View Program - 63rd Annual Drosophila Research Conference

Centrosomes are a key microtubule organizing center in the cell. They comprise a pair of centrioles surrounded by a matrix of proteins termed the pericentriolar material. Through microtubule (MT) nucleation centrosomes organize the mitotic spindle, cilia and flagella. To fulfill these functions, centrosomes must be motile to achieve proper positioning within the cell. Very little is understood about the different mechanisms of centrosome motility. Typically, motility is thought to be governed by the activity of MT motors, pushing or pulling on the microtubules anchored at the centrosome. In some cell types, centrioles lack PCM and microtubules, and are referred to as inactive centrioles. Inactive centrioles must be motile as their positioning is critical for asymmetric cell division. Despite this, the mechanisms of inactive centriole movement are not well understood. We investigated how inactive centrioles move in interphase cells. High resolution live imaging in Drosophila revealed that centrioles are microtubule cargo and move along the MT network in a manner involving Kinesin-1. Importantly, super resolution imaging demonstrated that Kinesin-1 localizes to the outside of the centriole in interphase cells. An RNAi screen identified Pericentrin-Like-Protein (Plp) as essential for interphase centriole movement. Through yeast-2-hybrid and an in vivo interaction assay we found that Plp interacts with the cargo binding region of the Kinesin-1 heavy chain. In vitro analysis showed that Plp and Kinesin-1 comigrate on MTs. Furthermore, autoinhibition of Kinesin-1 inhibits interaction with PLP. Relieving Kinesin-1 autoinhibition using hinge region mutations promotes robust PLP binding. Using random mutagenesis, we generated a series of mutations in Plp which ablate interaction with Kinesin-1, and significantly perturbs centriolar transport. Finally we show that centriole transport is essential for correct centrosome activity and inheritance in asymmetrically dividing neural stem cells. Our data support a model where Plp acts as a novel motor adaptor that links the centriole to the MT transport machinery, facilitating movement. In this work we propose the first detailed mechanism of how centrioles can move independently of their role as an MTOC. We will further discuss our recent in vitro and in vivo efforts to dissect the mechanism of Kinesin-1 activation by kinesin activators, which promote PLP binding and efficient centriole motility.

Pericentrin-Like-Protein is a Kinesin-1 Adaptor that drives Centriole Motility.