scholarly journals Rapid Regulation of Human Multidrug and Extrusion Transporters hMATE1 and hMATE2K

2020 ◽  
Vol 21 (14) ◽  
pp. 5157
Author(s):  
Marta Kantauskaitė ◽  
Anna Hucke ◽  
Moritz Reike ◽  
Sara Ahmed Eltayeb ◽  
Chuyan Xiao ◽  
...  
Keyword(s):  
Plasma Membranes ◽  
Membrane Domains ◽  
Renal Secretion ◽  
Madin Darby Canine Kidney ◽  
Fluorescent Substrate ◽  
Transcellular Transport ◽  
Human Embryonic Kidney Cells ◽  
Specific Regulation ◽  
Vectorial Transport ◽  
Darby Canine Kidney

Vectorial transport of organic cations (OCs) in renal proximal tubules is mediated by sequential action of human OC transporter 2 (hOCT2) and human multidrug and toxic extrusion protein 1 and 2K (hMATE1 and hMATE2K), expressed in the basolateral (hOCT2) and luminal (hMATE1 and hMATE2K) plasma membranes, respectively. It is well known that hOCT2 activity is subjected to rapid regulation by several signaling pathways, suggesting that renal OC secretion may be acutely adapted to physiological requirements. Therefore, in this work, the acute regulation of hMATEs stably expressed in human embryonic kidney cells was characterized using the fluorescent substrate 4-(4-(dimethylamino)styryl)-N-methylpyridinium (ASP+) as a marker. A specific regulation of ASP+ transport by hMATE1 and hMATE2K measured in uptake and efflux configurations was observed. In the example of hMATE1 efflux reduction by inhibition of casein kinase II, it was also shown that this regulation is able to modify transcellular transport of ASP+ in Madin–Darby canine kidney II cells expressing hOCT2 and hMATE1 on the basolateral and apical membrane domains, respectively. The activity of hMATEs can be rapidly regulated by some intracellular pathways, which sometimes are common to those found for hOCTs. Interference with these pathways may be important to regulate renal secretion of OCs.

Vectorial transport of bile salts across MDCK cells expressing both rat Na+-taurocholate cotransporting polypeptide and rat bile salt export pump

2005 ◽  
Vol 288 (1) ◽  
pp. G159-G167 ◽  
Author(s):  
Sachiko Mita ◽  
Hiroshi Suzuki ◽  
Hidetaka Akita ◽  
Bruno Stieger ◽  
Peter J. Meier ◽  
...  
Keyword(s):  
Bile Salt ◽  
Bile Salts ◽  
Mdck Cells ◽  
Bile Salt Export Pump ◽  
Madin Darby Canine Kidney ◽  
Transcellular Transport ◽  
Physiological Conditions ◽  
Vectorial Transport ◽  
Rat Bile ◽  
Darby Canine Kidney

Bile salts are predominantly taken up by hepatocytes via the basolateral Na+-taurocholate cotransporting polypeptide (NTCP/SLC10A1) and secreted into the bile by the bile salt export pump (BSEP/ABCB11). In the present study, we transfected rat Ntcp and rat Bsep into polarized Madin-Darby canine kidney cells and characterized the transport properties of these cells for eight bile salts. Immunohistochemical staining demonstrated that Ntcp was expressed at the basolateral domains, whereas Bsep was expressed at the apical domains. Basal-to-apical transport of taurocholate across the monolayer expressing only Ntcp and that coexpressing Ntcp/Bsep was observed, whereas the flux across the monolayer of control and Bsep-expressing cells was symmetrical. Basal-to-apical transport of taurocholate across Ntcp/Bsep-coexpressing monolayers was significantly higher than that across monolayers expressing only Ntcp. Kinetic analysis of this vectorial transport of taurocholate gave an apparent Km value of 13.9 ± 4.7 μM for cells expressing Ntcp alone, which is comparable with 22.2 ± 4.5 μM for cells expressing both Ntcp and Bsep and Vmax values of 15.8 ± 4.2 and 60.8 ± 9.0 pmol·min−1·mg protein−1 for Ntcp alone and Ntcp and Bsep-coexpressing cells, respectively. Transcellular transport of cholate, glycocholate, taurochenodeoxycholate, chenodeoxycholate, glycochenodeoxycholate, tauroursodeoxycholate, ursodeoxycholate, and glycoursodeoxycholate, but not that of lithocholate was also observed across the double transfectant. This double-expressing system can be used as a model to clarify vectorial transport of bile salts across hepatocytes under physiological conditions.

