The C ‐terminal tails of heterotrimeric kinesin‐2 motor subunits directly bind to α‐tubulin1: possible implications for cilia‐specific tubulin entry

Abstract
The assembly of microtubule‐based cytoskeleton propels the cilia and flagella growth. Previous studies have indicated that the kinesin‐2 family motors transport tubulin into the cilia through intraflagellar transport. Here, we report a direct interaction between the C‐terminal tail fragments of heterotrimeric kinesin‐2 and α‐tubulin1 isoforms in vitro. Blot overlay screen, affinity purification from tissue extracts, cosedimentation with subtilisin‐treated microtubule and LC‐ESI‐MS/MS characterization of the tail‐fragment‐associated tubulin identified an association between the tail domains and α‐tubulin1A/D isotype. The interaction was confirmed by FRET assay in tissue cultured cells. The overexpression of the recombinant tails in NIH3T3 cells affected the primary cilia growth which was rescued by coexpression of a α‐tubulin‐1 transgene. Furthermore, FRAP analysis in the olfactory cilia of Drosophila indicated that tubulin is transported in a non‐particulate form requiring kinesin‐2. These results provide additional new insight into the mechanisms underlying selective tubulin isoform enrichment in the cilia.

eTOC Synopsis
Tubulin entry into the cilia and flagella is essential for their growth. The article by Girotra et al., suggests that the tail domains of heterotrimeric Kinesin‐2 motor subunits can selectively bind certain α‐tubulin1 isotypes and regulate the entry of tubulin into cilia. The conjecture is supported by extensiv...

Source: Traffic - December 14, 2016 Category: Research Authors: Mukul Girotra, Shalini Srivastava, Anuttama Kulkarni, Ayan Barbora, Kratika Bobra, Debnath Ghosal, Pavithra Devan, Amol Aher, Akanksha Jain, Dulal Panda, Krishanu Ray Tags: Original Article Source Type: research