FKBP12 modulation of the binding of the skeletal ryanodine receptor onto the II-III loop of the dihydropyridine receptor.
In skeletal muscle, excitation-contraction coupling involves a functional interaction between the ryanodine receptor (RyR) and the dihydropyridine receptor (DHPR). The domain corresponding to Thr(671)-Leu(690) of the II-III loop of the skeletal DHPR alpha(1)-subunit is able to regulate RyR properties and calcium release from sarcoplasmic reticulum, whereas the domain corresponding to Glu(724)-Pro(760) antagonizes this effect. Two peptides, covering these sequences (peptide A(Sk) and C(Sk), respectively) were immobilized on polystyrene beads. We demonstrate that peptide A(Sk) binds to the skeletal isoform of RyR (RyR1) whereas peptide C(Sk) does not. Using surface plasmon resonance detection, we show that 1) domain Thr(671)-Leu(690) is the only sequence of the II-III loop binding with RyR1 and 2) the interaction of peptide A(Sk) with RyR1 is not modulated by Ca(2+) (pCa 9-2) nor by Mg(2+) (up to 10 mM). In contrast, this interaction is strongly potentiated by the immunophilin FKBP12 (EC(50) = 10 nM) and inhibited by both rapamycin (IC(50) = 5 nM) and FK506. Peptide A(Sk) induces a 300% increase of the opening probability of the RyR1 incorporated in lipid bilayer. Removal of FKBP12 from RyR1 completely abolishes this effect of domain A(Sk) on RyR1 channel behavior. These results demonstrate a direct interaction of the RyR1 with the discrete domain of skeletal DHPR alpha(1)-subunit corresponding to Thr(671)-Leu(690) and show that the association of FKBP12 with RyR1 specifically modulates this interaction.
O'Reilly
Fiona M
FM
Laboratoire Canaux Ioniques et Signalisation, Département de Biologie Moléculaire et Structurale, CEA-Grenoble, F-38054 Grenoble, France.
Robert
Mylène
M
Jona
Istvan
I
Szegedi
Csaba
C
Albrieux
Mireille
M
Geib
Sandrine
S
De Waard
Michel
M
Villaz
Michel
M
Ronjat
Michel
M
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D020746:Calcium Channels, L-Type / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000502:physiology*; D010446:Peptide Fragments / Q000737:chemistry / Q000494:pharmacology; D020033:Protein Isoforms / Q000737:chemistry / Q000378:metabolism; D021122:Protein Subunits; D011817:Rabbits; D020239:Receptor Cross-Talk; D019837:Ryanodine Receptor Calcium Release Channel / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000502:physiology; D022061:Tacrolimus Binding Protein 1A / Q000378:metabolism*
D020746:Calcium Channels, L-Type; D010446:Peptide Fragments; D020033:Protein Isoforms; D021122:Protein Subunits; D019837:Ryanodine Receptor Calcium Release Channel; D022061:Tacrolimus Binding Protein 1A