Localization and topology of a urate transporter/channel, a galectin, in epithelium-derived cells

2001 ◽  
Vol 281 (6) ◽  
pp. C1926-C1939 ◽  
Author(s):  
Joshua Z. Rappoport ◽  
Michael S. Lipkowitz ◽  
Ruth G. Abramson
Keyword(s):  
Lipid Bilayers ◽  
Plasma Membranes ◽  
Kidney Cells ◽  
Madin Darby Canine Kidney ◽  
Urate Transporter ◽  
And Topology ◽  
Cell Surface Biotinylation ◽  
Cytoplasmic Side ◽  
Darby Canine Kidney ◽  
Canine Kidney

Recombinant protein produced from a cDNA cloned in our laboratory (UAT) functions in lipid bilayers as a urate transporter/channel. Because UAT is a galectin, a family of proteins presumed to be soluble, the localization and topology of UAT were assessed in living cells. UAT was targeted to plasma membrane in multiple epithelium-derived cell lines and, in polarized cells, was targeted to both apical and basolateral membranes. The amino and carboxy termini of UAT were both detected on the cytoplasmic side of plasma membranes, whereas cell surface biotinylation studies demonstrated that UAT is not merely a cytosolic membrane-associated protein but contains at least one extracellular domain. Madin-Darby canine kidney cells were shown both functionally and immunologically to contain an apparent homolog of UAT; however, transfection with UAT did not modify urate uptake. Because coimmunoprecipitation studies revealed that UAT is capable of forming both homo- and heteromultimers, it is proposed that monomers of endogenous channels are in part replaced by monomers of the protein expressed subsequent to transfection, thereby maintaining constancy of urate uptake at basal levels.

scholarly journals Quantitative analysis of the lipidomes of the influenza virus envelope and MDCK cell apical membrane

2012 ◽  
Vol 196 (2) ◽  
pp. 213-221 ◽  
Author(s):  
Mathias J. Gerl ◽  
Julio L. Sampaio ◽  
Severino Urban ◽  
Lucie Kalvodova ◽  
Jean-Marc Verbavatz ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Plasma Membranes ◽  
Apical Membrane ◽  
Membrane Raft ◽  
Madin Darby Canine Kidney ◽  
Virus Envelope ◽  
Storage Lipids ◽  
Cell Plasma ◽  
And Storage ◽  
Darby Canine Kidney

The influenza virus (IFV) acquires its envelope by budding from host cell plasma membranes. Using quantitative shotgun mass spectrometry, we determined the lipidomes of the host Madin–Darby canine kidney cell, its apical membrane, and the IFV budding from it. We found the apical membrane to be enriched in sphingolipids (SPs) and cholesterol, whereas glycerophospholipids were reduced, and storage lipids were depleted compared with the whole-cell membranes. The virus membrane exhibited a further enrichment of SPs and cholesterol compared with the donor membrane at the expense of phosphatidylcholines. Our data are consistent with and extend existing models of membrane raft-based biogenesis of the apical membrane and IFV envelope.

scholarly journals Transcellular Transport of Leptin by the Short Leptin Receptor Isoform ObRa in Madin-Darby Canine Kidney Cells*

