Abstract
Programs that direct cellular differentiation are dependent on the strict temporal expression of regulatory factors that can be provided by Rho GTPases. Ciliogenesis is a complex sequence of events involving the generation and docking of basal bodies at the apical membrane, followed by ciliary axoneme generation. Although a cilia proteome has been assembled, programs that direct ciliated cell differentiation are not well established, particularly in mammalian systems. Using mouse primary culture airway epithelial cells, we identified a critical stage of ciliogenesis requiring the temporal establishment of an apical web-like structure of actin for basal body docking and subsequent axoneme growth. Apical web formation and basal body docking were prevented by interruption of actin remodeling and were dependent on RhoA activation. Additional evidence for this program was provided by analysis of Foxj1-null mice that failed to dock basal bodies and lacked apical actin. Foxj1 expression coincided with actin web formation, activated RhoA and RhoB, and persisted despite RhoA inhibition, suggesting that Foxj1 promoted RhoA during ciliogenesis. Apical ezrin localization was also dependent on Foxj1, actin remodeling, and RhoA, but was not critical for ciliogenesis. Thus, temporal Foxj1 and RhoA activity are essential regulatory events for cytoskeletal remodeling during mammalian ciliogenesis.
Highlights
Mutations in genes with roles in ciliogenesis have been increasingly implicated in human disease (Ibanez-Tallon et al, 2003)
Using a highly refined model of ciliogenesis developed in primary culture mouse tracheal epithelial cells, we found that RhoA is required for the formation of a characteristic apical actin web in ciliated cells
Actin is enriched at the apical membrane of ciliated cells during differentiation of airway epithelial cells Previous observations that ezrin was selectively expressed in ciliated airway cells, and binds to actin when localized to the apical membrane led us to examine the temporal and spatial localization of actin during epithelial cell differentiation
Summary
Mutations in genes with roles in ciliogenesis have been increasingly implicated in human disease (Ibanez-Tallon et al, 2003). Polycystic kidney disease is caused by mutations in genes expressed in sensory cilia (Nauli et al, 2003), whereas primary ciliary dyskinesia (leading to chronic respiratory infections) is caused by mutations in dynein motor proteins in motile cilia (Olbrich et al, 2002). Environmental factors play roles as evidenced by cilia loss due to respiratory viruses and cigarette smoke, requiring intact mechanisms of ciliogenesis for repair (Ibricevic et al, 2006; Look et al, 2001; Sisson et al, 1994). Together, these disorders suggest a role for cilia in a wide range of physiologic and pathologic processes. Despite the identification of these mutations, relatively little is known about the biochemical pathways for ciliogenesis, in mammalian systems
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