A novel Ca2+-induced Ca2+ release mechanism in A7r5 cells regulated by calmodulin-like proteins.
Intracellular Ca2+ release is involved in setting up Ca2+ signals in all eukaryotic cells. Here we report that an increase in free Ca2+ concentration triggered the release of up to 41 +/- 3% of the intracellular Ca2+ stores in permeabilized A7r5 (embryonic rat aorta) cells with an EC50 of 700 nm. This type of Ca2+-induced Ca2+ release (CICR) was neither mediated by inositol 1,4,5-trisphosphate receptors nor by ryanodine receptors, because it was not blocked by heparin, 2-aminoethoxydiphenyl borate, xestospongin C, ruthenium red, or ryanodine. ATP dose-dependently stimulated the CICR mechanism, whereas 10 mm MgCl2 abolished it. CICR was not affected by exogenously added calmodulin (CaM), but CaM1234, a Ca2+-insensitive CaM mutant, strongly inhibited the CICR mechanism. Other proteins of the CaM-like neuronal Ca2+-sensor protein family such as Ca2+-binding protein 1 and neuronal Ca2+ sensor-1 were equally potent for inhibiting the CICR. Removal of endogenous CaM, using a CaM-binding peptide derived from the ryanodine receptor type-1 (amino acids 3614-3643) prevented subsequent activation of the CICR mechanism. A similar CICR mechanism was also found in 16HBE14o-(human bronchial mucosa) cells. We conclude that A7r5 and 16HBE14o-cells express a novel type of CICR mechanism that is silent in normal resting conditions due to inhibition by CaM but becomes activated by a Ca2+-dependent dissociation of CaM. This CICR mechanism, which may be regulated by members of the family of neuronal Ca2+-sensor proteins, may provide an additional route for Ca2+ release that could allow amplification of small Ca2+ signals.
Laboratorium voor Fysiologie, K.U. Leuven Campus Gasthuisberg O/N, Herestraat 49, B-3000 Leuven, Belgium. nael.nadifkasri@med.kuleuven.ac.be
Nael Nadif
NN
Kasri
Ilse
I
Sienaert
Jan B
JB
Parys
Geert
G
Callewaert
Ludwig
L
Missiaen
Andreas
A
Jeromin
Humbert
H
De Smedt
D000255:Adenosine Triphosphate / Q000494:pharmacology; D020816:Amino Acid Motifs; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002478:Cells, Cultured; D003001:Cloning, Molecular; D004305:Dose-Response Relationship, Drug; D006801:Humans; D008274:Magnesium / Q000378:metabolism; D051379:Mice; D010452:Peptide Biosynthesis; D017434:Protein Structure, Tertiary; D051381:Rats; D019837:Ryanodine Receptor Calcium Release Channel / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors
D002147:Calmodulin; D019837:Ryanodine Receptor Calcium Release Channel; D000255:Adenosine Triphosphate; D008274:Magnesium; D002118:Calcium