Testing the ion-current model for flagellar length sensing and IFT regulation

Abstract

Eukaryotic cilia and flagella are microtubule-based organelles whose relatively simple shape makes them ideal for investigating the mechanisms of fundamental size regulation. Most of the flagellar materials are transported from the cell body via an active transport process called intraflagellar transport (IFT) because no protein synthesis occurs in the flagellum. The rate of IFT injection has been shown to negatively correlate with flagellar length. However, it remains unknown how the cell measures the length of its flagella and controls IFT injection. One of the most-discussed theoretical models for length sensing to control IFT is the ion-current model, which posits that there is a uniform distribution of Ca2+ channels along the flagellum and that the Ca2+ current from the flagellum into the cell body increases linearly with flagellar length. In this model, the cell uses the Ca2+ current to negatively regulate IFT injection. The recent discovery that IFT entry into flagella is regulated by a CDPK has provided further impetus for the ion current model. To test this model, we measured and manipulated the levels of Ca2+ inside of Chlamydomonas flagella and quantified IFT injection. Although the concentration of Ca2+ inside of flagella was weakly correlated with the length of flagella, we found that IFT injection was reduced in calcium-deficient flagella, rather than increased as the model predicted, and that variation in IFT injection was uncorrelated with the occurrence of flagellar Ca2+ spikes. Thus, Ca2+ does not work as a negative regulator of IFT injection, hence it cannot form the basis of a stable length control system

Role of intraflagellar transport in transcriptional control during flagellar regeneration in Chlamydomonas

“…Flagellar assembly and length maintenance involves a kinesin-based transport mechanism known as intraflagellar transport or IFT (Kozminski et al, 1993;Cole et al, 1998;Rosenbaum and Witman, 2002;Bhogaraju et al, 2014) that actively transports tubulin and other building blocks to the site of assembly at the tip of the growing flagellum (Qin et al, 2004;Hao et al, 2011;Bhogaraju et al, 2013;Craft et al, 2015). The activity of the IFT pathway is a function of flagellar length, such that the rate of IFT decreases according to 1/L (Engel et al, 2009;Ludington et al, 2013), but the mechanism by which length regulates IFT remains unclear (Ludington et al, 2015;Ishikawa and Marshall, 2017;Hendel et al, 2018;Ishikawa et al, 2022). The transport of tubulin and other cargoes by IFT also varies as a function of length (Wren et al, 2013;Craft et al, 2015), but at least in the case of tubulin it remains unclear whether this represents regulation of a binding interaction or a length dependence of the number of binding sites (Wemmer et al, 2020).…”

Role of intraflagellar transport in transcriptional control during flagellar regeneration in Chlamydomonas

“…Flagellar assembly and length maintenance involves a kinesin-based transport mechanism known as intraflagellar transport or IFT (Kozminski 1993;Cole 1998;Rosenbaum 2002;Bhogaraju 2014 ) that actively transports tubulin and other building blocks to the site of assembly at the tip of the growing flagellum (Qin 2004;Hao 2011;Bhogaraju 2013;Craft 2015). The activity of the IFT pathway is a function of flagellar length, such that the rate of IFT decreases according to 1/L (Engel 2009;Ludington 2013), but the mechanism by which length regulates IFT remains unclear (Ludington 2015;Ishikawa 2017;Hendel 2018;Ishikawa 2022). The transport of tubulin and other cargos by IFT also varies as a function of length (Wren 2013;Craft 2015), but at least in the case of tubulin it remains unclear whether this represents regulation of a binding interaction or a length dependence of the number of binding sites (Wemmer 2020).…”