Endocrinology ◽  
2000 ◽  
Vol 141 (6) ◽  
pp. 1955-1961 ◽  
Author(s):  
Stanley M. Hileman ◽  
Jens Tornøe ◽  
Jeffrey S. Flier ◽  
Christian Bjørbæk
Keyword(s):  
Cell Surface ◽  
Leptin Receptor ◽  
Mdck Cells ◽  
Short Form ◽  
Madin Darby Canine Kidney ◽  
Apical Cell Membrane ◽  
Transcellular Transport ◽  
The Brain ◽  
Darby Canine Kidney ◽  
Canine Kidney

Abstract Leptin is an adipocyte-derived hormone that acts in specific regions of the brain to regulate body weight and neuroendocrine function. The mechanism by which leptin enters the brain is unknown. We previously reported that rat brain microvessels, which constitute the blood-brain barrier, contain large amounts of messenger RNA encoding a short form of the leptin receptor (ObRa), suggesting that this site may be important for receptor-mediated transport of leptin into the brain. The purpose of this study was to determine whether ObRa is capable of transcellular transport of intact leptin. A transwell system in which Madin-Darby Canine Kidney (MDCK) cells stably expressing ObRa are grown in a monolayer was used to determine receptor distribution on apical or basolateral cell surfaces and the capacity for directional transport of 125I-leptin. Binding of 125I-leptin was greater on the apical vs. the basolateral cell surface and transport of 125I-leptin occurred only in the apical to basolateral direction. 11% of transported radioactivity appearing in the basolateral chamber represented intact leptin as assessed by TCA precipitation analysis and by SDS-PAGE. Parental MDCK cells did not express leptin receptors and did not bind or transport 125I-leptin. Epidermal growth factor (EGF) binding and transport via endogenous EGF receptors in MDCK cells also was assessed. In contrast to leptin, specific binding of 125I-EGF occurred primarily on the basolateral cell surface and transport of 125I-EGF occurred predominantly in the basolateral to apical direction. These data show that ObRa is preferentially targeted to the apical cell membrane in MDCK cells and that leptin transport occurs, albeit at a low rate, in a unidirectional manner in the apical to basolateral direction. These findings may be relevant to the putative role of ObRa in receptor-mediated transport of leptin from the circulation into the brain.

scholarly journals Polarized delivery of viral glycoproteins to the apical and basolateral plasma membranes of Madin-Darby canine kidney cells infected with temperature-sensitive viruses.

1985 ◽  
Vol 100 (1) ◽  
pp. 136-151 ◽  
Author(s):  
M J Rindler ◽  
I E Ivanov ◽  
H Plesken ◽  
D D Sabatini
Keyword(s):  
Plasma Membrane ◽  
G Protein ◽  
Plasma Membranes ◽  
Temperature Sensitive ◽  
Kidney Cells ◽  
Apical Surface ◽  
Nonpermissive Temperature ◽  
Madin Darby Canine Kidney ◽  
Darby Canine Kidney ◽  
Canine Kidney

The intracellular route followed by viral envelope glycoproteins in polarized Madin-Darby canine kidney cells was studied by using temperature-sensitive mutants of vesicular stomatitis virus (VSV) and influenza, in which, at the nonpermissive temperature (39.5 degrees C), the newly synthesized glycoproteins (G proteins) and hemagglutinin (HA), respectively, are not transported out of the endoplasmic reticulum. After infection with VSV and incubation at 39.5 degrees C for 4-5 h, synchronous transfer of G protein to the plasma membrane was initiated by shifting to the permissive temperature (32.5 degrees C). Immunoelectron microscopy showed that under these conditions the protein moved to the Golgi apparatus and from there directly to a region of the lateral plasma membrane near this organelle. G protein then seemed to diffuse progressively to basal regions of the cell surface and, only after it had accumulated in the basolateral domain, it began to appear on the apical surface near the intercellular junctions. The results of these experiments indicate that the VSV G protein must be sorted before its arrival at the cell surface, and suggest that passage to the apical domain occurs only late in infection when tight junctions are no longer an effective barrier. In complementary experiments, using the temperature-sensitive mutant of influenza, cultures were first shifted from the nonpermissive temperature (39.5 degrees C) to 18.5 degrees C, to allow entrance of the glycoprotein into the Golgi apparatus (see Matlin, K.S., and K. Simons, 1983, Cell, 34:233-243). Under these conditions HA accumulated in Golgi stacks and vesicles but did not reach the plasma membrane. When the temperature was subsequently shifted to 32.5 degrees C, HA rapidly appeared in discrete regions of the apical surface near, and often directly above, the Golgi elements, and later diffused throughout this surface. To ensure that the anti-HA antibodies had access to lateral domains, monolayers were treated with a hypertonic medium to dilate the intercellular spaces. Some labeling was then observed in the lateral plasma membranes soon after the shift, but this never increased beyond 1.0 gold particle/micron, whereas characteristic densities of labeling in apical surfaces soon became much higher (approximately 10 particles/micron). Our results suggest that the bulk of HA follows a direct pathway leading from the Golgi to regions of the apical surface close to trans-Golgi cisternae.

