Protein phosphatase-1 is a novel regulator of the interaction between IRBIT and the inositol 1,4,5-trisphosphate receptor

Abstract

IRBIT is an IP3R [IP3 (inositol 1,4,5-trisphosphate) receptor]-binding protein that competes with IP3 for binding to the IP3R. Phosphorylation of IRBIT is essential for the interaction with the IP3R. The unique N-terminal region of IRBIT, residues 1-104 for mouse IRBIT, contains a PEST (Pro-Glu-Ser-Thr) domain with many putative phosphorylation sites. In the present study, we have identified a well-conserved PP1 (protein phosphatase-1)-binding site preceeding this PEST domain which enabled the binding of PP1 to IRBIT both in vitro and in vivo. IRBIT emerged as a mediator of its own dephosphorylation by associated PP1 and, hence, as a novel substrate specifier for PP1. Moreover, IRBIT-associated PP1 specifically dephosphorylated Ser68 of IRBIT. Phosphorylation of Ser68 was required for subsequent phosphorylation of Ser71 and Ser74, but the latter two sites were not targeted by PP1. We found that phosphorylation of Ser71 and Ser74 were sufficient to enable inhibition of IP3 binding to the IP3R by IRBIT. Finally, we have shown that mutational inactivation of the docking site for PP1 on IRBIT increased the affinity of IRBIT for the IP3R. This pinpoints PP1 as a key player in the regulation of IP3R-controlled Ca2+ signals.

IRBIT, Inositol 1,4,5-Triphosphate (IP3) Receptor-binding Protein Released with IP3, Binds Na+/H+ Exchanger NHE3 and Activates NHE3 Activity in Response to Calcium

“…Phosphorylation at Ser-68 is apparently required for subsequent phosphorylation of Ser-71 and Ser-74. Although the protein kinase phosphorylating Ser-68 in vivo remains unknown, an in silico analysis suggested several candidate protein kinases, including CaMKII (35). Our present study showed that IRBIT-dependent regulation of NHE3 activity was sensitive to the inhibition of CaM and CaMKII.…”

“…This NH 2 -terminal region contains a PEST domain enriched with serine residues and phosphorylation of several serine residues is essential for the interaction with IP 3 R (5, 34). A recent study showed that a small region of IRBIT preceding the PEST domain binds protein phosphatase-1, which dephosphorylates Ser-68 of IRBIT, suggesting that IRBIT is regulated by phosphorylation and dephosphorylation (35). Phosphorylation at Ser-68 is apparently required for subsequent phosphorylation of Ser-71 and Ser-74.…”

Regulation of nuclear envelope permeability in cell death and survival

“…The association of the IP3R inhibitory protein, IRBIT, to IP3Rs is also regulated by phosphorylation. 56 Dephosphorylated IRBIT, which might predominate in kinase-inhibited cells, cannot bind to IP3R. As a consequence, the receptor becomes sensitive to IP3 concentrations that exist in cells under resting conditions, which might result in a lower threshold for Ca 2+ release from the NE/ER.…”

Splicing variation of Long-IRBIT determines the target selectivity of IRBIT family proteins

“…It is notable that the first lysine (K) for the consensus PP1 docking sequence is not included in LongV4. It was reported that the mutation of IRBIT at the PP1 docking site affected the binding affinity of IP 3 R1 (25). The PP1 docking site is located near the multiple phosphorylation sites of the IRBIT family.…”