scholarly journals Differential modulation of the molecular dynamics of the type IIa and IIc sodium phosphate cotransporters by parathyroid hormone

2011 ◽  
Vol 301 (4) ◽  
pp. C850-C861 ◽  
Author(s):  
Luca Lanzano ◽  
Tim Lei ◽  
Kayo Okamura ◽  
Hector Giral ◽  
Yupanqui Caldas ◽  
...  
Keyword(s):  
Parathyroid Hormone ◽  
Proximal Tubule ◽  
Sodium Phosphate ◽  
Time Course ◽  
Apical Membrane ◽  
Response Times ◽  
Complete Removal ◽  
Myosin Vi ◽  
The Difference ◽  
Renal Proximal Tubule Cells

The kidney is a key regulator of phosphate homeostasis. There are two predominant renal sodium phosphate cotransporters, NaPi2a and NaPi2c. Both are regulated by parathyroid hormone (PTH), which decreases the abundance of the NaPi cotransporters in the apical membrane of renal proximal tubule cells. The time course of PTH-induced removal of the two cotransporters from the apical membrane, however, is markedly different for NaPi2a compared with NaPi2c. In animals and in cell culture, PTH treatment results in almost complete removal of NaPi2a from the brush border (BB) within 1 h whereas for NaPi2c this process in not complete until 4 to 8 h after PTH treatment. The reason for this is poorly understood. We have previously shown that the unconventional myosin motor myosin VI is required for PTH-induced removal of NaPi2a from the proximal tubule BB. Here we demonstrate that myosin VI is also necessary for PTH-induced removal of NaPi2c from the apical membrane. In addition, we show that, while at baseline the two cotransporters have similar diffusion coefficients within the membrane, after PTH addition the diffusion coefficient for NaPi2a initially exceeds that for NaPi2c. Thus NaPi2c appears to remain “tethered” in the apical membrane for longer periods of time after PTH treatment, accounting, at least in part, for the difference in response times to PTH of NaPi2a versus NaPi2c.

Fluoride mobilizes intracellular calcium and promotes Ca2+ influx in rat proximal tubules

1991 ◽  
Vol 261 (2) ◽  
pp. F318-F327 ◽  
Author(s):  
J. H. Dominguez ◽  
J. G. Garcia ◽  
J. K. Rothrock ◽  
D. English ◽  
C. Mann
Keyword(s):  
Signal Transduction ◽  
Parathyroid Hormone ◽  
Proximal Tubule ◽  
G Protein ◽  
Pertussis Toxin ◽  
Time Course ◽  
Proximal Tubules ◽  
Signal Transduction Pathways ◽  
Pi Hydrolysis

In the renal proximal tubule, external Ca2+ ([Ca2+]o) is required for parathyroid hormone to elevate cytosolic Ca2+ ([Ca2+]i). However, other hormones increase [Ca2+]i in the absence of [Ca2+]o. These differences may arise from a diversity of signal transduction pathways acting on external and internal Ca2+ pools. However, Ca2+ influx may be necessary to expedite and maintain the rise of [Ca2+]i for a period after the initial surge. In this study, F- was used to probe the roles of intracellular Ca2+ mobilization, Ca2+ influx, and phosphoinositide (PI) hydrolysis on the surge of [Ca2+]i in rat proximal tubules. In the presence of external Ca2+; 1-20 mM F- evoked incremental rises of [Ca2+]i in tubules loaded with aequorin. Whereas 10 mM F- increased [Ca2+]i in the absence of [Ca2+]o, the time constant for the [Ca2+]i surge was increased. These findings are consistent with a role of Ca2+ influx on the effect of F- on [Ca2+]i. Indeed, 10 mM F- also enhanced the uptake of 45Ca2+, and promoted Ca2+ influx in aequorin- and fura-2-loaded, Ca(2+)-deprived tubules. In tubules, F- also activated PI hydrolysis with a time course that paralleled Ca2+ mobilization. The effect of F- on [Ca2+]i was not altered when the 39-kDa pertussis toxin substrate was inactivated with the toxin. This G protein was most likely Gi, because prostaglandin E2, an activator of Gi in tubules, dissociated the pertussis toxin-sensitive protein. The results support the notion that activation of a signal-transduction complex, the F- substrate, causes Ca2+ influx, mobilizes internal Ca2+, and activates PI hydrolysis in rat proximal tubules.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals PTH transiently increases the percent mobile fraction of Npt2a in OK cells as determined by FRAP

