Effect of TBCD and its regulatory interactor Arl2 on tubulin and microtubule integrity
- 25 August 2010
- journal article
- research article
- Published by Wiley in Cytoskeleton
- Vol. 67 (11), 706-714
- https://doi.org/10.1002/cm.20480
Abstract
Assembly of the α/β tubulin heterodimer requires the participation of a series of chaperone proteins (TBCA‐E) that function downstream of the cytosolic chaperonin (CCT) as a heterodimer assembly machine. TBCD and TBCE are also capable of acting in a reverse reaction in which they disrupt native heterodimers. Homologs of TBCA‐E exist in all eukaryotes, and the amino acid sequences of α‐ and β‐tubulin isotypes are rigidly conserved among vertebrates. However, the efficiency with which TBCD effects tubulin disruption in vivo depends on its origin: bovine (but not human) TBCD efficiently destroys tubulin and microtubules upon overexpression in cultured cells. Here we show that recombinant bovine TBCD is produced in HeLa cells as a stoichiometric cocomplex with β‐tubulin, consistent with its behavior in vitro and in vivo. In contrast, expression of human TBCD using the same host/vector system results in the generation of TBCD that is not complexed with β‐tubulin. We show that recombinant human TBCD functions indistinguishably from its nonrecombinant bovine counterpart in in vitro CCT‐driven folding reactions, in tubulin disruption reactions, and in tubulin GTPase activating protein assays in which TBCD and TBCC stimulate GTP hydrolysis by β‐tubulin at a heterodimer concentration far below that required for polymerization into microtubules. We conclude that bovine and human TBCD have functionally identical roles in de novo tubulin heterodimer assembly, and show that the inability of human TBCD to disrupt microtubule integrity upon overexpression in vivo can be overcome by siRNA‐mediated suppression of expression of the TBCD regulator Arl2 (ADP ribosylation factor‐like protein).Keywords
This publication has 29 references indexed in Scilit:
- TBCD Links Centriologenesis, Spindle Microtubule Dynamics, and Midbody Abscission in Human CellsPLOS ONE, 2010
- Pathways of chaperone-mediated protein folding in the cytosolNature Reviews Molecular Cell Biology, 2004
- Membrane Targeting: Getting Arl to the GolgiCurrent Biology, 2004
- Cytosolic Arl2 Is Complexed with Cofactor D and Protein Phosphatase 2AOnline Journal of Public Health Informatics, 2003
- Tubulin Folding Cofactors: Half a Dozen for a DimerCurrent Biology, 2002
- The Arabidopsis PILZ group genes encode tubulin-folding cofactor orthologs required for cell division but not cell growthGenes & Development, 2002
- Molecular Chaperones in the Cytosol: from Nascent Chain to Folded ProteinScience, 2002
- Type II chaperonns, prefoldin, and the tubulin-specific chaperonesAdvances in protein chemistry, 2001
- Tubulin folding cofactor D is a microtubule destabilizing proteinFEBS Letters, 2000
- Principles that Govern the Folding of Protein ChainsScience, 1973