Protein Kinase C-α Activity Modulates Transepithelial Permeability and Cell Junctions in the LLC-PK1 Epithelial Cell Line

Rosson, Dan; O’Brien, Thomas G.; Kampherstein, Jennifer A.; Szállási, Zoltán; Bogi, Krisztina; Blumberg, Peter M.; Mullin, James M.

doi:10.1074/jbc.272.23.14950

Cited by 81 publications

(50 citation statements)

References 24 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…In pulmonary epithelial monolayers, PKC␣ activation modulates TNF␣-induced increase in paracellular permeability (31). Similarly, it has been reported (17,18) that PKC␣ and PKC␦ expression were correlated with tight junctional leakiness in renal epithelial cells. Marano et al (33) reported that phorbol ester increased paracellular permeability in IEC-18 intestinal cells by a mechanism involving translocation of PKC␣ from the cytosolic to membrane compartment and phosphorylation of the tight junctional protein occludin.…”

Section: Discussionmentioning

confidence: 99%

“…A substantial body of experimental data indicates that PKC regulates paracellular permeability in epithelial (17,18) and endothelial (31) monolayers. The PKC activators diacylglycerol and phorbol esters increase paracellular permeability and diminish TER (15,32).…”

Section: Discussionmentioning

confidence: 99%

“…The PKC activators diacylglycerol and phorbol esters increase paracellular permeability and diminish TER (15,32). PKC regulates the sorting and assembly of the TJ proteins ZO-1 and the development of TER in confluent renal epithelial cells (17,18). In pulmonary epithelial monolayers, PKC␣ activation modulates TNF␣-induced increase in paracellular permeability (31).…”

Section: Discussionmentioning

confidence: 99%

“…For example, decreased transepithelial electrical resistance (TER) and increased paracellular flux are observed in cells exposed to the PKC agonist (12-O-tetradecanoylphorbol-13-acetate) (16). Both PKC␣ and -␦ are implicated in the PKC-dependent regulation of TJ assembly and barrier function following activation of a G-protein-coupled receptor (17)(18)(19)(20). Freeze fracture electron micrographs reveal abnormal TJ morphology in transfectants overexpressing mutant forms of the Rho family of GTPases, suggesting involvement of Rho proteins in TJ assembly (21).…”

mentioning

confidence: 99%

See 3 more Smart Citations

Protein Kinase C Signaling Regulates ZO-1 Translocation and Increased Paracellular Flux of T84 Colonocytes Exposed toClostridium difficile Toxin A

Chen

Pothoulakis

LaMont

2002

Journal of Biological Chemistry

155

View full text Add to dashboard Cite

Clostridium difficile is an anaerobic pathogen that causes antibiotic-associated diarrhea and colitis in humans (1). Two large molecular weight proteins, toxins A and B, secreted by this microbe are glucosyltransferases with substrate specificity for Rho family GTPases (Rho, Rac, and Cdc42) (2, 3). Rho proteins belong to the Ras superfamily of GTPases that regulate cell morphology, gene transcription, and cell proliferation (4). Rho glucosylation by toxins reduces the binding affinity of Rho to its downstream effectors, resulting in inhibition of Rho signaling, disassembly of actin filaments, and cell rounding (5). However, recent studies indicate that C. difficile toxins induce early cellular responses including activation of mitogen-activated protein kinases (6), generation of reactive oxygen metabolites (7), and induction of calcium influx (8) that occur prior to measurable Rho glucosylation, indicating that these and perhaps other Rho-independent signaling events are also involved in toxicity.Dysfunction of paracellular permeability is a major contributor to the increased fluid secretion and diarrhea in C. difficile infection and other inflammatory bowel diseases (9 -11). Tight junctions (TJs), 1 localized in the uppermost basolateral surface of polarized enterocytes, serve as a barrier regulating selective passage of fluid, ions, and lipids through the paracellular pathway (12, 13). Tight junction fibrils are composed of the transmembrane proteins occludin, claudins, and the junctional adhesion molecule whose cytosolic domains interact with the peripheral junctional proteins of the zonula occludens family (ZO-1, -2, and -3) (13). The C-terminal domains of ZOs, in turn, bind to perijunctional actomyosin filaments that support the TJ complex (14). TJ assembly and permeability are regulated by a network of signaling pathways including protein kinase C (PKC), heterotrimeric G proteins, Ras and Rho GTP-binding proteins, protein kinase A, tyrosine kinase, and calcium (15). For example, decreased transepithelial electrical resistance (TER) and increased paracellular flux are observed in cells exposed to the PKC agonist (12-O-tetradecanoylphorbol-13-acetate) (16). Both PKC␣ and -␦ are implicated in the PKC-dependent regulation of TJ assembly and barrier function following activation of a G-protein-coupled receptor (17)(18)(19)(20). Freeze fracture electron micrographs reveal abnormal TJ morphology in transfectants overexpressing mutant forms of the Rho family of GTPases, suggesting involvement of Rho proteins in TJ assembly (21). In polarized intestinal epithelial cells (T84 cells) infected with a chimeric protein containing Clostridium botulinum C3 exoenzyme that specifically inactivates RhoA, perijunctional actin rings are disrupted, leading to a decrease in TER and redistribution of .The increase in TJ permeability and redistribution of TJ proteins in T84 cells exposed to C. difficile toxins has been correlated with depolymerization of the actin cytoskeleton (23,24). However, we previously observed a rapid drop ...

