Primary ciliary dyskinesia (PCD) is a genetic disorder in which impaired ciliary function leads to chronic airway disease. Exome sequencing of a PCD subject identified an apparent homozygous frameshift variant, c.887_890delTAAG (p.Val296Glyfs*13), in exon 5; this frameshift introduces a stop codon in amino acid 308 of the growth arrest-specific protein 2-like 2 (GAS2L2). Further genetic screening of unrelated PCD subjects identified a second proband with a compound heterozygous variant carrying the identical frameshift variant and a large deletion (c.867_*343þ1207del; p.?) starting in exon 5. Both individuals had clinical features of PCD but normal ciliary axoneme structure. In this research, using human nasal cells, mouse models, and X.laevis embryos, we show that GAS2L2 is abundant at the apical surface of ciliated cells, where it localizes with basal bodies, basal feet, rootlets, and actin filaments. Cultured GAS2L2deficient nasal epithelial cells from one of the affected individuals showed defects in ciliary orientation and had an asynchronous and hyperkinetic (GAS2L2-deficient ¼ 19.8 Hz versus control ¼ 15.8 Hz) ciliary-beat pattern. These results were recapitulated in Gas2l2 À/À mouse tracheal epithelial cell (mTEC) cultures and in X. laevis embryos treated with Gas2l2 morpholinos. In mice, the absence of Gas2l2 caused neonatal death, and the conditional deletion of Gas2l2 impaired mucociliary clearance (MCC) and led to mucus accumulation. These results show that a pathogenic variant in GAS2L2 causes a genetic defect in ciliary orientation and impairs MCC and results in PCD.
ulticiliated cells (MCCs) contain tens of motile cilia that beat to drive fluid flow across epithelial surfaces. Multiciliated cells are present in the respiratory tract, brain ventricles and reproductive systems. Defects in motile-cilia formation or beating lead to the development of hydrocephaly, lethal respiratory symptoms and fertility defects 1-4. A centriole, or basal body, serves as a template for the cilium axoneme. Centriole duplication is tightly controlled in cycling cells so that a single new procentriole forms adjacent to each of the two parent centrioles 5. However, MCC progenitors with two parent centrioles produce tens to hundreds of additional new centrioles to nucleate multiple motile cilia 1. Steric constraints imposed by the 'centriolar' pathway seem to restrict the number of procentrioles that can be nucleated by the parent centrioles. Centriole amplification is therefore thought to rely on the assembly of dozens of MCCspecific organelles called deuterosomes, which each nucleate tens of procentrioles 6-14. Deuterosomes are assembled during centriole amplification and support the growth of approximately 90% of the procentrioles formed in mammalian MCCs 12,14. Deuterosomes have been proposed to be nucleated from the younger parent centriole 12 but can form spontaneously in a cloud of pericentriolar material (PCM) in MCCs depleted of the parent centrioles 15-17. Many of the proteins required for centriole formation in MCCs are common to centriole duplication 11-15,18-23. However, DEUP1 (CCDC67, alternate gene name) has been identified as a deuterosome-specific protein that arose from a gene-duplication event of the centriolar gene Cep63. Recent data suggest that Deup1 evolved to enable the formation of deuterosomes and the generation of large numbers of centrioles 14. In this manuscript, we interrogate the function of the deuterosome in MCCs from mouse and from Xenopus laevis. Surprisingly, our findings reveal that deuterosomes are dispensable for centriole amplification and multiciliogenesis both in vitro and in vivo. Moreover, we show that neither deuterosomes nor parent centrioles are required for MCCs to amplify the correct number of centrioles. These findings raise new questions about the evolutionary role of deuterosome during multiciliogenesis and the mechanisms regulating centriole number in MCCs. Results Generation of a Deup1-knockout mouse. To examine the role of the deuterosome in multiciliogenesis we created a Deup1-knockout mouse by replacing a region from within exon 2 to within exon 7 of the Deup1 gene with a LacZ reporter (Extended Data Fig. 1a). Reverse Transcription-quantitative PCR on brain and testes samples showed that the messenger RNA levels of Deup1 were reduced by at least tenfold in Deup1-knockout compared with control mice (Extended Data Fig. 1b,c). To examine the process of multiciliogenesis in Deup1 −/− cells, we utilized in vitro cultures of mouse tracheal epithelial cells (mTECs) or ependymal cells 24,25. Consistent with the absence of Deup1 mRNA, DEUP1 foci were a...
Kim et al. show that CLAMP regulates planar cell polarity (PCP) signaling. Its depletion causes a loss of the atypical cadherin Celsr2, a loss of PCP protein asymmetry, and a defect in cilia polarity and oriented cell division. CLAMP also, via its role in PCP, regulates the accumulation of an asymmetric pool of microtubules.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.