title: A conserved docking surface on calcineurin mediates interaction with substrates and immunosuppressants.
abstract: The phosphatase calcineurin, a target of the immunosuppressants cyclosporin A and FK506, dephosphorylates NFAT transcription factors to promote immune activation and development of the vascular and nervous systems. NFAT interacts with calcineurin through distinct binding motifs: the PxIxIT and LxVP sites. Although many calcineurin substrates contain PxIxIT motifs, the generality of LxVP-mediated interactions is unclear. We define critical residues in the LxVP motif, and we demonstrate its binding to a hydrophobic pocket at the interface of the two calcineurin subunits. Mutations in this region disrupt binding of mammalian calcineurin to NFATC1 and the interaction of yeast calcineurin with substrates including Rcn1, which contains an LxVP motif. These mutations also interfere with calcineurin-immunosuppressant binding, and an LxVP-based peptide competes with immunosuppressant-immunophilin complexes for binding to calcineurin. These studies suggest that LxVP-type sites are a common feature of calcineurin substrates, and that immunosuppressant-immunophilin complexes inhibit calcineurin by interfering with this mode of substrate recognition.
authors.affiliation: Department of Vascular Biology and Inflammation, Centro Nacional de Investigaciones Cardiovasculares, Madrid, Spain. a.rodriguez@uam.es
authors.forename: Antonio
authors.initials: A
authors.lastname: Rodríguez
authors.forename: Jagoree
authors.initials: J
authors.lastname: Roy
authors.forename: Sara
authors.initials: S
authors.lastname: Martínez-Martínez
authors.forename: María Dolores
authors.initials: MD
authors.lastname: López-Maderuelo
authors.forename: Perla
authors.initials: P
authors.lastname: Niño-Moreno
authors.forename: Leticia
authors.initials: L
authors.lastname: Ortí
authors.forename: David
authors.initials: D
authors.lastname: Pantoja-Uceda
authors.forename: Antonio
authors.initials: A
authors.lastname: Pineda-Lucena
authors.forename: Martha S
authors.initials: MS
authors.lastname: Cyert
authors.forename: Juan Miguel
authors.initials: JM
authors.lastname: Redondo
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D019703:Calcineurin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D065095:Calcineurin Inhibitors; D003001:Cloning, Molecular; D003198:Computer Simulation; D017124:Conserved Sequence; D017930:Genes, Reporter; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D020104:Immunophilins / Q000378:metabolism; D007166:Immunosuppressive Agents / Q000378:metabolism* / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D019169:Jurkat Cells; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D050778:NFATC Transcription Factors / Q000378:metabolism; D010455:Peptides / Q000378:metabolism; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D015398:Signal Transduction; D013499:Surface Properties; D022061:Tacrolimus Binding Protein 1A / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection
chemical_list: D065095:Calcineurin Inhibitors; D007166:Immunosuppressive Agents; D047908:Intracellular Signaling Peptides and Proteins; D050778:NFATC Transcription Factors; D010455:Peptides; C486302:RCN1 protein, S cerevisiae; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; C070393:CNA1 protein, S cerevisiae; D019703:Calcineurin; C525673:PPP3CA protein, human; C525676:PPP3R1 protein, human; D022061:Tacrolimus Binding Protein 1A; D020104:Immunophilins