Biochemical and Biophysical Research Communications
Phosphatidylinositol (3,4,5)-trisphosphate binds to sortilin and competes with neurotensin: Implications for very low density lipoprotein binding
Introduction
Sortilin is a multi-ligand binding protein with numerous binding partners [1]. Sortilin binds neurotensin (NT) at two sites: the carboxy terminus (C-term) site 1, and the amino terminus (N-term) site 2 [2]. Sortilin also binds B100-containing very low density lipoprotein (VLDL) and LDL [3], [4]. We previously demonstrated that sortilin facilitates the secretion of VLDL-B100 by insulin-sensitive McArdle RH7777 (McA) cells [5]. We have further shown that insulin enhances B100 binding to sortilin in McA cells [6] preceding autophagic destruction [7]. Considering the association of sortilin with VLDL binding and with VLDL-B100 degradation in response to insulin, we explored the possibility for insulin-generated phosphatidylinositol (3,4,5)-trisphosphate (PIP3) to interact directly with sortilin. We specifically tested the potential of PIP3 binding to NT defined binding sites on sortilin. Our results suggest a possible role for PIP3 present on VLDL to directly modulate lipoprotein-sortilin interactions.
Section snippets
Materials
Plasma from fasted Sprague Dawley rats was obtained from BioreclamationIVT, (Westbury, NY) and used to prepare rat lipoproteins. Bovine serum albumin (BSA) (A7285) was from Sigma-Aldrich Corp. (St. Louis, MO). The following materials and reagents were obtained from Echelon Biosciences, Inc. (St. Lake City, UT) including PIP3 PolyPIPosomes (Y-P039); dibutanoyl PIP3 (diC4-PIP3) (P-3904); dipalmitoyl PIP3 (diC16-PIP3) (P-3916); purified anti-PIP3 IgG (Z-P345b), and purified anti-PIP3 IgM (ZP345)
Sortilin is a PIP3 binding protein
Sortilin binding to NT has been characterized and involves N-term (site 2) and C-term (site 1) binding sites for binding to a single molecule of sortilin [2], [15]. A compound was identified with strong theoretical binding to site 2 (cpd984), and in biologic experiments cpd984 incubation increased VLDL-B100 secretion by McA cells [5]. In Fig. 1A, N-term NT is depicted in site 2 located across the central space of sortilin from PIP3 computationally docked into site 1 of hsortilin. The
Acknowledgements
We thank Dr. Stephen Sligar for his generous contribution of MSP1D1 protein and Dr. James Morrissey's laboratory in facilitating the preparation of PIP3 nanodiscs. This study was supported by grants from the National Institutes of Health DK100163 (JDS) and GM101132 (RAF).
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Correction: A small-molecule competitive inhibitor of phosphatidic acid binding by the AAA+ protein NSF/Sec18 blocks the SNARE-priming stage of vacuole fusion (Journal of Biological Chemistry (2019) 294(46) (17168–17185), (S002192582030716X), (10.1074/jbc.ra119.008865))
2022, Journal of Biological ChemistrySorting through the extensive and confusing roles of sortilin in metabolic disease
2022, Journal of Lipid ResearchCitation Excerpt :However, remarkably, cpd984 enhances neurotensin binding to sortilin (115). Interestingly, cpd984 also enhances the binding of other ligands to sortilin through the C-terminal neurotensin binding site, including phosphatidylinositol (3,4,5)-triphosphate (155), and likely apoB-100 (115). As discussed previously, the group developed various insulin-sensitive McA hepatoma cell lines in which sortilin levels were differentially reduced with siRNA.
A small-molecule competitive inhibitor of phosphatidic acid binding by the AAA+ protein NSF/Sec18 blocks the SNARE-priming stage of vacuole fusion
2019, Journal of Biological ChemistryCitation Excerpt :Approximately 2000 RU of 5% PA nanodiscs were immobilized noncovalently using 100 mm NiSO4 flowed at 10 μl/s followed by a blank buffer injection of HEPES, pH 7.4, 150 mm NaCl (HBS buffer). Injections were performed in HBS buffer at a flow rate of 30 μl/min with an association time of 90 s and dissociation time of 300 s, and binding was measured in relative response units (RU) as described (44). Regeneration with EDTA was performed at flow rate 30 μl/s for 120 s using 100 μm EDTA buffer.
Phosphatidic acid induces conformational changes in Sec18 protomers that prevent SNARE priming
2019, Journal of Biological ChemistryCitation Excerpt :Approximately 2000 response units of 5% PA nanodiscs were immobilized noncovalently using 1 μm NiSO4 flowed at 10 μl/s followed by a blank buffer injection of HEPES, pH 7.4, 150 mm NaCl (HBS buffer). Injections were performed in HBS buffer at a flow rate of 30 μl/min with an association time of 90 s, dissociation time of 300 s, and binding was measured in relative response units as described (67). Regeneration with EDTA was performed at flow rate 30 μl/s for 120 s using 100 nm EDTA buffer.
Anti-sortilin1 Antibody Up-Regulates Progranulin via Sortilin1 Down-Regulation
2020, Frontiers in Neuroscience