scholarly journals Distinct claudins and associated PDZ proteins form different autotypic tight junctions in myelinating Schwann cells

2002 ◽  
Vol 159 (2) ◽  
pp. 361-372 ◽  
Author(s):  
Sebastian Poliak ◽  
Sean Matlis ◽  
Christoph Ullmer ◽  
Steven S. Scherer ◽  
Elior Peles
Keyword(s):  
Tight Junction ◽  
Tight Junctions ◽  
Schwann Cells ◽  
Pdz Domain ◽  
Adaptor Protein ◽  
Dependent Manner ◽  
Guanylate Kinase ◽  
Pdz Proteins ◽  
Junction Protein ◽  
Zona Occludens

The apposed membranes of myelinating Schwann cells are joined by several types of junctional specializations known as autotypic or reflexive junctions. These include tight, gap, and adherens junctions, all of which are found in regions of noncompact myelin: the paranodal loops, incisures of Schmidt-Lanterman, and mesaxons. The molecular components of autotypic tight junctions have not been established. Here we report that two homologues of Discs Lost–multi PDZ domain protein (MUPP)1, and Pals-associated tight junction protein (PATJ), are differentially localized in myelinating Schwann cells and associated with different claudins. PATJ is mainly found at the paranodal loops, where it colocalized with claudin-1. MUPP1 and claudin-5 colocalized in the incisures, and the COOH-terminal region of claudin-5 interacts with MUPP1 in a PSD-95/Disc Large/zona occludens (ZO)-1 (PDZ)-dependent manner. In developing nerves, claudin-5 and MUPP1 appear together in incisures during the first postnatal week, suggesting that they coassemble during myelination. Finally, we show that the incisures also contain four other PDZ proteins that are found in epithelial tight junctions, including three membrane-associated guanylate-kinase proteins (membrane-associated guanylate-kinase inverted-2, ZO-1, and ZO-2) and the adaptor protein Par-3. The presence of these different tight junction proteins in regions of noncompact myelin may be required to maintain the intricate cytoarchitecture of myelinating Schwann cells.

scholarly journals The Maguk protein, Pals1, functions as an adapter, linking mammalian homologues of Crumbs and Discs Lost

2002 ◽  
Vol 157 (1) ◽  
pp. 161-172 ◽  
Author(s):  
Michael H. Roh ◽  
Olga Makarova ◽  
Chia-Jen Liu ◽  
Shin ◽  
Seonok Lee ◽  
...  
Keyword(s):  
Tight Junctions ◽  
Protein Targeting ◽  
Pdz Domain ◽  
Guanylate Kinase ◽  
Src Homology 3 ◽  
Discs Large ◽  
Domain Of Unknown Function ◽  
Src Homology ◽  
Zona Occludens ◽  
Maguk Proteins

Membrane-associated guanylate kinase (Maguk) proteins are scaffold proteins that contain PSD-95–Discs Large–zona occludens-1 (PDZ), Src homology 3, and guanylate kinase domains. A subset of Maguk proteins, such as mLin-2 and protein associated with Lin-7 (Pals)1, also contain two L27 domains: an L27C domain that binds mLin-7 and an L27N domain of unknown function. Here, we demonstrate that the L27N domain targets Pals1 to tight junctions by binding to a PDZ domain protein, Pals1-associated tight junction (PATJ) protein, via a unique Maguk recruitment domain. PATJ is a homologue of Drosophila Discs Lost, a protein that is crucial for epithelial polarity and that exists in a complex with the apical polarity determinant, Crumbs. PATJ and a human Crumbs homologue, CRB1, colocalize with Pals1 to tight junctions, and CRB1 interacts with PATJ albeit indirectly via binding the Pals1 PDZ domain. In agreement, we find that a Drosophila homologue of Pals1 participates in identical interactions with Drosophila Crumbs and Discs Lost. This Drosophila Pals1 homologue has been demonstrated recently to represent Stardust, a crucial polarity gene in Drosophila. Thus, our data identifies a new multiprotein complex that appears to be evolutionarily conserved and likely plays an important role in protein targeting and cell polarity.

scholarly journals The SARS Coronavirus E Protein Interacts with PALS1 and Alters Tight Junction Formation and Epithelial Morphogenesis

2010 ◽  
Vol 21 (22) ◽  
pp. 3838-3852 ◽  
Author(s):  
Kim-Tat Teoh ◽  
Yu-Lam Siu ◽  
Wing-Lim Chan ◽  
Marc A. Schlüter ◽  
Chia-Jen Liu ◽  
...  
Keyword(s):  
Tight Junction ◽  
Tight Junctions ◽  
Mammalian Cells ◽  
Pdz Domain ◽  
Ectopic Expression ◽  
Binding Motif ◽  
Dependent Manner ◽  
Tight Junction Formation ◽  
Golgi Region ◽  
Carboxy Terminal