scholarly journals Selective Alterations in Biosynthetic and Endocytic Protein Traffic in Madin-Darby Canine Kidney Epithelial Cells Expressing Mutants of the Small GTPase Rac1

2000 ◽  
Vol 11 (1) ◽  
pp. 287-304 ◽  
Author(s):  
Tzuu-Shuh Jou ◽  
Som-Ming Leung ◽  
Linette M. Fung ◽  
Wily G. Ruiz ◽  
W. James Nelson ◽  
...  
Keyword(s):  
Apical Membrane ◽  
Mdck Cells ◽  
Basolateral Membrane ◽  
Membrane Traffic ◽  
Small Gtpase ◽  
Apical Plasma Membrane ◽  
Membrane Domains ◽  
Madin Darby Canine Kidney ◽  
Darby Canine Kidney ◽  
Canine Kidney

Madin-Darby canine kidney (MDCK) cells expressing constitutively active Rac1 (Rac1V12) accumulate a large central aggregate of membranes beneath the apical membrane that contains filamentous actin, Rac1V12, rab11, and the resident apical membrane protein GP-135. To examine the roles of Rac1 in membrane traffic and the formation of this aggregate, we analyzed endocytic and biosynthetic trafficking pathways in MDCK cells expressing Rac1V12 and dominant inactive Rac1 (Rac1N17). Rac1V12 expression decreased the rates of apical and basolateral endocytosis, whereas Rac1N17 expression increased those rates from both membrane domains. Basolateral-to-apical transcytosis of immunoglobulin A (IgA) (a ligand for the polymeric immunoglobulin receptor [pIgR]), apical recycling of pIgR-IgA, and accumulation of newly synthesized GP-135 at the apical plasma membrane were all decreased in cells expressing Rac1V12. These effects of Rac1V12 on trafficking pathways to the apical membrane were the result of the delivery and trapping of these proteins in the central aggregate. In contrast to abnormalities in apical trafficking events, basolateral recycling of transferrin, degradation of EGF internalized from the basolateral membrane, and delivery of newly synthesized pIgR from the Golgi to the basolateral membrane were all relatively unaffected by Rac1V12 expression. Rac1N17 expression had little or no effect on these postendocytic or biosynthetic trafficking pathways. These results show that in polarized MDCK cells activated Rac1 may regulate the rate of endocytosis from both membrane domains and that expression of dominant active Rac1V12 specifically alters postendocytic and biosynthetic membrane traffic directed to the apical, but not the basolateral, membrane.

scholarly journals Modulation of Endocytic Traffic in Polarized Madin-Darby Canine Kidney Cells by the Small GTPase RhoA

