Identification and mapping of central pair proteins by proteomic analysis

Cilia or flagella of eukaryotes are small micro-hair like structures that are indispensable to single-cell motility and play an important role in mammalian biological processes. Cilia or flagella are composed of nine doublet microtubules surrounding a pair of singlet microtubules called the central pair (CP). Together, this arrangement forms the canonical and highly conserved 9+2 axonemal structure. The CP, which is a unique structure exclusive to motile cilia, is a pair of structurally dimorphic singlet microtubules decorated with numerous associated proteins. Mutations of CP-associated proteins cause several different physical symptoms termed ciliopathies. Thus, it is crucial to understand the architecture of the CP. However, the protein composition of the CP was poorly understood. This was because identification of CP proteins was mostly limited by available Chlamydomonas mutants of CP proteins. In this study, we conducted a comprehensive CP proteome analysis using several CP mutants and identified 37 novel CP protein candidates. By using Chlamydomonas strains lacking specific CP sub-structures, we also present a more complete model of localization of known and newly identified CP proteins. This work has established a new foundation for CP protein analysis for future studies.