Inhibitory action of linoleamide and oleamide toward sarco/endoplasmic reticulum Ca2+-ATPase.
SERCA maintains intracellular Ca2+homeostasis by sequestering cytosolic Ca2+into SR/ER stores. Two primary fatty acid amides (PFAAs), oleamide (18:19-cis) and linoleamide (18:29,12-cis), induce an increase in intracellular Ca2+levels, which might be caused by their inhibition of SERCA.



Three major SERCA isoforms, rSERCA1a, hSERCA2b, and hSERCA3a, were individually overexpressed in COS-1 cells, and the inhibitory action of PFAAs on Ca2+-ATPase activity of SERCA was examined.



The Ca2+-ATPase activity of each SERCA was inhibited in a concentration-dependent manner strongly by linoleamide (IC5015-53μM) and partially by oleamide (IC508.3-34μM). Inhibition by other PFAAs, such as stearamide (18:0) and elaidamide (18:19-trans), was hardly or slightly observed. With increasing dose, linoleamide decreased the apparent affinity for Ca2+and the apparent maximum velocity of Ca2+-ATPase activity of all SERCAs tested. Oleamide also lowered these values for hSERCA3a. Meanwhile, oleamide uniquely reduced the apparent Ca2+affinity of rSERCA1a and hSERCA2b: the reduction was considerably attenuated above certain concentrations of oleamide. The dissociation constants for SERCA interaction varied from 6 to 45μM in linoleamide and from 1.6 to 55μM in oleamide depending on the isoform.



Linoleamide and oleamide inhibit SERCA activity in the micromolar concentration range, and in a different manner. Both amides mainly suppress SERCA activity by lowering the Ca2+affinity of the enzyme.



Our findings imply a novel role of these PFAAs as modulators of intracellular Ca2+homeostasis via regulation of SERCA activity.
Department of Chemistry, Kyorin University School of Medicine, Mitaka, Tokyo 181-8611, Japan. Electronic address: s_yamamoto@ks.kyorin-u.ac.jp.
Sachiko
S
Yamamoto
Department of Materials Science, The University of Shiga Prefecture, Hassaka, Hikone 522-8533, Japan.
Munenori
M
Takehara
Department of Chemistry, Kyorin University School of Medicine, Mitaka, Tokyo 181-8611, Japan.
Makoto
M
Ushimaru
D000818:Animals; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism; D007700:Kinetics; D008041:Linoleic Acids / Q000494:pharmacology*; D009829:Oleic Acids / Q000494:pharmacology*; D011817:Rabbits; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism
D004791:Enzyme Inhibitors; D007527:Isoenzymes; D008041:Linoleic Acids; D009829:Oleic Acids; C420162:linoleamide; C029407:oleylamide; D053498:Sarcoplasmic Reticulum Calcium-Transporting ATPases
Calcium homeostasis; Inhibitor; Negative regulator; Primary fatty acid amide; Sarco/endoplasmic reticulum Ca(2+)-ATPase; Signaling lipid