Interactions between TULP3 tubby domain and ARL13B amphipathic helix promote lipidated protein transport to cilia

The primary cilium is a nexus for cell signaling and relies on specific protein traf-ficking for function. The tubby family protein TULP3 transports integral membrane proteins into cilia through interactions with the intraflagellar transport complex-A (IFT-A) and phos-phoinositides. It was previously shown that short motifs called ciliary localization sequences (CLSs) are necessary and sufficient for TULP3-dependent ciliary trafficking of transmembrane cargoes. However, the mechanisms by which TULP3 regulates ciliary compartmentalization of nonintegral, membrane-associated proteins and whether such trafficking requires TULP3-dependent CLSs is unknown. Here we show that TULP3 is required for ciliary transport of the Joubert syndrome-linked palmitoylated GTPase ARL13B through a CLS. An N-terminal am-phipathic helix, preceding the GTPase domain of ARL13B, couples with the TULP3 tubby domain for ciliary trafficking, irrespective of palmitoylation. ARL13B transport requires TULP3 binding to IFT-A but not to phosphoinositides, indicating strong membrane-proximate inter-actions, unlike transmembrane cargo transport requiring both properties of TULP3. TULP3-mediated trafficking of ARL13B also regulates ciliary enrichment of farnesylated and myris-toylated downstream effectors of ARL13B. The lipidated cargoes show distinctive depletion kinetics from kidney epithelial cilia with relation to Tulp3 deletion-induced renal cystogene-sis. Overall, these findings indicate an expanded role of the tubby domain in capturing analo-gous helical secondary structural motifs from diverse cargoes.

Automated summary

TULP3 protein plays a key role in transporting integral membrane proteins into cilia by interacting with the IFT-A and phosphoinositides. It is also involved in the ciliary transport of the Joubert syndrome-linked palmitoylated GTPase ARL13B, and regulates the ciliary enrichment of downstream effectors of ARL13B.