title: 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 Forster's resonance energy transfer 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-tubulin1 transgene. Furthermore, fluorescent recovery after photobleach 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.
authors.affiliation: Department of Biological Sciences, Tata Institute of Fundamental Research, Mumbai, India.
authors.forename: Mukul
authors.initials: M
authors.lastname: Girotra
authors.affiliation: Department of Biosciences and Biotechnology, Indian Institute of Technology Bombay, Mumbai, India.
authors.forename: Shalini
authors.initials: S
authors.lastname: Srivastava
authors.affiliation: Department of Biological Sciences, Tata Institute of Fundamental Research, Mumbai, India.
authors.forename: Anuttama
authors.initials: A
authors.lastname: Kulkarni
authors.affiliation: Department of Biological Sciences, Tata Institute of Fundamental Research, Mumbai, India.
authors.forename: Ayan
authors.initials: A
authors.lastname: Barbora
authors.affiliation: Department of Biological Sciences, Tata Institute of Fundamental Research, Mumbai, India.
authors.forename: Kratika
authors.initials: K
authors.lastname: Bobra
authors.affiliation: Department of Biological Sciences, Tata Institute of Fundamental Research, Mumbai, India.
authors.forename: Debnath
authors.initials: D
authors.lastname: Ghosal
authors.affiliation: Department of Biological Sciences, Tata Institute of Fundamental Research, Mumbai, India.
authors.forename: Pavithra
authors.initials: P
authors.lastname: Devan
authors.affiliation: Department of Biological Sciences, Tata Institute of Fundamental Research, Mumbai, India.
authors.forename: Amol
authors.initials: A
authors.lastname: Aher
authors.affiliation: Department of Biological Sciences, Tata Institute of Fundamental Research, Mumbai, India.
authors.forename: Akanksha
authors.initials: A
authors.lastname: Jain
authors.affiliation: Department of Biosciences and Biotechnology, Indian Institute of Technology Bombay, Mumbai, India.
authors.forename: Dulal
authors.initials: D
authors.lastname: Panda
authors.affiliation: Department of Biological Sciences, Tata Institute of Fundamental Research, Mumbai, India.
authors.forename: Krishanu
authors.initials: K
authors.lastname: Ray
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport / Q000502:physiology; D002923:Cilia / Q000378:metabolism*; D004330:Drosophila; D005407:Flagella / Q000378:metabolism; D016547:Kinesins; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism; D041681:NIH 3T3 Cells; D011485:Protein Binding / Q000502:physiology; D000072417:Protein Domains / Q000502:physiology; D021122:Protein Subunits / Q000378:metabolism*; D014404:Tubulin / Q000378:metabolism*
chemical_list: D008869:Microtubule-Associated Proteins; D021122:Protein Subunits; D014404:Tubulin; D016547:Kinesins
keywords: 
  Drosophila
  ; Kif3A; Kif3B; antenna; cilia; kinesin-like-protein 64D; kinesin-like-protein 68D; mouse