2009 ◽  
Vol 297 (6) ◽  
pp. F1560-F1565 ◽  
Author(s):  
Edward J. Weinman ◽  
Deborah Steplock ◽  
Boyoung Cha ◽  
Olga Kovbasnjuk ◽  
Nicholas A. Frost ◽  
...  
Keyword(s):  
Proximal Tubule ◽  
Binding Proteins ◽  
Apical Membrane ◽  
Wild Type ◽  
Lateral Mobility ◽  
Proximal Tubule Cells ◽  
Ok Cells ◽  
Mobile Fraction ◽  
Tubule Cells ◽  
Renal Proximal Tubule Cells

Renal sodium-dependent phosphate transporter 2a (Npt2a) binds to a number of PDZ adaptor proteins including sodium-hydrogen exchanger regulatory factor-1 (NHERF-1), which regulates its retention in the apical membrane of renal proximal tubule cells and the response to parathyroid hormone (PTH). The present experiments were designed to study the lateral mobility of enhanced green fluorescent protein (EGFP)-Npt2a in proximal tubule-like opossum kidney (OK) cells using fluorescence recovery after photobleaching (FRAP) and to determine the role of PDZ binding proteins in mediating the effects of PTH. The mobile fraction of wild-type Npt2a (EGFP-Npt2a-TRL) under basal conditions was ∼17%. Treatment of the cells with Bis(sulfosuccinimidyl) suberate, a water-soluble cross-linker, abolished recovery nearly completely, indicating that recovery represented lateral diffusion in the plasma membrane and not the exocytosis or synthesis of unbleached transporter. Substitution of the C-terminal amino acid PDZ binding sequence TRL with AAA (EGFP-Npt2a-AAA) resulted in a nearly twofold increase in percent mobile fraction of Npt2a. Treatment of cells with PTH resulted in a rapid increase in the percent mobile fraction to >30% followed by a time-dependent decrease to baseline or below. PTH had no effect on the mobility of EGFP-Npt2a-AAA expressed in native OK cells or on wild-type EGFP-Npt2a-TRL expressed in OK-H cells deficient in NHERF-1. These findings indicate that the association of Npt2a with PDZ binding proteins limits the lateral mobility of the transporter in the apical membrane of renal proximal tubule cells. Treatment with PTH, presumably by dissociating NHERF-1/Npt2a complexes, transiently increases the mobility of Npt2a, suggesting that freeing of Npt2a from the cytoskeleton precedes PTH-mediated endocytosis.

Posttranscriptional regulation of the proximal tubule NaPi-II transporter in response to PTH and dietary Pi

1999 ◽  
Vol 277 (5) ◽  
pp. F676-F684 ◽  
Author(s):  
Heini Murer ◽  
Ian Forster ◽  
Nati Hernando ◽  
Georg Lambert ◽  
Martin Traebert ◽  
...  
Keyword(s):  
Parathyroid Hormone ◽  
Proximal Tubule ◽  
Posttranscriptional Regulation ◽  
Apical Membrane ◽  
Tubular Reabsorption ◽  
Transport Capacity ◽  
Phosphate Intake ◽  
Proximal Tubular ◽  
Dietary Phosphate ◽  
Proximal Tubular Reabsorption

The rate of proximal tubular reabsorption of phosphate (Pi) is a major determinant of Pi homeostasis. Deviations of the extracellular concentration of Piare corrected by many factors that control the activity of Na-Pi cotransport across the apical membrane. In this review, we describe the regulation of proximal tubule Pi reabsorption via one particular Na-Pi cotransporter (the type IIa cotransporter) by parathyroid hormone (PTH) and dietary phosphate intake. Available data indicate that both factors determine the net amount of type IIa protein residing in the apical membrane. The resulting change in transport capacity is a function of both the rate of cotransporter insertion and internalization. The latter process is most likely regulated by PTH and dietary Pi and is considered irreversible since internalized type IIa Na-Picotransporters are subsequently routed to the lysosomes for degradation.

scholarly journals Defective Parathyroid Hormone Regulation of NHE3 Activity and Phosphate Adaptation in Cultured NHERF-1-/-Renal Proximal Tubule Cells

2004 ◽  
Vol 279 (36) ◽  
pp. 37815-37821 ◽  
Author(s):  
Rochelle Cunningham ◽  
Deborah Steplock ◽  
Fengying Wang ◽  
Huijun Huang ◽  
Xiaofei E ◽  
...  
Keyword(s):  
Parathyroid Hormone ◽  
Proximal Tubule ◽  
Renal Proximal Tubule ◽  
Hormone Regulation ◽  
Proximal Tubule Cells ◽  
Tubule Cells ◽  
Renal Proximal Tubule Cells

scholarly journals Porcine myosin-VI: characterization of a new mammalian unconventional myosin.

