scholarly journals Putative linear motifs mediate the trafficking to apical and basolateral membranes

2020 ◽  
Author(s):  
Laszlo Dobson ◽  
András Zeke ◽  
Levente Szekeres ◽  
Tamás Langó ◽  
Gábor Tusnády
Keyword(s):  
Epithelial Cells ◽  
Drug Transport ◽  
Basolateral Membrane ◽  
Transmembrane Proteins ◽  
Transport Processes ◽  
Membrane Domains ◽  
Protein Distribution ◽  
Basolateral Membranes ◽  
Linear Motifs ◽  
Cellular Components

AbstractCell polarity refers to the asymmetric organisation of cellular components in various cells. Epithelial cells are the best known examples of polarized cells, featuring apical and basolateral membrane domains. Despite huge efforts, the exact rules governing the protein distribution in such domains are still elusive. In this study we examined linear motifs accumulating in these parts and based on the results we prepared ‘Classical’ and Convolutional Neural Networks to classify human transmembrane proteins localizing into apical/basolateral membranes. Asymmetric expression of drug transporters results in vectorial drug transport, governing the pharmacokinetics of numerous substances, yet the data on how proteins are sorted in epithelial cells is very scattered. The provided dataset may offer help to experimentalists to characterize novel molecular targets to regulate transport processes more precisely.

Localization of a novel 210 kDa protein in Xenopus tight junctions

1997 ◽  
Vol 110 (8) ◽  
pp. 1005-1012 ◽  
Author(s):  
C.S. Merzdorf ◽  
D.A. Goodenough
Keyword(s):  
Monoclonal Antibody ◽  
Epithelial Cells ◽  
Tight Junction ◽  
Tight Junctions ◽  
Basolateral Membrane ◽  
Immunofluorescent Staining ◽  
Junctional Complex ◽  
Permeability Barrier ◽  
Membrane Domains ◽  
Kidney Liver

The tight junction is the most apical member of the intercellular junctional complex. It functions as a permeability barrier between epithelial cells and maintains the integrity of the apical and basolateral membrane domains. In order to study tight junctions in Xenopus laevis, a polyclonal antibody was raised which recognized Xenopus ZO-1. Monoclonal antibody 19B1 (mAb 19B1) was generated in rats using a crude membrane preparation from Xenopus lung as antigen. mAb 19B1 gave immunofluorescent staining patterns identical to those seen with anti-ZO-1 on monolayers of Xenopus A6 kidney epithelial cells and on frozen sections of Xenopus kidney, liver, and embryos. Electron microscopy showed that the 19B1 antigen colocalized with ZO-1 at the tight junction. Western blotting and immunoprecipitation demonstrated that ZO-1 is an approximately 220 kDa protein in Xenopus, while mAb 19B1 identified an approximately 210 kDa antigen on immunoblots. Immunoprecipitates of ZO-1 were not recognized by mAb 19B1 by western analysis. The solubility properties of the 19B1 antigen suggested that it is a peripheral membrane protein. Thus, the antigen recognized by the new monoclonal antibody 19B1 is not ZO-1 and represents a different Xenopus tight junction associated protein.

Selective permeability barrier to urea in shark rectal gland

2005 ◽  
Vol 289 (1) ◽  
pp. F83-F89 ◽  
Author(s):  
Joshua D. Zeidel ◽  
John C. Mathai ◽  
John D. Campbell ◽  
Wily G. Ruiz ◽  
Gerard L. Apodaca ◽  
...  
Keyword(s):  
Activation Energy ◽  
Epithelial Cells ◽  
Water Flow ◽  
Water Permeability ◽  
Apical Membrane ◽  
Basolateral Membrane ◽  
Water Flux ◽  
Rectal Gland ◽  
Basolateral Membranes ◽  
Urea Permeability

