Subtype-selective regulation of IP3 receptors by thimerosal via cysteine residues within the IP3-binding core and suppressor domain.

Inositol 1,4,5-trisphosphate receptors (IP3R) and ryanodine receptors are the most widely expressed intracellular Ca2+ channels and both are regulated by sulphydryl reagents. In DT40 cells stably expressing single subtypes of mammalian IP3R, low concentrations of thimerosal, which oxidises thiols to form a thiomercurylethyl complex, increased the sensitivity of IP3-evoked Ca2+ release via IP3R1 and IP3R2, but inhibited IP3R3. Activation of IP3R is initiated by IP3 binding to the IP3-binding core (IBC, residues 224-604) and proceeds via re-arrangement of an interface between the IBC and suppressor domain (SD, residues 1-223). Thimerosal (100 µM) stimulated IP3 binding to the isolated N-terminal (NT, residues 1-604) of IP3R1 and IP3R2, but not to that of IP3R3. Binding of a competitive antagonist (heparin) or partial agonist (dimeric-IP3) to NT1 was unaffected by thimerosal, suggesting that the effect of thimerosal is specifically related to IP3R activation. IP3 binding to NT1 in which all Cys were replaced by Ala was insensitive to thimerosal, so too were NT1 in which Cys were replaced in either the SD or IBC. This demonstrates that thimerosal interacts directly with Cys in both the SD and IBC. Chimeric proteins in which the SD of the IP3R was replaced by the structurally related A-domain of a ryanodine receptor were functional, but thimerosal inhibited both IP3 binding to the chimeric NT and IP3-evoked Ca2+ release from the chimeric IP3R. This is t...

Source: BJ Signal - January 2, 2013 Category: Biochemistry Authors: S A. Khan, A M Rossi, A M Riley, B V.L. Potter, C W. Taylor Tags: BJ Signal Source Type: research