1999 ◽  
Vol 10 (12) ◽  
pp. 4369-4384 ◽  
Author(s):  
Som-Ming Leung ◽  
Raul Rojas ◽  
Christopher Maples ◽  
Christopher Flynn ◽  
Wily G. Ruiz ◽  
...  
Keyword(s):  
Mdck Cells ◽  
Immunoglobulin A ◽  
Small Gtpase ◽  
Membrane Domains ◽  
Madin Darby Canine Kidney ◽  
Immunoglobulin Receptor ◽  
Early Endosomes ◽  
Biochemical Analyses ◽  
Darby Canine Kidney ◽  
Canine Kidney

Efficient postendocytic membrane traffic in polarized epithelial cells is thought to be regulated in part by the actin cytoskeleton. RhoA modulates assemblies of actin in the cell, and it has been shown to regulate pinocytosis and phagocytosis; however, its effects on postendocytic traffic are largely unexplored. To this end, we expressed wild-type RhoA (RhoAWT), dominant active RhoA (RhoAV14), and dominant inactive RhoA (RhoAN19) in Madin-Darby canine kidney (MDCK) cells expressing the polymeric immunoglobulin receptor. RhoAV14 expression stimulated the rate of apical and basolateral endocytosis, whereas RhoAN19 expression decreased the rate from both membrane domains. Polarized basolateral recycling of transferrin was disrupted in RhoAV14-expressing cells as a result of increased ligand release at the apical pole of the cell. Degradation of basolaterally internalized epidermal growth factor was slowed in RhoAV14-expressing cells. Although apical recycling of immunoglobulin A (IgA) was largely unaffected in cells expressing RhoAV14, transcytosis of basolaterally internalized IgA was severely impaired. Morphological and biochemical analyses demonstrated that a large proportion of IgA internalized from the basolateral pole of RhoAV14-expressing cells remained within basolateral early endosomes and was slow to exit these compartments. RhoAN19 and RhoAWT expression had little effect on these postendocytic pathways. These results indicate that in polarized MDCK cells activated RhoA may modulate endocytosis from both membrane domains and postendocytic traffic at the basolateral pole of the cell.

scholarly journals Receptor-mediated vectorial transcytosis of epidermal growth factor by Madin-Darby canine kidney cells.

1987 ◽  
Vol 105 (4) ◽  
pp. 1595-1601 ◽  
Author(s):  
E Maratos-Flier ◽  
C Y Kao ◽  
E M Verdin ◽  
G L King
Keyword(s):  
Growth Factor ◽  
Epidermal Growth Factor ◽  
Egf Receptor ◽  
Mdck Cells ◽  
Egf Receptors ◽  
Madin Darby Canine Kidney ◽  
Transcellular Transport ◽  
Epidermal Growth ◽  
Darby Canine Kidney ◽  
Canine Kidney

Transcellular transport of a variety of ligands may be an important mechanism by which regulatory substances reach their site of action. We have studied the transcellular transport of two 6,000-mol-wt proteins, epidermal growth factor (EGF) and insulin, across polarized Madin-Darby canine kidney (MDCK) cells grown on dual-sided chambers on a nitrocellulose filter substrate. When grown on these chambers, MDCK cells are polarized and express distinct basal and apical surfaces. MDCK cells are capable of unidirectional transport of EGF from the basal-to-apical direction, 50% of bound EGF transported in 2 h. Transport was inhibited by the addition of unlabeled EGF in a dose-dependent manner. Anti-EGF receptor Ab, which inhibited binding, also inhibited transport. No transport in the apical-to-basal direction is noted. Insulin transport is not observed in either direction. Transport correlates with the presence of ligand-specific receptors on the cell surface. Hence, EGF receptors (Ro = 48,000, Kd = 3.5 X 10(-10) M) are found only on the basal surface of the MDCK cells and neither surface expresses insulin receptors. Characterization of the EGF receptors on MDCK cells, as assessed by affinity, molecular mass, and anti-receptor antibody binding reveals that this receptor has similar characteristics to EGF receptors previously described on a variety of cells. Hence, the EGF receptor can function as a transporter of EGF across an epithelial cell barrier.

Sign in / Sign up

Export Citation Format

Share Document