scholarly journals Stimulation of fructose transport across the intestinal brush-border membrane by PMA is mediated by GLUT2 and dynamically regulated by protein kinase C

2000 ◽  
Vol 350 (1) ◽  
pp. 149 ◽  
Author(s):  
Philip A. HELLIWELL ◽  
Michael RICHARDSON ◽  
Julie AFFLECK ◽  
George L. KELLETT
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Intestinal Brush Border Membrane ◽  
Kinase C ◽  
Intestinal Brush Border ◽  
Stimulation Of

Phorbol ester inhibition of chicken intestinal brush-border sodium-proton exchange

1991 ◽  
Vol 260 (6) ◽  
pp. C1264-C1272 ◽  
Author(s):  
E. B. Chang ◽  
M. W. Musch ◽  
D. Drabik-Arvans ◽  
M. C. Rao
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phorbol Ester ◽  
Brush Border ◽  
Time Course ◽  
Brush Border Membrane ◽  
Phorbol Esters ◽  
Kinase C ◽  
Na Uptake ◽  
Exchange Activity

Phorbol esters, specific activators of protein kinase C, inhibit amiloride-sensitive Na uptake from the mucosal medium in intact intestinal mucosa as well as in isolated chicken villus enterocytes. In isolated cells, maximal inhibition is observed at 60 s, and influx returns to control values within 15 min. This effect can be measured either as initial 22Na influx rates or by following changes in intracellular pH using the pH-sensitive fluorescent dye 5,6-carboxyfluorescein. The effects of amiloride and phorbol esters were not additive, suggesting inhibition of a common transport system, i.e., Na-H exchange. In brush-border membrane vesicles (BBMV) made from villus enterocytes, amiloride-sensitive Na-H exchange activity was significantly inhibited in phorbol ester-treated cells. The degree of inhibition of 22Na uptake by BBMV had the same time course and dose-effect relationship as phorbol ester-inhibited cellular Na uptake. Similarly, the time course of protein kinase C translocation from cytosol to particulate or brush-border membrane fractions correlated with Na uptake measurements made in whole cells and BBMV. These results suggest that protein kinase C activation in chicken villus enterocytes inhibits brush-border membrane Na-H exchange activity.

Protein kinase C in mouse kidney: subcellular distribution and endogenous substrates

1988 ◽  
Vol 66 (4) ◽  
pp. 262-272 ◽  
Author(s):  
Avihu Boneh ◽  
Harriet S. Tenenhouse
Keyword(s):  
Protein Kinase C ◽  
Catalytic Activity ◽  
Protein Kinase ◽  
Brush Border ◽  
Subcellular Distribution ◽  
Brush Border Membrane ◽  
Mouse Kidney ◽  
Kidney Cortex ◽  
Kinase C ◽  
Endogenous Substrates

The subcellular distribution, kinetic properties, and endogenous substrates of calcium-activated, phospholipid-dependent protein kinase (protein kinase C) were examined in mouse kidney cortex. Protein kinase C associated with the particulate, mitochondrial, and brush border membrane fractions was assayed after solubilization in 0.2% Triton X-100 under conditions shown to be noninhibitory to catalytic activity. Of recovered activity, 52% was associated with the cytosolic fraction; mitochondrial and brush border membrane associated protein kinase C constituted 12 and 3%, respectively, of the activity recovered in the particulate fraction. Protein kinase C associated with brush border membranes exhibited a high affinity for ATP (apparent Km = 62 ± 10 μM) and the highest apparent maximal velocity (1146 ± 116 pmol P/(mg protein∙min)) of the renal fractions examined. Maximal stimulation of protein kinase C by diacylglycerol (in the presence of phosphatidylserine) was achieved at both 25 and 300 μM calcium in all renal fractions. These results are consistent with previous reports demonstrating that diacylglycerol increases the apparent affinity of protein kinase C for calcium. Phorbol 12-myristate 13-acetate, but not 4α-phorbol, was able to substitute for diacylglycerol and stimulate cytosolic and particulate renal protein kinase C. 1-(5-Isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride, a specific inhibitor of protein kinase C, led to significant inhibition of catalytic activity in all renal subcellular fractions. Endogenous substrates for protein kinase C were demonstrated in renal cytosolic (26, 45, 63, and 105 kilodaltons (kDa)), particulate (26, 33, 68, and 105 kDa), mitochondrial (43 kDa), and brush border membrane (26, 41, 52, 88, and 105 kDa) fractions. The possible physiological significance of protein kinase C in mammalian kidney is discussed.