1994 ◽  
Vol 127 (2) ◽  
pp. 425-440 ◽  
Author(s):  
T Hasson ◽  
M S Mooseker
Keyword(s):  
Proximal Tubule ◽  
Cell Line ◽  
Actin Binding ◽  
Proximal Tubule Cell ◽  
Myosin Vi ◽  
Proximal Tubule Cells ◽  
Unconventional Myosin ◽  
Tubule Cells ◽  
The Difference

We have cloned a new mammalian unconventional myosin, porcine myosin-VI from the proximal tubule cell line, LLC-PK1 (CL4). Porcine myosin-VI is highly homologous to Drosophila 95F myosin heavy chain, and together these two myosins comprise a sixth class of myosin motors. Myosin-VI exhibits ATP-sensitive actin-binding activities characteristic of myosins, and it is associated with a calmodulin light chain. Within LLC-PK1 cells, myosin-VI is soluble and does not associate with the major actin-containing domains. Within the kidney, however, myosin-VI is associated with sedimentable structures and specifically locates to the actin- and membrane-rich apical brush border domain of the proximal tubule cells. This motor was not enriched within the glomerulus, capillaries, or distal tubules. Myosin-VI associates with the proximal tubule cytoskeleton in an ATP-sensitive fashion, suggesting that this motor is associated with the actin cytoskeleton within the proximal tubule cells. Given the difference in association of myosin-VI with the apical cytoskeleton between LLC-PK1 cells and adult kidney, it is likely that this cell line does not fully differentiate to form functional proximal tubule cells. Myosin-VI may require the presence of additional elements, only found in vivo in proximal tubule cells, to properly locate to the apical domain.

Parathyroid hormone and dibutyryl cAMP inhibit Na+/H+ exchange in renal brush border vesicles

1985 ◽  
Vol 248 (2) ◽  
pp. F212-F218 ◽  
Author(s):  
A. M. Kahn ◽  
G. M. Dolson ◽  
M. K. Hise ◽  
S. C. Bennett ◽  
E. J. Weinman
Keyword(s):  
Parathyroid Hormone ◽  
Proximal Tubule ◽  
Brush Border ◽  
Sodium Phosphate ◽  
Brush Border Membrane ◽  
Gradient Centrifugation ◽  
Membrane Vesicles ◽  
Enzymatic Digestion ◽  
Dibutyryl Camp ◽  
Ficoll Gradient

Parathyroid hormone (PTH) and cAMP inhibit sodium, water, and bicarbonate reabsorption in the proximal tubule. We wished to determine whether these agents directly inhibit proximal tubular Na+/H+ exchange. A suspension of rabbit proximal tubules was prepared by enzymatic digestion and Ficoll gradient centrifugation. Oxygen consumption at 37 degrees C was stable over 60 min, averaged 20 nmol X mg protein-1 X min-1, and was inhibited 60% by ouabain. Over 96% of cells excluded trypan blue. From this suspension, brush border membrane vesicles were isolated. The vesicles were enriched 12.7 times in alkaline phosphatase relative to a cortical homogenate and demonstrated pH gradient-stimulated, amiloride-sensitive Na+/H+ countertransport and sodium-phosphate and sodium-D-glucose cotransport. When the tubule suspension was exposed to PTH or dibutyryl cAMP, the activity of Na+/H+ countertransport in the resultant brush border vesicles was inhibited. Neither PTH nor dibutyryl cAMP affected the amiloride-insensitive component of sodium transport or sodium-phosphate or sodium-D-glucose cotransport. The effect of PTH on Na+/H+ counter-transport could not be explained by an alteration in fluidity of the brush border membrane. These experiments demonstrate that PTH and dibutyryl cAMP directly inhibit Na+/H+ countertransport in the brush border membrane of the rabbit proximal tubule.

scholarly journals Functional colocalization of water channels and proton pumps in endosomes from kidney proximal tubule.

