Calumenin, a multiple EF-hands Ca2+-binding protein, interacts with ryanodine receptor-1 in rabbit skeletal sarcoplasmic reticulum.
Calumenin is a multiple EF-hand Ca2+-binding protein located in endo/sarcoplasmic reticulum of mammalian tissues. In the present study, we cloned two rabbit calumenin isoforms (rabbit calumenin-1 and -2, GenBank Accession Nos. SY225335 and AY225336, respectively) by RT-PCR. Both isoforms contain a 19 aa N-terminal signal sequence, 6 EF-hand domains, and a C-terminal ER/SR retrieval signal, HDEF. Both calumenin isoforms exist in rabbit cardiac and skeletal muscles, but calumenin-2 is the main isoform in skeletal muscle. Presence of calumenin in rabbit sarcoplasmic reticulum (SR) was identified by Western blot analysis. GST-pull down and co-immunoprecipitation experiments showed that ryanodine receptor 1 (RyR1) interacted with calumenin-2 in millimolar Ca2+ concentration range. Experiments of gradual EF-hand deletions suggest that the second EF-hand domain is essential for calumenin binding to RyR1. Adenovirus-mediated overexpression of calumenin-2 in C2C12 myotubes led to increased caffeine-induced Ca2+ release, but decreased depolarization-induced Ca2+ release. Taken together, we propose that calumenin-2 in the SR lumen can directly regulate the RyR1 activity in Ca2+-dependent manner.
Jung
Dai Hyun
DH
Department of Life Science, Gwangju Institute of Science and Technology, Gwangju 500-712, Republic of Korea.
Mo
Sang Hyun
SH
Kim
Do Han
DH
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D003001:Cloning, Molecular; D020832:EF Hand Motifs; D047468:Immunoprecipitation; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000737:chemistry / Q000378:metabolism*; D025941:Protein Interaction Mapping; D020033:Protein Isoforms / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D011817:Rabbits; D019837:Ryanodine Receptor Calcium Release Channel / Q000187:drug effects / Q000378:metabolism*; D012519:Sarcoplasmic Reticulum / Q000737:chemistry / Q000378:metabolism*
D002135:Calcium-Binding Proteins; D020033:Protein Isoforms; D019837:Ryanodine Receptor Calcium Release Channel; D002118:Calcium