Intraflagellar transport proteins 172, 80, 57, 54, 38,  and 20 form a stable tubulin‐binding IFT‐B2 complex. (24th February 2016)

• Record Type: Journal Article
• Authors: Taschner, Michael; Weber, Kristina; Mourão, André; Vetter, Melanie; Awasthi, Mayanka; Stiegler, Marc; Bhogaraju, Sagar; Lorentzen, Esben
• Is Part Of: EMBO journal. Volume 35:Number 7(2016)
• Page Start: 773
• Page End: 790
• Publication Date: 2016-02-24
• Subjects: cilia -- IFT54 -- IFT‐B -- intraflagellar transport -- tubulin transpor; Molecular biology -- Periodicals; 572.805
• DOI: 10.15252/embj.201593164
• Languages: English
• ISSNs: 0261-4189

Abstract

Abstract: Intraflagellar transport (IFT) relies on the IFT complex and is required for ciliogenesis. The IFT‐B complex consists of 9–10 stably associated core subunits and six "peripheral" subunits that were shown to dissociate from the core structure at moderate salt concentration. We purified the six "peripheral" IFT‐B subunits of Chlamydomonas reinhardtii as recombinant proteins and show that they form a stable complex independently of the IFT‐B core. We suggest a nomenclature of IFT‐B1 (core) and IFT‐B2 (peripheral) for the two IFT‐B subcomplexes. We demonstrate that IFT88, together with the N‐terminal domain of IFT52, is necessary to bridge the interaction between IFT‐B1 and B2. The crystal structure of IFT52N reveals highly conserved residues critical for IFT‐B1/IFT‐B2 complex formation. Furthermore, we show that of the three IFT‐B2 subunits containing a calponin homology (CH) domain (IFT38, 54, and 57), only IFT54 binds αβ‐tubulin as a potential IFT cargo, whereas the CH domains of IFT38 and IFT57 mediate the interaction with IFT80 and IFT172, respectively. Crystal structures of IFT54 CH domains reveal that tubulin binding is mediated by basic surface‐exposed residues. Synopsis: Thought to be peripheral components of the IFT‐B complex, IFT172, 80, 57, 54, and 20 form a stable sub‐complex (IFT‐B2). IFT‐B2 interacts with tubulin/microtubules and connects them to the core IFT‐B1 complex through interaction with IFT88/52. Peripheral IFT‐B complex proteins IFT172, 80, 57, 54, and 20 form a stable IFT‐B2 complex that we reconstitute and purify. IFT‐B2 interacts with the IFT‐B1 (core) complex via IFT88/52 to form the full IFT‐B complex. IFT‐B2 binds tubulin/MT via the N‐terminal calponin homology (CH) domain of IFT54. High‐resolution crystal structures of IFT52N and IFT54N reveal residues critical to IFT‐B complex assembly and tubulin binding. Abstract : Thought to be peripheral components of the IFT‐B complex, IFT172, 80, 57, 54, and 20 form a stable sub‐complex (IFT‐B2). IFT‐B2 interacts with tubulin/microtubules and connects them to the core IFT‐B1 complex through interaction with IFT88/52.