1989 ◽  
Vol 93 (5) ◽  
pp. 885-902 ◽  
Author(s):  
R G Ye ◽  
L B Shi ◽  
W I Lencer ◽  
A S Verkman
Keyword(s):  
Proximal Tubule ◽  
Time Course ◽  
Apical Membrane ◽  
Water Channels ◽  
Fluorescence Signal ◽  
Proton Pumps ◽  
Osmotic Gradient ◽  
Proton Atpase ◽  
Osmotic Water

The apical membrane of mammalian proximal tubule undergoes rapid membrane cycling by exocytosis and endocytosis. Osmotic water and ATP-driven proton transport were measured in endocytic vesicles from rabbit and rat proximal tubule apical membrane labeled in vivo with the fluid phase marker fluorescein-dextran. Osmotic water permeability (Pf) was determined from the time course of fluorescein-dextran fluorescence after exposure of endosomes to an inward osmotic gradient in a stopped-flow apparatus. Pf was 0.009 (rabbit) and 0.029 cm/s (rat) (23 degrees C) and independent of osmotic gradient size. Pf in rabbit endosomes was inhibited reversibly by HgCl2 (KI = 0.2 mM) and had an activation energy of 6.4 +/- 0.5 kcal/mol (15-35 degrees C). Endosomal proton ATPase activity was measured from the time course of internal pH, measured by fluorescein-dextran fluorescence, after the addition of external ATP. Endosomes contained an ATP-driven proton pump that was sensitive to N-ethylmaleimide and insensitive to vanadate and oligomycin. In response to saturating [ATP] the pump acidified the endosomal compartment at a rate of 0.17 (rat) and 0.029 pH unit/s (rabbit); at an external pH of 7.4, the steady-state pH was 6.4 (rat) and 6.5 (rabbit). To examine whether water channels and the proton ATPase were present in the same endosome, the time course of fluorescein-dextran fluorescence was measured in response to an osmotic gradient in the presence and absence of ATP. ATP did not alter endosome Pf, but decreased the amplitude of the fluorescence signal by 43 +/- 3% (rabbit) and 47 +/- 4% (rat).(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Shank2 redistributes with NaPilla during regulated endocytosis

2010 ◽  
Vol 299 (6) ◽  
pp. C1324-C1334 ◽  
Author(s):  
Evgenia Dobrinskikh ◽  
Hector Giral ◽  
Yupanqui A. Caldas ◽  
Moshe Levi ◽  
R. Brian Doctor
Keyword(s):  
Proximal Tubule ◽  
Time Course ◽  
Pdz Domain ◽  
Renal Proximal Tubule ◽  
Proximal Tubule Cells ◽  
Early Endosomes ◽  
Tubule Cells ◽  
Renal Proximal Tubule Cells ◽  
High Phosphate

Serum phosphate levels are acutely impacted by the abundance of sodium-phosphate cotransporter IIa (NaPiIIa) in the apical membrane of renal proximal tubule cells. PSD-95/Disks Large/Zonula Occludens (PDZ) domain-containing proteins bind NaPiIIa and likely contribute to the delivery, retention, recovery, and trafficking of NaPiIIa. Shank2 is a distinctive PDZ domain protein that binds NaPiIIa. Its role in regulating NaPiIIa activity, distribution, and abundance is unknown. In the present in vivo study, rats were maintained on a low-phosphate diet, and then plasma phosphate levels were acutely elevated by high-phosphate feeding to induce the recovery, endocytosis, and degradation of NaPiIIa. Western blot analysis of renal cortical tissue from rats given high-phosphate feed showed NaPiIIa and Shank2 underwent degradation. Quantitative immunofluorescence analyses, including microvillar versus intracellular intensity ratios and intensity correlation quotients, showed that Shank2 redistributed with NaPiIIa during the time course of NaPiIIa endocytosis. Furthermore, NaPiIIa and Shank2 trafficked through distinct endosomal compartments (clathrin, early endosomes, lysosomes) with the same temporal pattern. These in vivo findings indicate that Shank2 is positioned to coordinate the regulated endocytic retrieval and downregulation of NaPiIIa in rat renal proximal tubule cells.

Sign in / Sign up

Export Citation Format

Share Document