DYNLT (Tctex-1) forms a tripartite complex with dynein intermediate chain and RagA, hence linking this small GTPase to the dynein motor

FEBS J. 2015 Oct;282(20):3945-58. doi: 10.1111/febs.13388. Epub 2015 Aug 20.

Abstract

It has been suggested that DYNLT, a dynein light chain known to bind to various cellular and viral proteins, can function as a microtubule-cargo adaptor. Recent data showed that DYNLT links the small GTPase Rab3D to microtubules and, for this to occur, the DYNLT homodimer needs to display a binding site for dynein intermediate chain together with a binding site for the small GTPase. We have analysed in detail how RagA, another small GTPase, associates to DYNLT. After narrowing down the binding site of RagA to DYNLT we could identify that a β strand, part of the RagA G3 box involved in nucleotide binding, mediates this association. Interestingly, we show that both microtubule-associated DYNLT and cytoplasmic DYNLT are equally able to bind to the small GTPases Rab3D and RagA. Using NMR spectroscopy, we analysed the binding of dynein intermediate chain and RagA to mammalian DYNLT. Our experiments identify residues of DYNLT affected by dynein intermediate chain binding and residues affected by RagA binding, hence distinguishing the docking site for each of them. In summary, our results shed light on the mechanisms adopted by DYNLT when binding to protein cargoes that become transported alongside microtubules bound to the dynein motor.

Keywords: Tctex-1; dynein motor; dynein-mediated transport; retrograde transport; small GTPases.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Substitution
  • Animals
  • COS Cells
  • Chlorocebus aethiops
  • Cytoplasm / enzymology
  • Cytoplasm / metabolism*
  • Cytoplasmic Dyneins / chemistry
  • Cytoplasmic Dyneins / genetics
  • Cytoplasmic Dyneins / metabolism*
  • Dimerization
  • Dyneins / chemistry
  • Dyneins / genetics
  • Dyneins / metabolism*
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • Humans
  • Mice
  • Microtubules / enzymology
  • Microtubules / metabolism*
  • Models, Molecular*
  • Monomeric GTP-Binding Proteins / chemistry
  • Monomeric GTP-Binding Proteins / genetics
  • Monomeric GTP-Binding Proteins / metabolism*
  • Mutant Proteins / chemistry
  • Mutant Proteins / metabolism
  • Peptide Fragments / chemistry
  • Peptide Fragments / genetics
  • Peptide Fragments / metabolism
  • Protein Interaction Domains and Motifs
  • Protein Transport
  • Rats
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / metabolism
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / metabolism
  • rab3 GTP-Binding Proteins / chemistry
  • rab3 GTP-Binding Proteins / genetics
  • rab3 GTP-Binding Proteins / metabolism*

Substances

  • DYNLT1 protein, human
  • Mutant Proteins
  • Peptide Fragments
  • Recombinant Fusion Proteins
  • Recombinant Proteins
  • Green Fluorescent Proteins
  • RRAGA protein, human
  • Rab3d protein, mouse
  • Cytoplasmic Dyneins
  • Dync1i2 protein, rat
  • Dyneins
  • Monomeric GTP-Binding Proteins
  • rab3 GTP-Binding Proteins