Junctional Adhesion Molecule 3 Expression in the Mouse Airway Epithelium Is Linked to Multiciliated Cells

Abstract

Tight-junction (TJ) proteins are essential for establishing the barrier function between neighbor epithelial cells, but also for recognition of pathogens or cell migration. Establishing the expression pattern and localization of different TJ proteins will help to understand the development and physiology of the airway. Here we identify that the junctional adhesion molecule 3 (Jam3) expression is restricted to multiciliated cells (MCCs) in the airway epithelium. In vitro, Jam3 expression varies along airway basal stem cell (BSC) differentiation and upon DAPT treatment or IL6 exposure. However, Jam3 is not required for BSC differentiation to specific cell types. In addition, we found that MCC lacking Jam3 display normal cilia morphology and cilia beating frequency with a delay in BB assembly/positioning in MCCs during differentiation. Remarkably, Jam3 in MCC is mostly localized to subapical organelles, which are negative for the apical recycling endosome marker Rab11 and positive for EEA1. Our data show that Jam3 expression is connected to mature MCC in the airway epithelium and suggest a Jam3 role unrelated to its known barrier function.

Quantifying cilia beat frequency using high‐speed video microscopy: Assessing frame rate requirements when imaging different ciliated tissues

“…Ns = not significantly different (p > 0.05). Mateos-Quiros et al, 2021;Mikhailik et al, 2021;Reula et al, 2021;Smith et al, 2012;Zahid et al, 2020). The lack of a standardized fps for imaging motile cilia is especially problematic because it hinders high-speed video microscopy from being clinically adapted as a diagnostic tool to help uncover motile cilia diseases, such as primary ciliary dyskinesia (Bricmont et al, 2021;Shapiro et al, 2018).…”

“…The easiest way to assess motile cilia function is by quantifying CBF via highspeed video microscopy (Francis & Lo, 2013 ; O'Callaghan et al, 2012 ; Sisson et al, 2003 ) and this forms a sizable chunk of the cilia literature. However, no standardized frame rate exists when imaging cilia motility for subsequent CBF calculation, this has resulted in a large range of frames rates used to image cilia motility within the published literature (Abdelhamed et al, 2018 ; Abdelhamed et al, 2020 ; Bustamante‐Marin et al, 2019 ; Chen et al, 2016 ; Hagiwara et al, 2020 ; Hennessy et al, 1986 ; Liu et al, 2021 ; Mateos‐Quiros et al, 2021 ; Mikhailik et al, 2021 ; Reula et al, 2021 ; Smith et al, 2012 ; Zahid et al, 2020 ). The lack of a standardized fps for imaging motile cilia is especially problematic because it hinders high‐speed video microscopy from being clinically adapted as a diagnostic tool to help uncover motile cilia diseases, such as primary ciliary dyskinesia (Bricmont et al, 2021 ; Shapiro et al, 2018 ).…”

“…Determination of the optimum sampling rates for cilia video microscopy is also becoming increasingly important with the rapid advancements in digital camera technology (Liang & Wang, 2021; Pauls & Mukasyan, 2017). This has put high‐speed digital imaging within the budget of even the most modest of laboratories, with even consumer‐grade devices (e.g., iPhone) now having the possible frame rates required for cilia imaging (Chen et al, 2016; Mateos‐Quiros et al, 2021). Determining the optimal sampling rate for cilia video microscopy is needed due to the dramatic increase in imaging rates seen in the latest generation of scientific high‐speed video cameras (Etoh et al, 2013), where fps oversampling can greatly increase file sizes needing to be processed and archived (Aiello et al, 2019; Yaffe, 2019).…”

“…The most used method to assess motile cilia function is to quantify cilia beat frequency (CBF) via high‐speed video microscopy (Reula et al, 2021 ; Rubbo et al, 2019 ; Shapiro et al, 2018 ). However, a large heterogeneity exists within the published literature as to what frame rate per second (fps) to use when imaging cilia motility for subsequent CBF calculation; with published frame rates varying between 28–500 fps (Abdelhamed et al, 2018 ; Abdelhamed et al, 2020 ; Bustamante‐Marin et al, 2019 ; Chen et al, 2016 ; Hagiwara et al, 2020 ; Liu et al, 2021 ; Mateos‐Quiros et al, 2021 ; Mikhailik et al, 2021 ; Reula et al, 2021 ; Smith et al, 2012 ; Zahid et al, 2020 ). The large variation in sampling rates suggests some studies may be under‐sampling or oversampling cilia motility, which could result in inaccurate quantification of CBF or generation of large volumes of oversampled data requiring analysis and storage.…”

The Roles of Junctional Adhesion Molecules (JAMs) in Cell Migration

“…JAM-C distributed nonclassically in the apical membranes of Müller cells and retinal pigment epithelial (RPE) ( Daniele et al, 2007 ; Economopoulou et al, 2009 ), and JAM-C knockdown inhibits human RPE cell migration but not proliferation and decreases the permeability of monolayer hRPE ( Hou et al, 2012 ). JAM3 is expressed in multiciliated cells (MCCs) in the airway epithelium, and JAM3 lacking causes a delay in BB assembly/positioning during MCC differentiation ( Mateos-Quiros et al, 2021 ).…”