Flagellar-associated Protein FAP85 Is a Microtubule Inner Protein That Stabilizes Microtubules

Kirima, Junya; Oiwa, Kazuhiro

doi:10.1247/csf.17023

Cited by 32 publications

(26 citation statements)

References 40 publications

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“…This means that the degree of inherent tension inside the doublet can be changed by external cues. There are a few MIP candidates with calcium binding domain, such as Rib72a (11) and FAP85 (21). Thus, MIPs can exploit the tubulin conformational change as a tool to modify the rigidity of the doublet and, thus, ciliary bending (Fig.…”

Section: Resultsmentioning

confidence: 99%

Tubulin Lattice in Cilia is in a Stressed Form Regulated by Microtubule Inner Proteins

Ichikawa

Khalifa

Basu

et al. 2019

Preprint

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17Cilia, the hair-like protrusions that beat at high frequencies to propel a cell or move fluid around 18 the cell, are composed of radially bundled doublet microtubules. The doublet microtubule is 19 composed of a 13-protofilament A-tubule, a partial 10-protofilament B-tubule and microtubule 20 inner proteins (MIPs) inside the tubulin lattice. In this study, we present the near-atomic 21 resolution map of the Tetrahymena doublet microtubules. The map demonstrates that the 22 network of microtubule inner proteins is weaving into the tubulin lattice, forming an inner sheath 23 of proteins. In addition, we also obtain the tubulin lattice structure with missing MIPs by Sarkosyl 24 treatment. In this structure, the tubulin lattice showed significant longitudinal compaction and 25 lateral angle changes between protofilaments. These results are evidence that the binding of 26MIPs directly affects and stabilizes the tubulin lattice. It is also suggested that the doublet 27 microtubule is an intrinsically stressed filament and this stress could be exploited in the 28 regulation of ciliary waveforms. 29 30 rigidity, damage resistance and stability similar to what microtubule-associated proteins affects 52 the properties of singlets (10). 53Recently, Rib72a/b were characterized as components of MIPs inside the A-tubule of 54Tetrahymena (11) while FAP45 and FAP52 were identified as MIPs in the B-tubule in 55Chlamydomonas (10). The Rib72a/b knockout caused reduced Tetrahymena swimming speed. 56The B-tubule of Chlamydomonas FAP45 and FAP52 double knockout mutant were more 57 vulnerable to depolymerization. These studies highlight the global effects of MIPs on motility and 58 stability of cilia. The resolution of the doublet structures presented in these studies, however, 59 was insufficient to uncover how these proteins affect the tubulin lattice. In this study, we 60 obtained near-atomic resolution maps of the doublet and the fractionated A-tubule from 61Tetrahymena cilia to understand the influence of MIPs to the tubulin lattice. 62

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Section: Resultsmentioning

confidence: 99%

Tubulin Lattice in Cilia is in a Stressed Form Regulated by Microtubule Inner Proteins

Ichikawa

Khalifa

Basu

et al. 2019

Preprint

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“…This is nearly twice of the sum of RIB72A and RIB72B (133 kDa), suggesting that either multiple copies of each RIB72 proteins are involved, or that additional, unidentified proteins contribute to these MIP-supra-complexes. A recent biochemical and immuno-gold labeling study suggested that the EF-hand containing protein FAP85 is an A-tubule MIP and may be associated with the MIP1 structure in Chlamydomonas flagella (Kirima and Oiwa, 2017).…”

Section: Rib72a and Rib72b Are Conserved Mips Required For The Assembmentioning

confidence: 99%

“…Whereas singlet cytoplasmic microtubules are usually characterized by dynamic instability, doublet and triplet microtubules form a highly stable scaffold, which serves as a platform for a plethora of associated proteins. A common structural feature shared by doublets, triplets, and other highly stable microtubules, such as the subpellicular and ventral disc microtubules of the pathogens Plasmodium and Giardia, respectively, is the presence of a unique class of microtubule associated proteins, called Microtubule Inner Proteins (MIPs) that are regularly distributed along the luminal surface of the microtubules (Nicastro et al, 2006;Nicastro et al, 2011;Maheshwari et al, 2015;Ichikawa et al, 2017;Kirima and Oiwa, 2017). The 3D structures and periodicities of MIPs have been best described in doublet microtubules (DMTs) of motile cilia where they assemble into a complex arrangement of at least 6 conserved MIP structures (Nicastro et al, 2006;Nicastro et al, 2011;Ichikawa et al, 2017) (Fig.…”