Elasmobranchs such as the dogfish shark Squalus acanthius achieve osmotic homeostasis by maintaining urea concentrations in the 300- to 400-mM range, thus offsetting to some degree ambient marine osmolalities of 900–1,000 mosmol/kgH2O. These creatures also maintain salt balance without losing urea by secreting a NaCl-rich (500 mM) and urea-poor (18 mM) fluid from the rectal gland that is isotonic with the plasma. The composition of the rectal gland fluid suggests that its epithelial cells are permeable to water and not to urea. Because previous work showed that lipid bilayers that permit water flux do not block flux of urea, we reasoned that the plasma membranes of rectal gland epithelial cells must either have aquaporin water channels or must have some selective barrier to urea flux. We therefore isolated apical and basolateral membranes from shark rectal glands and determined their permeabilities to water and urea. Apical membrane fractions were markedly enriched for Na-K-2Cl cotransporter, whereas basolateral membrane fractions were enriched for Na-K-ATPase. Basolateral membrane osmotic water permeability (Pf) averaged 4.3 ± 1.3 × 10−3 cm/s, whereas urea permeability averaged 4.2 ± 0.8 × 10−7 cm/s. The activation energy for water flow averaged 16.4 kcal/mol. Apical membrane Pf averaged 7.5 ± 1.6 × 10−4 cm/s, and urea permeability averaged 2.2 ± 0.4 × 10−7 cm/s, with an average activation energy for water flow of 18.6 kcal/mol. The relatively low water permeabilities and high activation energies argue strongly against water flux via aquaporins. Comparison of membrane water and urea permeabilities with those of artificial liposomes and other isolated biological membranes indicates that the basolateral membrane urea permeability is fivefold lower than would be anticipated for its water permeability. These results indicate that the rectal gland maintains a selective barrier to urea in its basolateral membranes.

scholarly journals Basolateral membrane K+ channels in renal epithelial cells

2012 ◽  
Vol 302 (9) ◽  
pp. F1069-F1081 ◽  
Author(s):  
Kirk L. Hamilton ◽  
Daniel C. Devor
Keyword(s):  
Epithelial Cells ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Gene Families ◽  
Proper Function ◽  
Major Function ◽  
Basolateral Membranes ◽  
Family Approach ◽  
Membrane Transport Proteins ◽  
Significant Disease

The major function of epithelial tissues is to maintain proper ion, solute, and water homeostasis. The tubule of the renal nephron has an amazingly simple structure, lined by epithelial cells, yet the segments (i.e., proximal tubule vs. collecting duct) of the nephron have unique transport functions. The functional differences are because epithelial cells are polarized and thus possess different patterns (distributions) of membrane transport proteins in the apical and basolateral membranes of the cell. K+ channels play critical roles in normal physiology. Over 90 different genes for K+ channels have been identified in the human genome. Epithelial K+ channels can be located within either or both the apical and basolateral membranes of the cell. One of the primary functions of basolateral K+ channels is to recycle K+ across the basolateral membrane for proper function of the Na+-K+-ATPase, among other functions. Mutations of these channels can cause significant disease. The focus of this review is to provide an overview of the basolateral K+ channels of the nephron, providing potential physiological functions and pathophysiology of these channels, where appropriate. We have taken a “K+ channel gene family” approach in presenting the representative basolateral K+ channels of the nephron. The basolateral K+ channels of the renal epithelia are represented by members of the KCNK, KCNJ, KCNQ, KCNE, and SLO gene families.

Epithelial localization of a reptilian Na+/H+ exchanger homologous to NHE-1

1997 ◽  
Vol 272 (6) ◽  
pp. G1594-G1606 ◽  
Author(s):  
S. P. Harris ◽  
T. V. Strong ◽  
N. Wys ◽  
N. W. Richards ◽  
J. Pouyssegur ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cellular Localization ◽  
Cell Types ◽  
Transport Processes ◽  
Transepithelial Transport ◽  
Immunofluorescent Localization ◽  
Basolateral Membranes ◽  
Partial Length ◽  
High Level ◽  
Exchange Activity

