Planar Cell Polarity Effector Proteins Inturned and Fuzzy Form a Rab23 GEF Complex.

Andreas Gerondopoulos,Nicola L Stevenson,Tim P Levine,Helen Strutt,Tomoaki Sobajima,Francis A Barr,David Strutt,David J Stephens

doi:10.1016/j.cub.2019.07.090

Andreas Gerondopoulos, Nicola L Stevenson + Show 6 more

Open Access

https://doi.org/10.1016/j.cub.2019.07.090

Copy DOI

Abstract

SummaryA subset of Rab GTPases have been implicated in cilium formation in cultured mammalian cells [1, 2, 3, 4, 5, 6]. Rab11 and Rab8, together with their GDP-GTP exchange factors (GEFs), TRAPP-II and Rabin8, promote recruitment of the ciliary vesicle to the mother centriole and its subsequent maturation, docking, and fusion with the cell surface [2, 3, 4, 5]. Rab23 has been linked to cilium formation and membrane trafficking at mature cilia [1, 7, 8]; however, the identity of the GEF pathway activating Rab23, a member of the Rab7 subfamily of Rabs, remains unclear. Longin-domain-containing complexes have been shown to act as GEFs for Rab7 subfamily GTPases [9, 10, 11, 12]. Here, we show that Inturned and Fuzzy, proteins previously implicated as planar cell polarity (PCP) effectors and in developmentally regulated cilium formation [13, 14], contain multiple longin domains characteristic of the Mon1-Ccz1 family of Rab7 GEFs and form a specific Rab23 GEF complex. In flies, loss of Rab23 function gave rise to defects in planar-polarized trichome formation consistent with this biochemical relationship. In cultured human and mouse cells, Inturned and Fuzzy localized to the basal body and proximal region of cilia, and cilium formation was compromised by depletion of either Inturned or Fuzzy. Cilium formation arrested after docking of the ciliary vesicle to the mother centriole but prior to axoneme elongation and fusion of the ciliary vesicle and plasma membrane. These findings extend the family of longin domain GEFs and define a molecular activity linking Rab23-regulated membrane traffic to cilia and planar cell polarity.

Highlights

This pattern of homology indicates that Intu-Fuz are likely to form a functional pair, equivalent to Mon1-Ccz1 and Hps1Hps4
Identification of the Rab23 GTP exchange factors (GEFs) Complex Phylogenetic analysis shows that Rab23 is a member of the Rab7/32/38 family of Rabs acting in lysosome and lysosomerelated organelle trafficking [15]
Rab7 family GTPases are activated by a conserved group of two-subunit GEF complexes, where each subunit is characterized by a single copy of a common a-b-a sandwich fold, the longin domain that is close to the N terminus [9,10,11,12]

Summary

Introduction

This pattern of homology indicates that Intu-Fuz are likely to form a functional pair, equivalent to Mon1-Ccz and Hps1Hps. Interaction mapping revealed that Intu and Fuz form a binary complex (Figures 1C and S1C). In part, this is likely to be mediated by the multiple longin domains in both proteins (Figures S1A and S1B). By analogy with the other longin domain containing Rab GEFs [11, 22], we propose that the multiple longin domains are likely to form sequential pairwise interaction

Methods

Results

Discussion

Conclusion