Dynein-2 intermediate chains play crucial but distinct roles in primary cilia formation and function

Abstract

The dynein-2 microtubule motor is the retrograde motor for intraflagellar transport. Mutations in dynein-2 components cause skeletal ciliopathies, notably Jeune syndrome. Dynein-2 contains a heterodimer of two non-identical intermediate chains, WDR34 and WDR60. Here, we use knockout cell lines to demonstrate that each intermediate chain has a distinct role in cilium function. Using quantitative proteomics, we show that WDR34 KO cells can assemble a dynein-2 motor complex that binds IFT proteins yet fails to extend an axoneme, indicating complex function is stalled. In contrast, WDR60 KO cells do extend axonemes but show reduced assembly of dynein-2 and binding to IFT proteins. Both proteins are required to maintain a functional transition zone and for efficient bidirectional intraflagellar transport. Our results indicate that the subunit asymmetry within the dynein-2 complex is matched with a functional asymmetry between the dynein-2 intermediate chains. Furthermore, this work reveals that loss of function of dynein-2 leads to defects in transition zone architecture, as well as intraflagellar transport.

Cytoplasmic dynein-2 at a glance

“…Although there are several common features of IFT between model organisms, there are also key differences. In Chlamydomonas, kinesin-2 appears to mainly diffuse back to the ciliary base (Engel et al, 2012), whereas, in metazoans, kinesin-2 motors appear to be recycled to the ciliary base predominantly through retrograde IFT (Mijalkovic et al, 2017;Signor et al, 1999;Vuolo et al, 2018;Williams et al, 2014). Interestingly, in C. elegans, an additional dynein heavy chain, i.e.…”

“…WDR34 and WDR60 form a heterodimer (Asante et al, 2014;Hamada et al, 2018;Toropova et al, 2019;Vuolo et al, 2018) (see poster). Their C-terminal β-propeller domains each bind a copy of the heavy chain, and their extended N-terminal regions are held together by an array of light chain dimers (Toropova et al, 2019).…”

Intraflagellar transport trains and motors: Insights from structure

“…It will be interesting to decipher if passage through the transition zone requires a conformational change in the IFT train or Y-link. Recent studies show that mutations in IFT-A and dynein-2 subunits can perturb the localization of transition zone proteins, highlighting a connection between IFT and transition zone integrity [ [85] , [86] , [87] ].…”

“…Interestingly, dynein-2 heavy chain immunoprecipitated with IFT172 migrates differently by SDS-PAGE compared to that from crude extract, suggesting that it could be differentially modified [ 44 ]. Additional IFT-B proteins have been associated with dynein-2 in trypanosomes (IFT22/25/27) [ [139] , [140] , [141] ] and mammalian cells (IFT25/54/57/74/88/172) [ 86 ].…”

Emerging mechanisms of dynein transport in the cytoplasm versus the cilium

“…Dysfunction of IFT-A is typically associated with short cilia with accumulations at their tips, similar to the cellular phenotype of dynein-2 mutants [ 31 ]. The IFT-A proteins IFT139 and IFT140 have been found to co-immmunoprecipitate with dynein-2 in C. reinhardtii [ 75 ] and mammalian cells [ 135 ], respectively. In general, because the IFT-A and IFT-B proteins play multiple roles in IFT train assembly, cargo binding, and ciliogenesis, dissecting their specific contribution to dynein-2 regulation is an ongoing challenge.…”