Mechanism of microtubule plus-end tracking by the plant-specific SPR1 protein and its development as a versatile plus-end marker

Abstract

Microtubules are cytoskeletal polymers that perform diverse cellular functions. The plus ends of microtubules promote polymer assembly and disassembly and connect the microtubule tips to other cellular structures. The dynamics and functions of microtubule plus ends are governed by microtubule plus end-tracking proteins (+TIPs). Here we report that the Arabidopsis thaliana SPIRAL1 (SPR1) protein, which regulates directional cell expansion, is an autonomous +TIP. Using in vitro reconstitution experiments and total internal reflection fluorescence microscopy, we demonstrate that the conserved N-terminal region of SPR1 and its GGG motif are necessary for +TIP activity whereas the conserved C-terminal region and its PGGG motif are not. We further show that the N- and C-terminal regions, either separated or when fused in tandem (NC), are sufficient for +TIP activity and do not significantly perturb microtubule plus-end dynamics compared with full-length SPR1. We also found that exogenously expressed SPR1-GFP and NC-GFP label microtubule plus ends in plant and animal cells. These results establish SPR1 as a new type of intrinsic +TIP and reveal the utility of NC-GFP as a versatile microtubule plus-end marker.