Cell‐specific diversity in the expression and organization of cytoplasmic plaque proteins of apical junctions

Abstract

Tight and adherens junctions play critical roles in the barrier, adhesion, and signaling functions of epithelial and endothelial cells. How the molecular organization of these junctions is tuned to the widely diverse physiological requirements of each tissue type is not well understood. Here, we address this question by examining the expression, localization, and interactions of major cytoplasmic plaque proteins of tight and adherens junctions in different cultured epithelial and endothelial cell lines. Immunoblotting and immunofluorescence analyses show that the expression profiles of cingulin, paracingulin, ZO-1, ZO-2, ZO-3, PLEKHA7, afadin, PDZD11, p120-catenin, and α-catenin, as well as the transmembrane junctional proteins occludin, E-cadherin, and VE-cadherin, are significantly diverse when comparing kidney cells (MDCK, mCCD), keratinocytes (HaCaT), lung carcinoma (A427, A549), and endothelium-derived cells (bEnd.3, meEC, H5V). Proximity ligation and co-immunoprecipitation assays show that PLEKHA7 and PDZD11 are significantly more associated with the tight junction proteins cingulin and ZO-1 in aortic endothelium-derived (meEC) cells but not kidney collecting duct epithelial (mCCD) cells. These results provide evidence that the cytoplasmic plaques of tight and adherens junctions are diverse in their composition and molecular architecture and establish a conceptual framework by which we can rationally address the mechanisms of tissue-dependent junction physiology and signaling by cytoplasmic junctional proteins.

A Dock-and-Lock Mechanism Clusters ADAM10 at Cell-Cell Junctions to Promote α-Toxin Cytotoxicity

“…The Junctional Clustering of ADAM10 Correlates with Its Maturation, Cell Confluency, and a-Toxin-Dependent Cell Death Using an antibody validated on cells depleted of ADAM10 (Figures S1A-S1D), we examined ADAM10 immunofluorescence (IF) localization in confluent kidney (mouse cortical collecting duct, mCCD]) cells, grown on Transwell filters to enhance apico-basal polarization. ADAM10 was co-localized with PLEKHA7 at apical junctions (arrows, Figures 1A and 1B), which comprise the zonular junctions TJ and ZA, and where the respective markers ZO-1 and PLEKHA7 show partially overlapped localizations (Pulimeno et al, 2010;Vasileva et al, 2017). It should be noted that ZA proteins and some TJ proteins, including ZO-1 (Howarth et al, 1992), are localized at AJs in non-epithelial cells, such as myeloblastic-derived Hap1 cells.…”

PLEKHA5, PLEKHA6 and PLEKHA7 bind to PDZD11 to target the Menkes ATPase ATP7A to the cell periphery and regulate copper homeostasis

“…IB analysis showed distinct patterns of expression for each WW-PLEKHA in epithelial and non-epithelial cell lines (Fig. S3A and (Vasileva et al, 2017)). For IF analysis of endogenous proteins, we used epithelial cells from the collecting duct (mCCD) and the proximal tubule (MDCK) of the kidney, grown to full polarization either on Transwell filters or as cysts in Matrigel, and myeloblastic-derived Hap1 cells (Shah et al, 2018).…”

“…Culture conditions for mouse cortical collecting duct cells (mCCD), Madin-Darby canine kidney cells (MDCKII Tet-off), mouse brain microvascular endothelial (endothelioma) cell line (bEnd.3), mouse heart endothelial cell line (H5V), human lung carcinoma cell line (A427), ciliated aortic mouse embryonic endothelial cells (meEC), human keratinocyte cell line (HaCaT) (Vasileva et al, 2017), human umbilical vascular endothelial cells (HUVEC) (Rouaud et al, 2019), haploid human cells (Hap1) (Popov et al, 2015), mouse mammary epithelial cells (Eph4), human intestinal carcinoma cells (Caco-2) (Spadaro et al, 2017), human embryonic kidney epithelial cells (HEK293T) (Rouaud et al, 2020) were described previously.…”

Elevated Expression of PDZD11 Is Associated With Poor Prognosis and Immune Infiltrates in Hepatocellular Carcinoma

“…Current research reports that the interaction of PDZD11 with PLEKHA7 is significantly associated with tight and adherens junctions (Guerrera et al, 2016;Vasileva et al, 2017). However, to the best of our knowledge, no study has investigated the role of PDZD11 in liver cancer.…”

“…Recent work has also shown that cooperative binding of the tandem WW domains (e.g., WW1 and WW2) of PLEKHA7 to PDZD11 promoted the binding of the C-terminus of Tspan33 to PLEKHA7. Furthermore, the complex formation of PLEKHA7, PDZD11, ADAM10 and its molecular chaperone Tspan33 through promoting the junctional clustering of the α-toxin receptor ADAM10 makes cells more sensitive to the cytotoxic effects of Staphylococcus aureus α-toxin (Vasileva et al, 2017;Rouaud et al, 2020).…”