scholarly journals Stimulation of fructose transport across the intestinal brush-border membrane by PMA is mediated by GLUT2 and dynamically regulated by protein kinase C

2000 ◽  
Vol 350 (1) ◽  
pp. 149-154 ◽  
Author(s):  
Philip A. HELLIWELL ◽  
Michael RICHARDSON ◽  
Julie AFFLECK ◽  
George L. KELLETT
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Basolateral Membrane ◽  
Fold Increase ◽  
Sugar Absorption ◽  
Kinase C

Perfusion of rat jejunum in vitro with PMA increased fructose transport by 70% compared with control values and was blocked by the protein kinase C (PKC) inhibitor chelerythrine. The brush-border membrane contained both the fructose transporters GLUT5 and GLUT2; the presence of the latter was confirmed by luminal biotinylation. PMA increased the GLUT2 level 4-fold within minutes, so that the level was comparable with that of the basolateral membrane, but had no effect on GLUT5 level. GLUT2 was functional, accessible to luminal fructose and could be inhibited selectively by phloretin to permit determination of GLUT2- and GLUT5-mediated transport components. The 4-fold increase in GLUT2 level induced by PMA was matched by a 4-fold increase in GLUT2-mediated transport: there was a compensatory fall in the GLUT5-mediated rate. The pattern of dynamic trafficking was seen only for GLUT2, not GLUT5 or SGLT1, implying that GLUT2 trafficks to the brush-border membrane by a different pathway. Trafficking of GLUT2 to the brush-border membrane correlated with activation of PKC βII, implying that this isoenzyme is likely to control trafficking. Since PKC is activated by endogenous hormones, GLUT2 levels in vivo are 3–4-fold those in vitro; moreover, because PKC is inactivated as soon as intestine is excised, GLUT2 is lost from the brush-border within minutes in vitro. It is therefore difficult to detect GLUT2 in most in vitro preparations and its role in intestinal sugar absorption across the brush-border membrane has accordingly been overlooked.

Divalent cations and vitamin B12 uptake by intestinal brush-border membrane vesicles

1980 ◽  
Vol 239 (6) ◽  
pp. G452-G456
Author(s):  
R. C. Beesley ◽  
C. D. Bacheller
Keyword(s):  
Vitamin B12 ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Divalent Cations ◽  
Intrinsic Factor ◽  
Membrane Vesicles ◽  
Brush Border Membrane Vesicles ◽  
Tetraacetic Acid ◽  
Intestinal Brush Border Membrane ◽  
Intestinal Brush Border

Brush-border membrane vesicles from hamster intestine were employed to investigate uptake (binding) of vitamin B12 (B12). Ileal vesicles took up 25 times more B12 than did jejunal vesicles. Uptake of B12 by ileal vesicles was dependent on intrinsic factor (IF) and required Ca2+. Increasing the Ca2+ concentration caused an increase in uptake of B12 reaching a maximum at approximately 8 mM Ca2+. At high Ca2+ concentrations, 6–8 mM, Mg2+ had little effect on uptake of B12. At low Ca2+ concentrations, up to 2 mM, Mg2+ stimulated B12 uptake. Mg2+, Mn2+, and, to a lesser extent, Sr2+ stimulated Ca2+-dependent B12 uptake, but Zn2+, Ba2+, Na+, K+, and La3+ did not. B12 was apparently not metabolized and was bound as IF-B12 complex, which could be removed with (ethylenedinitrilo)tetraacetic acid (EDTA). Our results suggest that two types of divalent cation reactive sites are involved in binding of IF-B12. One is Ca2+ specific. The other is less specific reacting with Mg2+, Mn2+, Sr2+, and perhaps Ca2+ itself, thereby stimulating Ca2+-dependent binding of IF-B12 to its ileal receptor.

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