Intercellular tight junctions define epithelial apicobasal polarity and form a physical fence which protects underlying tissues from pathogen invasions. PALS1, a tight junction-associated protein, is a member of the CRUMBS3-PALS1-PATJ polarity complex, which is crucial for the establishment and maintenance of epithelial polarity in mammals. Here we report that the carboxy-terminal domain of the SARS-CoV E small envelope protein (E) binds to human PALS1. Using coimmunoprecipitation and pull-down assays, we show that E interacts with PALS1 in mammalian cells and further demonstrate that the last four carboxy-terminal amino acids of E form a novel PDZ-binding motif that binds to PALS1 PDZ domain. PALS1 redistributes to the ERGIC/Golgi region, where E accumulates, in SARS-CoV–infected Vero E6 cells. Ectopic expression of E in MDCKII epithelial cells significantly alters cyst morphogenesis and, furthermore, delays formation of tight junctions, affects polarity, and modifies the subcellular distribution of PALS1, in a PDZ-binding motif-dependent manner. We speculate that hijacking of PALS1 by SARS-CoV E plays a determinant role in the disruption of the lung epithelium in SARS patients.

GENETIC SUSCEPTIBILITY TO IBD OPERATES VIA CONTROL OF APICAL JUNCTIONAL COMPLEXES

2021 ◽  
Vol 27 (Supplement_1) ◽  
pp. S30-S30
Author(s):  
Isabelle Hébert-Milette ◽  
Chloé Lévesque ◽  
Guy Charron ◽  
John Rioux
Keyword(s):  
Tight Junction ◽  
Tight Junctions ◽  
Intestinal Inflammation ◽  
Protein Localization ◽  
Junctional Complex ◽  
Epithelial Permeability ◽  
3D Cultures ◽  
Apical Junctional Complex ◽  
Junction Protein ◽  
The Impact

Abstract Introduction Intestinal permeability is increased in unaffected 1st degree relatives of patients with inflammatory bowel disease (IBD), and is considered a risk factor for the development of IBD, likely increasing the interactions between intestinal microorganisms and the immune system. We recently reported that C1orf106, a gene located within a genomic region associated with IBD, regulates epithelial permeability. We further demonstrated that a rare coding variant within C1orf106 (p.Y333F) decreases protein stability and that lower levels of C1orf106 protein leads altered stability of adherens junctions (AJ) and to an increase in epithelial permeability. Hypothesis In addition to altering AJ, we believe that C1orf106 is also involved in the regulation of tight junction (TJ) formation, which also impacts epithelial permeability. Objectives The objectives of the project are to (a) validate the impact of C1orf106 on tight junctions and (b) verify the impact of C1orf106 IBD-associated variants on intestinal barrier integrity. Results We observed that knocking down the expression of C1orf106 in Caco-2 cells leads to a number of phenotypes in human epithelial monolayer (2D) and spheroid (3D) cultures that are associated with alterations in TJs. Specifically, when studying the dynamic reformation of TJ in 2D cultures after transient withdrawal of calcium, which is required for TJ stability, we observed that lower levels of C1orf106 resulted in (1) decreased recovery of barrier function as measured by transepithelial electrical resistance (TEER); (2) an alteration of tight junction protein localization; and (3) thickening of the circumferential actin belt. Moreover, in 3D cultures, we observed an altered spheroid formation associated with impaired epithelial polarization. In addition, our preliminary studies of human induced pluripotent stem cell (hiPSC)-derived epithelial cultures support that Y333F heterozygotes also have altered structure and function of their tight junctions. Conclusion Our observations indicate an important role of C1orf106 in apical junctional complex (AJC) formation likely mediated by a regulation of the circumferential actin belt. This can affect other functions of AJC, like the establishment of cell polarity. AJC formation is important for epithelial repair after an injury and its dysregulation impairs the formation of an impermeable epithelial barrier, which likely facilitates the passage of microorganisms and the induction and maintenance of intestinal inflammation.

scholarly journals Regulation of testicular tight junctions by gonadotrophins in the adult Djungarian hamster in vivo

Reproduction ◽  
2008 ◽  
Vol 135 (6) ◽  
pp. 867-877 ◽  
Author(s):  
Gerard A Tarulli ◽  
Sarah J Meachem ◽  
Stefan Schlatt ◽  
Peter G Stanton
Keyword(s):  
Tight Junction ◽  
Adaptor Protein ◽  
Tight Junction Protein ◽  
Mrna Levels ◽  
Tight Junction Proteins ◽  
Djungarian Hamster ◽  
Junction Protein ◽  
Protein Localisation ◽  
Junction Proteins

