Peptide fragments of the dihydropyridine receptor can modulate cardiac ryanodine receptor channel activity and sarcoplasmic reticulum Ca2+ release.
We show that peptide fragments of the dihydropyridine receptor II-III loop alter cardiac RyR (ryanodine receptor) channel activity in a cytoplasmic Ca2+-dependent manner. The peptides were AC (Thr-793-Ala-812 of the cardiac dihydropyridine receptor), AS (Thr-671-Leu-690 of the skeletal dihydropyridine receptor), and a modified AS peptide [AS(D-R18)], with an extended helical structure. The peptides added to the cytoplasmic side of channels in lipid bilayers at > or = 10 nM activated channels when the cytoplasmic [Ca2+] was 100 nM, but either inhibited or did not affect channel activity when the cytoplasmic [Ca2+] was 10 or 100 microM. Both activation and inhibition were independent of bilayer potential. Activation by AS, but not by AC or AS(D-R18), was reduced at peptide concentrations >1 mM in a voltage-dependent manner (at +40 mV). In control experiments, channels were not activated by the scrambled AS sequence (ASS) or skeletal II-III loop peptide (NB). Resting Ca2+ release from cardiac sarcoplasmic reticulum was not altered by peptide AC, but Ca2+-induced Ca2+ release was depressed. Resting and Ca2+-induced Ca2+ release were enhanced by both the native and modified AS peptides. NMR revealed (i) that the structure of peptide AS(D-R18) is not influenced by [Ca2+] and (ii) that peptide AC adopts a helical structure, particularly in the region containing positively charged residues. This is the first report of specific functional interactions between dihydropyridine receptor A region peptides and cardiac RyR ion channels in lipid bilayers.
Dulhunty
Angela F
AF
John Curtin School of Medical Research, Australian National University, Canberra, ACT 2601, Australia. angela.dulhunty@anu.edu.au
Curtis
Suzanne M
SM
Cengia
Louise
L
Sakowska
Magdalena
M
Casarotto
Marco G
MG
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000737:chemistry / Q000502:physiology*; D004305:Dose-Response Relationship, Drug; D017136:Ion Transport; D008051:Lipid Bilayers; D046911:Macromolecular Substances; D008861:Microsomes / Q000187:drug effects / Q000378:metabolism; D008969:Molecular Sequence Data; D009119:Muscle Contraction; D009124:Muscle Proteins / Q000737:chemistry / Q000502:physiology; D018482:Muscle, Skeletal / Q000737:chemistry; D009206:Myocardium / Q000737:chemistry; D010446:Peptide Fragments / Q000494:pharmacology*; D017433:Protein Structure, Secondary; D011817:Rabbits; D019837:Ryanodine Receptor Calcium Release Channel / Q000187:drug effects* / Q000378:metabolism; D012519:Sarcoplasmic Reticulum / Q000187:drug effects* / Q000378:metabolism
D020746:Calcium Channels, L-Type; D008051:Lipid Bilayers; D046911:Macromolecular Substances; D009124:Muscle Proteins; D010446:Peptide Fragments; D019837:Ryanodine Receptor Calcium Release Channel; D002118:Calcium