title: DYNLT (Tctex-1) forms a tripartite complex with dynein intermediate chain and RagA, hence linking this small GTPase to the dynein motor.
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.
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Químicas, Universidad Complutense de Madrid, Spain.
authors.forename: Javier
authors.initials: J
authors.lastname: Merino-Gracia
authors.affiliation: Departamento de Química Biológica, Instituto de Química-Física Rocasolano, CSIC, Serrano, Madrid, Spain.
authors.forename: María Flor
authors.initials: MF
authors.lastname: García-Mayoral
authors.affiliation: Dynamics of Cell Growth and Division, Institut de Biologie Cellulaire et de Génétique, Centre National de la Recherche Scientifique, Bordeaux, France.
authors.forename: Peter
authors.initials: P
authors.lastname: Rapali
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Químicas, Universidad Complutense de Madrid, Spain.
authors.forename: Ruth Ana
authors.initials: RA
authors.lastname: Valero
authors.affiliation: Departamento de Química Biológica, Instituto de Química-Física Rocasolano, CSIC, Serrano, Madrid, Spain.
authors.forename: Marta
authors.initials: M
authors.lastname: Bruix
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Químicas, Universidad Complutense de Madrid, Spain.
authors.forename: Ignacio
authors.initials: I
authors.lastname: Rodríguez-Crespo
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D003593:Cytoplasm / Q000201:enzymology / Q000378:metabolism*; D056445:Cytoplasmic Dyneins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D004398:Dyneins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D051379:Mice; D008870:Microtubules / Q000201:enzymology / Q000378:metabolism*; D008958:Models, Molecular; D020559:Monomeric GTP-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D054730:Protein Interaction Domains and Motifs; D021381:Protein Transport; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020692:rab3 GTP-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: C533376:DYNLT1 protein, human; D050505:Mutant Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D049452:Green Fluorescent Proteins; C484342:RRAGA protein, human; C116711:Rab3d protein, mouse; D056445:Cytoplasmic Dyneins; C584751:Dync1i2 protein, rat; D004398:Dyneins; D020559:Monomeric GTP-Binding Proteins; D020692:rab3 GTP-Binding Proteins
keywords: Tctex-1; dynein motor; dynein-mediated transport; retrograde transport; small GTPases