Section: Introductionmentioning

confidence: 99%

TetrahymenaRIB72A and RIB72B are Microtubule Inner Proteins in the ciliary doublet microtubules

Stoddard

Zhao

Bayless

et al. 2018

Preprint

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SUMMARYMicrotubule Inner Proteins (MIPs) bind to the luminal surface of highly stable microtubules. Combining cell biology and cryo-electron tomography, Stoddard et al. show that RIB72A and RIB72B are conserved MIPs in ciliary doublet microtubules and that they are important for proper ciliary motility. ABSTRACTDoublet and triplet microtubules are essential and highly stable core structures of centrioles, basal bodies, cilia and flagella. In contrast to dynamic cytoplasmic microtubules, their luminal surface is coated with regularly arranged Microtubule Inner Proteins (MIPs). However, the protein composition and biological function(s) of MIPs remain poorly understood. Using genetic, biochemical and imaging techniques we identified Tetrahymena RIB72A and RIB72B proteins as ciliary MIPs. Fluorescence imaging of tagged RIB72A and RIB72B showed that both proteins co-localize to Tetrahymena cilia and basal bodies, but assemble independently. Cryoelectron tomography of RIB72A and/or RIB72B knockout strains revealed major structural defects in the ciliary A-tubule involving MIP1, MIP4 and MIP6 structures. The defects of individual mutants were complementary in the double mutant. All mutants had reduced swimming speed and ciliary beat frequencies, and high-speed video imaging revealed abnormal highly curved cilia during power stroke. Our results show that RIB72A and RIB72B are crucial for the structural assembly of ciliary A-tubule MIPs and are important for proper ciliary motility.All rights reserved. No reuse allowed without permission.

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“…1a) 7,21 . To date, two MIPs in the A-tubule have been identified by recent studies 22,23 . MIPs have also been observed in the axonemes of higher organisms 7,21,24–26 , implying that these inner structures are essential for the integrity of DMTs in motile cilia and flagella.…”

Section: Introductionmentioning

confidence: 99%

Inner lumen proteins stabilize doublet microtubules in cilia and flagella

et al. 2019

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Motile cilia are microtubule-based organelles that play important roles in most eukaryotes. Although axonemal microtubules are sufficiently stable to withstand their beating motion, it remains unknown how they are stabilized while serving as tracks for axonemal dyneins. To address this question, we have identified two uncharacterized proteins, FAP45 and FAP52, as microtubule inner proteins (MIPs) in Chlamydomonas . These proteins are conserved among eukaryotes with motile cilia. Using cryo-electron tomography (cryo-ET) and high-speed atomic force microscopy (HS-AFM), we show that lack of these proteins leads to a loss of inner protrusions in B-tubules and less stable microtubules. These protrusions are located near the inner junctions of doublet microtubules and lack of both FAP52 and a known inner junction protein FAP20 results in detachment of the B-tubule from the A-tubule, as well as flagellar shortening. These results demonstrate that FAP45 and FAP52 bind to the inside of microtubules and stabilize ciliary axonemes.

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Flagellar-associated Protein FAP85 Is a Microtubule Inner Protein That Stabilizes Microtubules

Abstract: ABSTRACT. Genomics and proteomics studies in

Cited by 32 publications

References 40 publications

Tubulin Lattice in Cilia is in a Stressed Form Regulated by Microtubule Inner Proteins

Tubulin Lattice in Cilia is in a Stressed Form Regulated by Microtubule Inner Proteins

TetrahymenaRIB72A and RIB72B are Microtubule Inner Proteins in the ciliary doublet microtubules

Inner lumen proteins stabilize doublet microtubules in cilia and flagella

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