Direct Modification and Activation of a Nuclear Receptor–PIP ₂ Complex by the Inositol Lipid Kinase IPMK

Abstract

Phosphatidylinositol (4,5)-bisphosphate (PIP2) is best known as a plasma membrane-bound regulatory lipid. While PIP2 and phosphoinositide-modifying enzymes coexist in the nucleus, their roles in the nucleus remain unclear. Here we show that the nuclear inositol polyphosphate multikinase (IPMK), which functions both as an inositol- and a PI3-kinase, interacts with the nuclear receptor SF-1 (NR5A1) and phosphorylates its bound ligand, PIP2. IPMK failed to recognize SF-1/PIP2 after blocking or displacing PIP2 from SF-1’s large hydrophobic pocket. In contrast to IPMK, p110 catalytic subunits of type 1 PI3-kinases were inactive on SF-1/PIP2. These and other in vitro analyses demonstrated specificity of IPMK for the SF-1/PIP2 protein/lipid complex. Once generated, SF-1/PIP3 is readily dephosphorylated by the lipid phosphatase PTEN. Importantly, decreasing IPMK or increasing PTEN expression greatly reduced SF-1 transcriptional activity. This ability of lipid kinases and phosphatases to alter the activity and directly remodel a non-membrane protein/lipid complex such SF-1/PIP2, establishes a new pathway for promoting lipid-mediated signaling in the nucleus.

The signaling phospholipid PIP 3 creates a new interaction surface on the nuclear receptor SF-1

“…Given that each position on the inositol head group has unique stereochemistry (33), one might imagine that the flipped orientations of the exposed PIP 2 and PIP 3 head groups differentially recruit stereo-specific coregulators. This hypothesis is consistent with our previous data showing that, at least on a subset of SF-1 target genes, PIP 3 but not PIP 2 is required for maximal SF-1 activity (13,15).…”

“…However, in the few available structures of PLTPs, the phosphoinositide head groups are not accessible to solvent (24,25,37). In stark contrast, our SF-1/PIP n structures establish that the PIP n head groups have unrestricted solvent accessibility, thus allowing lipid-modifying enzymes, such as IPMK and the lipid phosphatase PTEN to act on these exposed head groups (13). When considered as a PLTP that shuttles phospholipids within the nucleus, NR5As expand on that function by directly integrating lipid-signaling information into a transcriptional output (Fig.…”

“…Our previous work suggests that SF-1 bound to PIP 3 is active whereas SF-1 bound to PIP 2 is less active (13). To compare these different phosphoinositide-bound states, the SF-1/PI(4,5)P 2 structure (SF-1/PIP 2 ) was solved by molecular replacement, using PDB ID code 1YOW as the search model, and compared with the SF-1/PIP 3 structure (Table S2).…”

“…Consistent with these early reports, lipid-modifying enzymes responsible for phosphoinositide metabolism were also found in the nucleus (4)(5)(6)(7); however, the function of PIP n in this cellular compartment remains poorly defined. The nuclear receptors (NRs) steroidogenic factor 1 (SF-1, NR5A1) and liver receptor homolog 1 (LRH-1, NR5A2) bind phosphoinositides as well as other phospholipids in their large hydrophobic pockets (8)(9)(10)(11)(12)(13). The ability of NR5As to interact with PIP n is well-conserved with the Caenorhabditis elegans ortholog nhr-25 able to bind both PIP 2 and PIP 3 (14).…”

“…That phosphoinositides might serve as endogenous NR5A ligands is suggested by the fact that elevating cellular pools of PIP 3 increases SF-1 activity (15) and that impairing PIP 3 uptake decreases SF-1 activity (12). Further, when purified from mammalian cells, the phosphoinositide PIP 2 is found associated with SF-1 and can be modified by the lipid kinase, IPMK, as well as the lipid phosphatase, PTEN (13). Taken together, these data suggest that signaling phosphoinositides are biologically relevant ligands for SF-1.…”