Dynein and dynactin leverage their bivalent character to form a high-affinity interaction.

Authors

Amanda E Siglin, Thomas Jefferson UniversityFollow
Shangjin Sun, University of Delaware, Newark, Delaware
Jeffrey K Moore, Washington University in Saint Louis, Saint Louis, Missouri
Sarah Tan, University of Texas, Austin, Texas
Martin Poenie, University of Texas, Austin, Texas
James D Lear, University of Pennsylvania, Philadelphia, Pennsylvania
Tatyana Polenova, University of Delaware, Newark, Delaware
John A Cooper, Washington University in Saint Louis, Saint Louis, MissouriFollow
John C Williams, Thomas Jefferson UniversityFollow

Document Type

Article

Publication Date

1-1-2013

Comments

This article has been peer reviewed. It was published in: PLoS ONE

Volume 8, Issue 4, 2013, Pages e59453.

The published version is available at DOI: 10.1371/journal.pone.0059453. Copyright © PLoS ONE

Abstract

Cytoplasmic dynein and dynactin participate in retrograde transport of organelles, checkpoint signaling and cell division. The principal subunits that mediate this interaction are the dynein intermediate chain (IC) and the dynactin p150(Glued); however, the interface and mechanism that regulates this interaction remains poorly defined. Herein, we use multiple methods to show the N-terminus of mammalian dynein IC, residues 10-44, is sufficient for binding p150(Glued). Consistent with this mapping, monoclonal antibodies that antagonize the dynein-dynactin interaction also bind to this region of the IC. Furthermore, double and triple alanine point mutations spanning residues 6 to 19 in the yeast IC homolog, Pac11, produce significant defects in spindle positioning. Using the same methods we show residues 381 to 530 of p150(Glued) form a minimal fragment that binds to the dynein IC. Sedimentation equilibrium experiments indicate that these individual fragments are predominantly monomeric, but admixtures of the IC and p150(Glued) fragments produce a 2:2 complex. This tetrameric complex is sensitive to salt, temperature and pH, suggesting that the binding is dominated by electrostatic interactions. Finally, circular dichroism (CD) experiments indicate that the N-terminus of the IC is disordered and becomes ordered upon binding p150(Glued). Taken together, the data indicate that the dynein-dynactin interaction proceeds through a disorder-to-order transition, leveraging its bivalent-bivalent character to form a high affinity, but readily reversible interaction.

Recommended Citation

Siglin, Amanda E; Sun, Shangjin; Moore, Jeffrey K; Tan, Sarah; Poenie, Martin; Lear, James D; Polenova, Tatyana; Cooper, John A; and Williams, John C, "Dynein and dynactin leverage their bivalent character to form a high-affinity interaction." (2013). Department of Biochemistry and Molecular Biology Faculty Papers. Paper 52.
https://jdc.jefferson.edu/bmpfp/52

PubMed ID

23577064