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Proteomic analysis of microtubule inner proteins (MIPs) in Rib72 null Tetrahymena cells reveals functional MIPs

By Amy S. Fabritius, Brian A Bayless, Sam Li, Daniel Stoddard, Westley Heydeck, Christopher C. Ebmeier, Lauren Anderson, Tess Gunnels, Chidambaram Nachiappan, Justen B. Whittall, William Old, David A. Agard, Daniela Nicastro, Mark Winey

Posted 03 Oct 2020
bioRxiv DOI: 10.1101/2020.10.02.324467

Motile cilia and flagella are built from stable populations of doublet microtubules that comprise their axonemes. Their unique stability is brought about, at least in part, by a network of Microtubule Inner Proteins (MIPs) found in the lumen of their doublet microtubules. Rib72A and Rib72B were identified as microtubule inner proteins (MIPs) in the motile cilia of Tetrahymena thermophila . Loss of these proteins leads to ciliary defects and loss of multiple MIPs. We performed mass spectrometry coupled with proteomic analysis and bioinformatics to identify the MIPs lost in RIB72A/B knockout (KO) Tetrahymena cells. From this analysis we identified a number of candidate MIPs and pursued one, Fap115, for functional characterization. We find that loss of Fap115 results in disrupted cell swimming and aberrant ciliary beating. Cryo-electron tomography reveals that Fap115 localizes to MIP6a in the A-tubule of the doublet microtubules. Overall, our results highlight the complex relationship between MIPs, ciliary structure, and ciliary function. ### Competing Interest Statement The authors have declared no competing interest.

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