Basolateral membranes of turtle (Pseudemys scripta) colon epithelial cells exhibit robust Na+/H+ exchange activity that can be activated by cell shrinkage and is blocked by amiloride [M. A. Post and D. C. Dawson. Am. J. Physiol. 262 Cell Physiol. 31):C1089-C1094, 1992]. The colonic epithelium actively absorbs Na+ and secretes K+ and HCO3-, but the role of basolateral Na+/H+ exchange, if any, in transepithelial transport is unknown. The current studies were undertaken to identify the gene product(s) responsible for the observed basolateral Na+/H+ exchange activity and to determine the cellular localization of the reptilian Na+/H+ exchange protein. We cloned and sequenced partial-length cDNAs that are likely to encode a reptilian homologue of the mammalian NHE-1 Na+/H+ exchanger isoform. The partial-length cDNAs were > 80% identical to mammalian NHE-1 homologues at the nucleotide level and recognized a transcript (approximately 5.8-6.0 kb) in RNA isolated from turtle colon, small intestine, stomach, kidney, urinary bladder, heart, and liver. In situ hybridization showed that mRNA encoding the reptile homologue of NHE-1 was expressed predominantly in the epithelial cells of these tissues. Immunofluorescent localization of the reptilian Na+/H+ exchanger protein using an antibody raised against a human NHE-1 fusion protein confirmed that protein expression paralleled abundant mRNA expression in epithelial cells of turtle stomach and colon, as well as in some nephron segments, and showed that the reptile NHE-1 homologue was localized exclusively to the basolateral membranes of these cells. The relatively high level of NHE-1 expression in epithelial cells, particularly those of the colon and stomach, suggests that NHE-1 function is important for the maintenance or regulation of ion transport processes that occur in these cell types.

scholarly journals Mechanisms and functional features of polarized membrane traffic in epithelial and hepatic cells

1998 ◽  
Vol 336 (2) ◽  
pp. 257-269 ◽  
Author(s):  
Mirjam M. P. ZEGERS ◽  
Dick HOEKSTRA
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Apical Membrane ◽  
Basolateral Membrane ◽  
Transport Processes ◽  
Protein Kinase Activity ◽  
Transport Pathway ◽  
Hepatic Cells ◽  
Functional Features ◽  
Specific Lipid

Epithelial cells express plasma-membrane polarity in order to meet functional requirements that are imposed by their interaction with different extracellular environments. Thus apical and basolateral membrane domains are distinguished that are separated by tight junctions in order to maintain the specific lipid and protein composition of each domain. In hepatic cells, the plasma membrane is also polarized, containing a sinusoidal (basolateral) and a bile canalicular (apical)-membrane domain. Relevant to the biogenesis of these domains are issues concerning sorting, (co-)transport and regulation of transport of domain-specific membrane components. In epithelial cells, specific proteins and lipids, destined for the apical membrane, are sorted in the trans-Golgi network (TGN), which involves their sequestration into cholesterol/sphingolipid ‘rafts ’, followed by ‘direct ’ transport to the apical membrane. In hepatic cells, a direct apical transport pathway also exists, as revealed by transport of sphingolipids from TGN to the apical membrane. This is remarkable, since in these cells numerous apical membrane proteins are ‘indirectly ’ sorted, i.e. they are first transferred to the basolateral membrane prior to their subsequent transcytosis to the apical membrane. This raises intriguing questions as to the existence of specific lipid rafts in hepatocytes. As demonstrated in studies with HepG2 cells, it has become evident that, in hepatic cells, apical transport pathways can be regulated by protein kinase activity, which in turn modulates cell polarity. Finally, an important physiological function of hepatic cells is their involvement in intracellular transport and secretion of bile-specific lipids. Mechanisms of these transport processes, including the role of multidrug-resistant proteins in lipid translocation, will be discussed in the context of intracellular vesicular transport. Taken together, hepatic cell systems provide an important asset to studies aimed at elucidating mechanisms of sorting and trafficking of lipids (and proteins) in polarized cells in general.

