Foxj1 regulates basal body anchoring to the cytoskeleton of ciliated pulmonary epithelial cells

Abstract

The forkhead box transcription factor Foxj1 is required for cilia formation and left-right axis determination. To define the role of Foxj1 in ciliogenesis, microarray analysis was performed to identify differentially expressed genes in the pulmonary epithelium of foxj1 +/+ and foxj1 -/-mice. In the absence of Foxj1, the expression of calpastatin, an inhibitor of the protease calpain, decreased. RNase protection confirmed the decrease in calpastatin expression and decreased calpastatin was detected in the proximal pulmonary epithelium of foxj1 -/-mice by immunohistochemistry. No change was detected in the expression of calpain 2 in the pulmonary epithelium by western blot or immunohistochemistry. By western blot and immunofluorescence, ezrin, a substrate for calpain, was also found to decrease in the pulmonary epithelium of foxj1 -/-mice. No change in ezrin gene expression was found by RT-PCR. A decrease in ezrin binding phosphoprotein-50 (EBP-50) was also detected by immunofluorescence in the foxj1 -/-mouse pulmonary epithelium. Immunoelectron microscopy demonstrated ezrin associated with the basal bodies of cilia in the pulmonary epithelium. Treatment of tracheal explants from foxj1 -/-mice with a calpain inhibitor resulted in a partial reappearance of cilia observed in these mice. Additionally, following treatment of foxj1 -/-tracheal explants with calpain inhibitor, basal bodies were observed in an apical location along with relocalization of ezrin and EBP-50. Regulation of calpain activity by calpastatin thus provides a mechanism for regulating the anchoring of basal bodies to the apical cytoskeleton in ciliated cells. In the absence of Foxj1, decreased calpastatin expression with decreased ezrin and EBP-50 results in an inability of basal bodies to anchor to the apical cytoskeleton and subsequent failure of axonemal formation.

Lack of cadherins Celsr2 and Celsr3 impairs ependymal ciliogenesis, leading to fatal hydrocephalus

“…Studies of ciliogenesis in the quail oviduct showed that localization of basal bodies to the apical cell domain depends on actin dynamics 35 . In line with this, ciliogenesis is defective in Foxj1 mutant mice, due to abnormal basal body anchoring to the actin-based apical cytoskeleton, and failure of RhoA-mediated apical actin enrichment [44][45][46][47] . The hypothesis that PCP signaling regulates apical actin network formation required for docking of basal bodies and normal ciliogenesis predicts that mutations in other genes that control PCP in mice should similarly lead to cilia dysfunction and hydrocephalus.…”

Generation of multiciliated cells in functional airway epithelia from human induced pluripotent stem cells

“…This mucociliary clearance protects the airway from infection. Forkhead box protein J1 (FOXJ1) is a transcription factor associated with the activation of genes necessary for the anchoring of basal bodies at the apical surface and initiating ciliogenesis (41,42). In our differentiation protocol, FOXJ1 gene expression is highly up-regulated after switching to ALI at day 17 following initiation of differentiation (Fig.…”

Lung development and repair: Contribution of the ciliated lineage

“…Evidence suggests that Foxj1 functions in airway cells during late stages of ciliogenesis and is not sufficient to drive cilia formation in nonciliated adult epithelial cells (27,28). Therefore, FOXJ1-GFP expression in cells without cilia most likely marks embryonic cells already committed to the ciliated lineage.…”

“…Both strains use a promoter region of FOXJ1, a gene that encodes a forkhead domain transcription factor required for late stages of ciliogenesis (27,28). Our analysis of FOXJ1-GFP embryonic lungs provides a unique overview of the appearance of ciliated cells during development.…”