This study aimed to assess the effect of gonadotrophin suppression and FSH replacement on testicular tight junction dynamics and blood–testis barrier (BTB) organisation in vivo, utilising the seasonal breeding Djungarian hamster. Confocal immunohistology was used to assess the cellular organisation of tight junction proteins and real-time PCR to quantify tight junction mRNA. The effect of tight junction protein organisation on the BTB permeability was also investigated using a biotin-linked tracer. Tight junction protein (claudin-3, junctional adhesion molecule (JAM)-A and occludin) localisation was present but disorganised after gonadotrophin suppression, while mRNA levels (claudin-11, claudin-3 and occludin) were significantly (two- to threefold) increased. By contrast, both protein localisation and mRNA levels for the adaptor protein zona occludens-1 decreased after gonadotrophin suppression. FSH replacement induced a rapid reorganisation of tight junction protein localisation. The functionality of the BTB (as inferred by biotin tracer permeation) was found to be strongly associated with the organisation and localisation of claudin-11. Surprisingly, JAM-A was also recognised on spermatogonia, suggesting an additional novel role for this protein in trans-epithelial migration of germ cells across the BTB. It is concluded that gonadotrophin regulation of tight junction proteins forming the BTB occurs primarily at the level of protein organisation and not gene transcription in this species, and that immunolocalisation of the organised tight junction protein claudin-11 correlates with BTB functionality.

Campylobacter jejuni inhibits the absorptive transport functions of Caco-2 cells and disrupts cellular tight junctions

Microbiology ◽  
2005 ◽  
Vol 151 (7) ◽  
pp. 2451-2458 ◽  
Author(s):  
Amanda MacCallum ◽  
Simon P. Hardy ◽  
Paul H. Everest
Keyword(s):  
Tight Junction ◽  
Tight Junctions ◽  
Campylobacter Jejuni ◽  
Fluid Transport ◽  
Cell Function ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Ussing Chambers ◽  
Cell Monolayers ◽  
Junction Protein

Caco-2 cells are models of absorptive enterocytes. The net transport of fluid from apical to basolateral surfaces results in ‘domes' forming in differentiated monolayers. Here, the effect of Campylobacter jejuni on this process has been examined. C. jejuni caused no changes in short-circuit current upon infection of Caco-2 cell monolayers in Ussing chambers. Thus, no active secretory events could be demonstrated using this model. It was therefore hypothesized that C. jejuni could inhibit the absorptive function of enterocytes and that this may contribute to diarrhoeal disease. C. jejuni infection of fluid-transporting (‘doming’) Caco-2 cells resulted in a significant reduction in dome number, which correlated with a decrease in tight junction integrity in infected monolayers, when measured as transepithelial electrical resistance. Defined mutants of C. jejuni also reduced dome numbers in infected monolayers. C. jejuni also altered the distribution of the tight junction protein occludin within cell monolayers. The addition to monolayers of extracellular gentamicin prevented these changes, indicating the contribution of extracellular bacteria to this process. Thus, tight junction integrity is required for fluid transport in Caco-2 cell monolayers as leaky tight junctions cannot maintain support of transported fluid at the basolateral surface of infected cell monolayers. Inhibition of absorptive cell function, changes in epithelial resistance and rearrangement of tight junctional proteins such as occludin represent a potential diarrhoeal mechanism of C. jejuni.

scholarly journals Sorting nexin 27 rescues neuroligin 2 from lysosomal degradation to control inhibitory synapse number

2019 ◽  
Vol 476 (2) ◽  
pp. 293-306 ◽  
Author(s):  
Caroline S. Binda ◽  
Yasuko Nakamura ◽  
Jeremy M. Henley ◽  
Kevin A. Wilkinson
Keyword(s):  
Adaptor Protein ◽  
Inhibitory Synapse ◽  
Endosomal Trafficking ◽  
Dependent Manner ◽  
Lysosomal Degradation ◽  
Sorting Nexin ◽  
The Core ◽  
Cargo Proteins ◽  
Vacuolar Protein ◽  
Zona Occludens