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

See 2 more Smart Citations

Protein Kinase C Signaling Regulates ZO-1 Translocation and Increased Paracellular Flux of T84 Colonocytes Exposed toClostridium difficile Toxin A

Chen

Pothoulakis

LaMont

2002

Journal of Biological Chemistry

155

View full text Add to dashboard Cite

show abstract

“…While this work was being written, the article by Rosson et al (36) came to our notice; it describes the involvement of 2 E. Batlle and A. Garcia de Herreros, unpublished observations.…”

Section: Cpkc-␣ Regulates Cell Growth and Invasionmentioning

confidence: 99%

Protein Kinase C-α Activity Inversely Modulates Invasion and Growth of Intestinal Cells

Batlle¹,

Verdú²,

Domı́nguez³

et al. 1998

Journal of Biological Chemistry

View full text Add to dashboard Cite

The phorbol ester phorbol 12-myristate 13-acetate induces remarkable phenotypic changes in intestinal HT-29 M6 cells; these changes consist of loss of homotypic adhesion and inactivation of E-cadherin. In parallel, cell growth is retarded. We have transfected HT-29 M6 cells with an activated form of the conventional protein kinase C␣ (cPK-C␣). Expression of this isoform induced the acquisition of a scattered phenotype, similar to that adopted by cells after addition of phorbol 12-myristate 13-acetate, with very low cell-to-cell aggregation and undetectable levels of functional E-cadherin. These cell clones were highly motile and rapidly invaded embryonic chick heart fragments. Furthermore, cells expressing activated-cPK-C␣ showed decreased proliferation in comparison to control clones. We have also studied how these two apparently antagonistic changes affect the tumorigenic ability of HT-29 M6 cells. When the different cell clones were xenografted into athymic mice, the effect on cell growth seemed to predominate. Expression of activated-cPK-C␣ significantly reduced the size of the tumors; the cells with the highest level of expression did not even form subcutaneous tumors. Besides their smaller size, the morphology of these tumors was clearly different from those originated by HT-29 M6 cells, and they could be defined as infiltrative on anatomo-pathological basis. These results indicate that cPK-C␣ controls both cell-to-cell adhesion and proliferation of intestinal cells.

show abstract

Translocation of MARCKS and reorganization of the cytoskeleton by PMA correlates with the ion selectivity, the confluence, and transformation state of kidney epithelial cell lines

et al. 1999

View full text Add to dashboard Cite

The role of protein kinase C (PKC) in the regulation of the cytoskeleton of epithelial cells with tightly sealed contacts, poor contacts, and without contacts were investigated by incubating them with a protein kinase C activator phorbol myristoyl acetate (PMA). The morphology and organization of the membrane skeleton and stress fibers as well as the localization of an actin-bundling PKC substrate MARCKS in confluent MDCK cells originating from the distal tubulus of dog kidney, LLC-PK1 cells originating from the proximal tubulus of pig kidney, src-transformed MDCK cells, epidermoid carcinoma A431 cells, and MDCK cells grown in low calcium medium (LC medium) in low density were visualized with phase contrast and immunofluorescence microscopy. Four different responses to the PMA-treatment in actin-based structures of cultured epithelial cells were observed: 1) disintegration of the membrane skeleton in confluent MDCK cells; 2) depolymerization of the stress fibers in confluent MDCK and LLC-PK1 cells; 3) formation of the membrane skeleton in A431 cells, and 4) formation of the stress fibers and membrane skeleton in LC-MDCK cells. Thus, it seems that in fully confluent tightly sealed epithelium, activation of PKC has a deleterious effect on actin-based structures, whereas in cells without contacts or loose contacts, activation of PKC by PMA results in improvement of actin-based cytoskeletal structures. The main difference between the two kidney cell lines used is their selectivity to ion transport: the monolayer of LLC-PK1 cells is anion selective and MDCK cells cation selective. We propose a model where alterations in the ionic milieu within the MDCK cells by means of cation channels affect the disintegration of the membrane skeleton. The distribution of MARCKS followed the distribution of fodrin in both cell lines upon PMA-treatment, suggesting that phosphorylation of MARCKS by PKC may contribute in the regulation of the integrity of the membrane skeleton.

show abstract

Protein Kinase C-α Activity Modulates Transepithelial Permeability and Cell Junctions in the LLC-PK1 Epithelial Cell Line

Cited by 81 publications

References 24 publications

Protein Kinase C Signaling Regulates ZO-1 Translocation and Increased Paracellular Flux of T84 Colonocytes Exposed toClostridium difficile Toxin A

Protein Kinase C Signaling Regulates ZO-1 Translocation and Increased Paracellular Flux of T84 Colonocytes Exposed toClostridium difficile Toxin A

Protein Kinase C-α Activity Inversely Modulates Invasion and Growth of Intestinal Cells

Translocation of MARCKS and reorganization of the cytoskeleton by PMA correlates with the ion selectivity, the confluence, and transformation state of kidney epithelial cell lines

Contact Info

Product

Resources

About