechanisms of CsA-induced toxicity. By means of yeast two-hybrid screens with human kidney cDNA, we discovered a novel interaction between CypB and the membrane Na/K-ATPase β1 subunit protein (Na/K-β1) that was confirmed by pull-down, co-immunoprecipitation and confocal microscopy, in proximal tubule-derived HK-2 cells. The Na/K-ATPase pump, a key plasma membrane transporter, is responsible for maintenance of electrical Na+ and K+ gradients across the membrane. We showed that CypB silencing produced similar effects on Na/K-ATPase activity than CsA treatment in HK-2 cells. It was also observed an enrichment of both alpha and beta subunits in the ER, what suggested a possible failure on the maturation and routing of the pump from this compartment towards the plasma membrane. These data indicate that CypB through its interaction with Na/K-β1 might regulate maturation and trafficking of the pump through the secretory pathway, offering new insights into the relationship between cyclophilins and the nephrotoxic effects of CsA.
PLoS ONE 11/2010; 5(11):e13930. DOI:10.1371/journal.pone.0013930 · 3.23 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Objective: Endogenous digoxin has been related to hemispheric dominance. Right hemispheric dominant individuals were hyperdigoxinemic, left hemispheric d o m i n a n t i n d i v i d u a l s w e r e h y p o d i g o x i n e m i c a n d b i h e m i s p h e r i c d o m i n a n t i n d i v i d u a l s w e r e normodigoxinemic. The possibility of endogenous digoxin synthesis by actinide based primitive organism like archaea with a mevalonate pathway and cholesterol catabolism was considered. An actinide dependent shadow biosphere of archaea and viroids in normal humans and disease states is described. The intracellular endosymbionts archaea and their intron derived viroids constitute the third element regulating the human body. Ayurveda, the traditional Indian System of Medicine, deals with the theory of the three tridosha states (both physical and psychological): Vata, Pitta, and Kapha. The Kapha state has been demonstrated as equivalent to right hemispheric dominant hyperdigoxinemic state. The Pitta state has been demonstrated as equivalent to the left hemispheric dominant hypodigoxinemic state. The Vata state has been demonstrated as equivalent to the bihemispheric dominant normodigoxinemic state  . The study assessed actinidic archaea and viroids in the tridosha states of Ayurveda. The results are presented in this paper. Methods: The following groups were included in the study:-(I) right handed-left hemispheric dominant-pitta group, (II) left handed-right hemispheric dominant-kapha group and (III) amphidextrous-bihemispheric dominant-vata group of individuals. Cholesterol substrate was added to the plasma of the patients and the generation of cytochrome F420, free RNA, free DNA, polycyclic aromatic hydrocarbon, hydrogen peroxide, serotonin, pyruvate, ammonia, glutamate, cytochrome C, hexokinase, ATP synthase, HMG CoA reductase, digoxin and bile acids were studied. The changes with the addition of antibiotics and rutile to the patient's plasma were also studied. Results: The plasma of the bihemispheric dominant vata group showed detectable levels of the above mentioned parameters after incubation for 1 hour and addition of cholesterol substrate resulted in still further increase in these parameters. The addition of antibiotics to the bihemispheric dominant vata group caused a decrease in all the parameters while addition of rutile increased their levels. The plasma of right hemispheric dominant kapha group showed a significant increase in the above mentioned parameters as compared to bihemispheric dominant vata group. The addition of antibiotics to the right hemispheric dominant kapha group caused a decrease in all the parameters while addition of rutile increased their levels but the extent of change was more in right hemispheric dominant kapha group as compared to bihemispheric dominant vata group. The plasma of left hemispheric dominant pitta group showed a significant decrease in the above mentioned parameters as compared to the bihemispheric dominant vata group. The addition of antibiotics to the left hemispheric dominant pitta group caused a decrease in all the parameters while addition of rutile increased their levels but the extent of change was less in left hemispheric dominant pitta group as compared to bihemispheric dominant vata group. Conclusion: The third element formed of intracellular archaea and viroidal symbiosis determines hemispheric dominance and tridoshas. Also archaeal cholesterol synthesis and cholesterol catabolism determines hemispheric dominance and tridoshas. The archaea and viroidal density is high in right hemispheric dominant kapha group, intermediate in bihemispheric dominant vata group and low in left hemispheric dominant pitta group.
[Show abstract][Hide abstract] ABSTRACT: Aim: A hypothesis regarding cholesterol based abiogenesis and its role in the evolution of universe is elucidated. Endomyocardial fibrosis along with the root wilt disease of coconut is endemic to Kerala with its radioactive actinide beach sands. Actinides like rutile as well as organisms like phytoplasmas and viroids have been implicated in the etiology of these diseases. Actinidic archaea has been related to the pathogenesis of schizophrenia, malignancy, metabolic syndrome x, autoimmune disease and neuronal degeneration. An actinide dependent shadow biosphere of archaea and viroids in the above mentioned disease states is described. Actinide based primitive organism like archaea have a mevalonate pathway and cholesterol catabolism. This points to cholesterol as the primal prebiotic molecule and evolution of actinidic archaea and viroids from a primitive isoprenoid organism. The role of magnetotactic actinidic archaea in the origin of the biological universe is discussed. Methods: Plasma from fasting heparinised blood was used and the experimental protocol was as follows (I) Plasma+phosphate buffered saline, (II) same as I+cholesterol substrate, (III) same as II+rutile 0.1 mg/ml, (IV) same as II+ciprofloxacine and doxycycline each in a concentration of 1 mg/ml. The following estimations were carried out:-Cytochrome F420, free RNA, free DNA, polycyclic aromatic hydrocarbon, hydrogen peroxide, dopamine, serotonin, pyruvate, ammonia, glutamate, cytochrome C, hexokinase, ATP synthase, HMG CoA reductase, digoxin and bile acids. Results: Plasma of control subjects showed increased levels of the above mentioned parameters with after incubation for 1 hour and addition of cholesterol substrate resulted in still further significant increase in these parameters. The plasma of patients showed similar results but the extent of increase was more. The addition of antibiotics to the control plasma caused a decrease in all the parameters while addition of rutile increased their levels. The addition of antibiotics and rutile to the patient's plasma produced the same changes but the extent of change was more in patient's sera as compared to controls. Conclusion: An actinide dependent shadow biosphere of archaea and viroids in the above mentioned disease states is described. Metal actinides in beach sands have been postulated to play a role in abiogenesis. A hypothesis of cholesterol as the primal prebiotic molecule synthesised on actinide surfaces with all other biomolecules arising from it and a self replicating cholesterol lipid organism as the initial life form is presented. A cholesterol based theory of abiogenesis and evolution of actinidic archaea and viroids from the primitive isoprenoid organism is discussed. The role of magnetotactic actinidic archaea in the evolution of universe is elucidated. The role of actinidic archaea in global warming and evolutionary cycles is also discussed.
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