Abstract Retromer is an evolutionarily conserved endosomal trafficking complex that mediates the retrieval of cargo proteins from a degradative pathway for sorting back to the cell surface. To promote cargo recycling, the core retromer trimer of VPS (vacuolar protein sorting)26, VPS29 and VPS35 recognises cargo either directly, or through an adaptor protein, the most well characterised of which is the PDZ [postsynaptic density 95 (PSD95), disk large, zona occludens] domain-containing sorting nexin SNX27. Neuroligins (NLGs) are postsynaptic trans-synaptic scaffold proteins that function in the clustering of postsynaptic proteins to maintain synaptic stability. Here, we show that each of the NLGs (NLG1–3) bind to SNX27 in a direct PDZ ligand-dependent manner. Depletion of SNX27 from neurons leads to a decrease in levels of each NLG protein and, for NLG2, this occurs as a result of enhanced lysosomal degradation. Notably, while depletion of the core retromer component VPS35 leads to a decrease in NLG1 and NLG3 levels, NLG2 is unaffected, suggesting that, for this cargo, SNX27 acts independently of retromer. Consistent with loss of SNX27 leading to enhanced lysosomal degradation of NLG2, knockdown of SNX27 results in fewer NLG2 clusters in cultured neurons, and loss of SNX27 or VPS35 reduces the size and number of gephyrin clusters. Together, these data indicate that NLGs are SNX27–retromer cargoes and suggest that SNX27–retromer controls inhibitory synapse number, at least in part through trafficking of NLG2.

scholarly journals The Heat-Shock Protein Apg-2 Binds to the Tight Junction Protein ZO-1 and Regulates Transcriptional Activity of ZONAB

2006 ◽  
Vol 17 (3) ◽  
pp. 1322-1330 ◽  
Author(s):  
Anna Tsapara ◽  
Karl Matter ◽  
Maria S. Balda
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Tight Junction ◽  
Intracellular Signaling ◽  
Transcriptional Activity ◽  
Adaptor Protein ◽  
Sh3 Domain ◽  
Nucleic Acid Binding ◽  
Junction Protein

The tight junction adaptor protein ZO-1 regulates intracellular signaling and cell proliferation. Its Src homology 3 (SH3) domain is required for the regulation of proliferation and binds to the Y-box transcription factor ZO-1-associated nucleic acid binding protein (ZONAB). Binding of ZO-1 to ZONAB results in cytoplasmic sequestration and hence inhibition of ZONAB's transcriptional activity. Here, we identify a new binding partner of the SH3 domain that modulates ZO-1–ZONAB signaling. Expression screening of a cDNA library with a fusion protein containing the SH3 domain yielded a cDNA coding for Apg-2, a member of the heat-shock protein 110 (Hsp 110) subfamily of Hsp70 heat-shock proteins, which is overexpressed in carcinomas. Regulated depletion of Apg-2 in Madin-Darby canine kidney cells inhibits G1/S phase progression. Apg-2 coimmunoprecipitates with ZO-1 and partially localizes to intercellular junctions. Junctional recruitment and coimmunoprecipitation with ZO-1 are stimulated by heat shock. Apg-2 competes with ZONAB for binding to the SH3 domain in vitro and regulates ZONAB's transcriptional activity in reporter gene assays. Our data hence support a model in which Apg-2 regulates ZONAB function by competing for binding to the SH3 domain of ZO-1 and suggest that Apg-2 functions as a regulator of ZO-1–ZONAB signaling in epithelial cells in response to cellular stress.

Two classes of tight junctions are revealed by ZO-1 isoforms

1993 ◽  
Vol 264 (4) ◽  
pp. C918-C924 ◽  
Author(s):  
M. S. Balda ◽  
J. M. Anderson
Keyword(s):  
Tight Junction ◽  
Tight Junctions ◽  
Rna Splicing ◽  
Seminiferous Tubules ◽  
Junction Protein ◽  
Acid Domain ◽  
Ribonuclease Protection ◽  
Tissue Compartments ◽  
Junctional Resistance ◽  
Amino Acid Domain

The tight junction forms the intercellular barrier separating tissue compartments. The characteristics of this barrier are remarkably diverse among different epithelia and endothelia and are not explained by our limited knowledge of its molecular composition. Two isoforms of the 220-kDa tight junction protein ZO-1 result from alternative RNA splicing and differ by an internal 80-amino acid domain, termed alpha (E. Willott, M. S. Balda, M. Heintzman, B. Jameson, and J. M. Anderson. Am. J. Physiol. 262 (Cell Physiol. 31): C1119-C1124, 1992). Using antibodies specific for each isoform and double-labeled immunofluorescence microscopy, we observed that the ZO-1 alpha- isoform is restricted to junctions of endothelial cells and highly specialized epithelial cells of both seminiferous tubules (Sertoli cells) and renal glomeruli (podocytes); in contrast, the ZO-1 alpha+ isoform is expressed in cells of all other epithelia examined. Both immunoblotting and ribonuclease protection analysis confirmed this pattern of expression. This distribution does not correlate with differences in junctional resistance or ultrastructural complexity. Instead, we observe a correlation with junctional plasticity; ZO-1 alpha- is expressed in structurally dynamic junctions, whereas ZO-1 alpha+ is expressed in those which are less dynamic. This is the first molecular distinction among tight junctions and reveals a fundamental dichotomy with implications for how the paracellular barriers of endothelia and epithelia are regulated.

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