Getting tubulin to the tip of the cilium: One IFT train, many different tubulin cargo‐binding sites?

Abstract

Cilia are microtubule-based hair-like structures that project from the surfaces of eukaryotic cells. Cilium formation relies on intraflagellar transport (IFT) to move ciliary proteins such as tubulin from the site of synthesis in the cell body to the site of function in the cilium. A large protein complex (the IFT complex) is believed to mediate interactions between cargoes and the molecular motors that walk along axonemal microtubules between the ciliary base and tip. A recent study using purified IFT complexes has identified a tubulin-binding module in the two core IFT proteins IFT74 and IFT81 that likely serves to bind and transport tubulin within cilia. Here, we calculate the amount of tubulin required to support the observed cilium assembly kinetics and explore the possibility of multiple tubulin binding sites within the IFT complex.

Intraflagellar transport proteins 172, 80, 57, 54, 38, and 20 form a stable tubulin‐binding IFT‐B2 complex

“…We recently showed that the CH domain of IFT81 together with a basic domain of IFT74 (both subunits of the IFT‐B1 complex) forms a tubulin‐binding module that likely functions in the IFT of αβ‐tubulin to the tips of cilia (Bhogaraju et al , 2013a). However, calculations using the kinetics of Chlamydomonas ciliogenesis suggest that each IFT complex likely carries at least two αβ‐tubulin heterodimers to sustain the observed fast initial rate of cilium growth (Bhogaraju et al , 2014). Prime candidates for additional tubulin cargo‐binding sites within the IFT complex are the CH domains of the IFT38, IFT54, and IFT57 subunits of the newly identified IFT‐B2 complex (Taschner et al , 2012; Schou et al , 2013).…”

“…In vitro studies suggested that tubulin cargo directly binds to the IFT complex via a dedicated module consisting of a calponin homology (CH) domain of IFT81 and a highly basic domain of IFT74 (Bhogaraju et al , 2013a). However, given that tubulin is the most abundant ciliary protein, it has been suggested that additional tubulin‐binding sites may exist within the IFT complex (Bhogaraju et al , 2014). Other axonemal components known to travel by IFT in Chlamydomonas are the nexin–dynein regulatory complex proteins DRC2 and DRC4 and the central pair protein PF16 (Wren et al , 2013).…”

Together, the IFT81 and IFT74 N-termini form the main module for intraflagellar transport of tubulin

“…Within IFT-B, there are three proteins (IFT57, IFT54 and IFT38) in addition to IFT81 that have calponin-homology domains and could interact with tubulin or actin (Bhogaraju et al, 2014). Recent in-vitro-binding studies, in which bacterially expressed Chlamydomonas proteins were used, indicate that IFT38 and IFT57 do not bind tubulin but IFT54 binds soluble tubulin with a K d within the low µM range, similar to that of IFT81-IFT74 (Taschner et al, 2016).…”

The Intraflagellar Transport Machinery

“…Second, Drosophila melanogaster does not contain homologs of these proteins (Cole and Snell 2009), but nevertheless assembles sensory cilia by IFT (e.g., Sarpal et al 2003). Last, the presence of only one dedicated tubulin-binding site in the IFT complex is in theory not sufficient to explain the fast initial kinetics of ciliogenesis in Chlamydomonas after deflagellation (Bhogaraju et al 2014). Interestingly, three other IFT proteins within the IFT-B2 complex (IFT57, IFT54, and IFT38) contain amino-terminal CH domains just like IFT81 (Taschner et al 2012;Schou et al 2013), but two of those (IFT57 and IFT38) are engaged in intra-IFT interactions with IFT172 and IFT80 and do not bind tubulin (see Fig.…”

“…About 350,000 tubulin dimers are required to construct a full-length flagellum in Chlamydomonas (Bhogaraju et al 2014), and this process takes 90 min to be complete (Rosenbaum et al 1969). Tubulin was shown to move along the axoneme with IFT particles in C. reinhardtii (Craft et al 2015) and C. elegans (Hao et al 2011), and a first insight into how it is recognized as a cargo came from studies on the amino-terminal domains of IFT81 and IFT74.…”