scholarly journals Viscoelasticity of basal plasma membranes and cortices derived from MDCK II cells

2021 ◽  
Author(s):  
Andreas Janshoff
Keyword(s):  
Mechanical Properties ◽  
Epithelial Cells ◽  
Plasma Membranes ◽  
Basolateral Membrane ◽  
Scaling Law ◽  
Focal Adhesions ◽  
Living Cells ◽  
The Body ◽  
Basolateral Membranes ◽  
Apical Side

In mature epithelial cells, however, cells adhere to one another through tight junctions, adherens junctions and desmosomes thereby displaying a pronounced apical-basal polarity. In vivo, the apical membrane has a larger surface area and faces the outer surface of the body or the lumen of internal cavities, whereas the basolateral membrane is oriented on the side away from the lumen and forms focal adhesions with the extracellular matrix. The mechanical properties of cells are largely determined by the architecture and dynamics of their viscoelastic cortex, which consists of a contractile, cross-linked actin mesh attached to the plasma membrane via linker proteins. Measuring the mechanical properties of adherent, polarized epithelial cells is usually limited to the upper, i.e., apical side of the cells due to their accessibility on culture dishes. Moreover, contributions from the cell interior comprising various filament types, organelles, and the crowded cytoplasm usually impede examination of the cortex alone. Here, we investigate the viscoelastic properties of basolateral membranes derived from polarized MDCK II epithelia in response to external deformation and compare them to living cells probed at the apical side. Therefore, we grew MDCK II cells on porous surfaces to confluency and removed the upper cell body by sandwich cleavage. The free-standing, defoliated cortices were subject to force indentation and relaxation experiments permitting a precise assessment of cortical viscoelasticity. A new theoretical framework to describe the force cycles is developed and applied to obtain the time-dependent area compressibility modulus of cell cortices from adherent cells. Compared to the viscoelastic response of living cells the basolateral membranes are substantially less fluid and stiffer but obey to the same universal scaling law if excess area is taken into account.

Chloride transport in rabbit esophageal epithelial cells

2002 ◽  
Vol 282 (4) ◽  
pp. G663-G675 ◽  
Author(s):  
Solange Abdulnour-Nakhoul ◽  
Nazih L. Nakhoul ◽  
Canan Caymaz-Bor ◽  
Roy C. Orlando
Keyword(s):  
Epithelial Cells ◽  
Apical Membrane ◽  
Chloride Transport ◽  
Ussing Chamber ◽  
Basolateral Membrane ◽  
Rate Of Change ◽  
Luminal Cell ◽  
Transport Pathways ◽  
Basolateral Membranes ◽  
Esophageal Epithelial Cells

We investigated Cl− transport pathways in the apical and basolateral membranes of rabbit esophageal epithelial cells (EEC) using conventional and ion-selective microelectrodes. Intact sections of esophageal epithelium were mounted serosal or luminal side up in a modified Ussing chamber, where transepithelial potential difference and transepithelial resistance could be determined. Microelectrodes were used to measure intracellular Cl− activity (a[Formula: see text]), basolateral or apical membrane potentials ( V mBL or V mC), and the voltage divider ratio. When a basal cell was impaled, V mBL was −73 ± 4.3 mV and a[Formula: see text] was 16.4 ± 2.1 mM, which were similar in presence or absence of bicarbonate. Removal of serosal Cl−caused a transient depolarization of V mBL and a decrease in a[Formula: see text] of 6.5 ± 0.9 mM. The depolarization and the rate of decrease of a[Formula: see text] were inhibited by ∼60% in the presence of the Cl−-channel blocker flufenamate. Serosal bumetanide significantly decreased the rate of change of a[Formula: see text] on removal and readdition of serosal Cl−. When a luminal cell was impaled, V mC was −65 ± 3.6 mV and a[Formula: see text] was 16.3 ± 2.2 mM. Removal of luminal Cl− depolarized V mC and decreased a[Formula: see text] by only 2.5 ± 0.9 mM. Subsequent removal of Cl− from the serosal bath decreased a[Formula: see text]in the luminal cell by an additional 6.4 ± 1.0 mM. A plot of V mBL measurements vs. log a[Formula: see text]/log a[Formula: see text] (a[Formula: see text] is the activity of Cl− in a luminal or serosal bath) yielded a straight line [slope ( S) = 67.8 mV/decade of change in a[Formula: see text]/a[Formula: see text]]. In contrast, V mC correlated very poorly with log a[Formula: see text]/a[Formula: see text] ( S = 18.9 mV/decade of change in a[Formula: see text]/a[Formula: see text]). These results indicate that 1) in rabbit EEC, a[Formula: see text] is higher than equilibrium across apical and basolateral membranes, and this process is independent of bicarbonate; 2) the basolateral cell membrane possesses a conductive Cl− pathway sensitive to flufenamate; and 3) the apical membrane has limited permeability to Cl−, which is consistent with the limited capacity for transepithelial Cl− transport. Transport of Cl− at the basolateral membrane is likely the dominant pathway for regulation of intracellular Cl−.

Dependence of intracellular Na+ concentration on apical and basolateral membrane Na+ influx in frog skin

1985 ◽  
Vol 249 (5) ◽  
pp. F662-F671
Author(s):  
J. S. Stoddard ◽  
S. I. Helman
Keyword(s):  
Epithelial Cells ◽  
Frog Skin ◽  
Apical Membrane ◽  
Basolateral Membrane ◽  
Na Transport ◽  
Maximal Inhibition ◽  
Basolateral Membranes ◽  
Isotopic Method ◽  
Rate Of Increase ◽  
Kinetics Of

An isotopic method was developed to measure the intracellular Na+ content of the transepithelial Na+ transport pool of frog skin. Isolated epithelia (no corium) were labeled with 24Na either asymmetrically, from apical (Aa) or basolateral (Ab) solutions, or symmetrically (Aab). Transport pool Na+ could be identified from the kinetics of washout of 24Na carried out in the presence of 1 mM ouabain, 100 microM amiloride, and 1 mM furosemide that served to trap cold Na+ and 24Na within the transport pool. In control epithelia, Aab averaged 64.1 neq/cm2 (13.9 mM), and maximal inhibition of apical membrane Na+ entry with 100 microM amiloride caused Aab to decrease to 24.3 neq/cm2 (5.3 mM). Ouabain caused Aab to increase markedly to 303 neq/cm2 in 30 min, whereas amiloride inhibition of apical membrane Na+ entry reduced markedly the rate of increase of Aab caused by ouabain (7.3 neq X cm-2 X min-1 in control and 1.7 neq X cm-2 X min-1 in the presence of amiloride). These data, in part, confirmed the existence of an important basolateral membrane permeability to Na+ that was measured in separate studies of the bidirectional 24Na fluxes at the basolateral membranes of the cells. Both sets of data were supportive of the idea that a significant Na+ recycling exists at the basolateral membranes of the cells that contributes to the Na+ load on the pump and Na+ recycling participates in the regulation of the Na+ concentration of the Na+ transport pool of these epithelial cells.

scholarly journals Epithelial polarity requires septin coupling of vesicle transport to polyglutamylated microtubules

2008 ◽  
Vol 180 (2) ◽  
pp. 295-303 ◽  
Author(s):  
Elias T. Spiliotis ◽  
Stephen J. Hunt ◽  
Qicong Hu ◽  
Makoto Kinoshita ◽  
W. James Nelson
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Basolateral Membrane ◽  
Vesicle Transport ◽  
Epithelial Polarity ◽  
Membrane Domains ◽  
Polarized Epithelia ◽  
Membrane Growth ◽  
Basolateral Plasma Membrane ◽  
Golgi Network

In epithelial cells, polarized growth and maintenance of apical and basolateral plasma membrane domains depend on protein sorting from the trans-Golgi network (TGN) and vesicle delivery to the plasma membrane. Septins are filamentous GTPases required for polarized membrane growth in budding yeast, but whether they function in epithelial polarity is unknown. Here, we show that in epithelial cells septin 2 (SEPT2) fibers colocalize with a subset of microtubule tracks composed of polyglutamylated (polyGlu) tubulin, and that vesicles containing apical or basolateral proteins exit the TGN along these SEPT2/polyGlu microtubule tracks. Tubulin-associated SEPT2 facilitates vesicle transport by maintaining polyGlu microtubule tracks and impeding tubulin binding of microtubule-associated protein 4 (MAP4). Significantly, this regulatory step is required for polarized, columnar-shaped epithelia biogenesis; upon SEPT2 depletion, cells become short and fibroblast-shaped due to intracellular accumulation of apical and basolateral membrane proteins, and loss of vertically oriented polyGlu microtubules. We suggest that septin coupling of the microtubule cytoskeleton to post-Golgi vesicle transport is required for the morphogenesis of polarized epithelia.

scholarly journals The establishment of polarized membrane traffic in Xenopus laevis embryos.

1992 ◽  
Vol 118 (6) ◽  
pp. 1359-1369 ◽  
Author(s):  
S J Roberts ◽  
D S Leaf ◽  
H P Moore ◽  
J C Gerhart
Keyword(s):  
Epithelial Cells ◽  
Basolateral Membrane ◽  
Constitutive Expression ◽  
Membrane Traffic ◽  
Peptide Hormone ◽  
Membrane Domains ◽  
Rna Transcripts ◽  
Transmembrane Glycoprotein ◽  
Polarized Epithelial Cells ◽  
Oocyte Membrane

Delineation of apical and basolateral membrane domains is a critical step in the epithelialization of the outer layer of cells in the embryo. We have examined the initiation of polarized membrane traffic in Xenopus and show that membrane traffic is not polarized in oocytes but polarized membrane domains appear at first cleavage. The following proteins encoded by injected RNA transcripts were used as markers to monitor membrane traffic: (a) VSV G, a transmembrane glycoprotein preferentially inserted into the basolateral surface of polarized epithelial cells; (b) GThy-1, a fusion protein of VSV G and Thy-1 that is localized to the apical domains of polarized epithelial cells; and (c) prolactin, a peptide hormone that is not polarly secreted. In immature oocytes, there is no polarity in the expression of VSV G or GThy-1, as shown by the constitutive expression of both proteins at the surface in the animal and vegetal hemispheres. At meiotic maturation, membrane traffic to the surface is blocked; the plasma membrane no longer accepts the vesicles synthesized by the oocyte (Leaf, D. L., S. J. Roberts, J. C. Gerhart, and H.-P. Moore. 1990. Dev. Biol. 141:1-12). When RNA transcripts are injected after fertilization, VSV G is expressed only in the internal cleavage membranes (basolateral orientation) and is excluded from the outer surface (apical orientation, original oocyte membrane). In contrast, GThy-1 and prolactin, when expressed in embryos, are inserted or released at both the outer membrane derived from the oocyte and the inner cleavage membranes. Furthermore, not all of the cleavage membrane comes from an embryonic pool of vesicles--some of the cleavage membrane comes from vesicles synthesized during oogenesis. Using prolactin as a marker, we found that a subset of vesicles synthesized during oogenesis was only released after fertilization. However, while embryonic prolactin was secreted from both apical and basolateral surfaces, the secretion of oogenic prolactin was polarized. Oogenic prolactin was secreted only into the blastocoel (from the cleavage membrane), none could be detected in the external medium (from the original oocyte membrane). These results provide the first direct evidence that the oocyte synthesizes a cache of vesicles for specific recruitment to the embryonic cleavage membranes which are polarized beginning with the first cleavage division.

Sign in / Sign up

Export Citation Format

Share Document