Proteome of acidic phospholipid-binding proteins: spatial and temporal regulation of Coronin 1A by phosphoinositides.
Reversible interactions between acidic phospholipids in the cellular membrane and proteins in the cytosol play fundamental roles in a wide variety of physiological events. Here, we present a novel approach to the identification of acidic phospholipid-binding proteins using nano-liquid chromatography-tandem mass spectrometry. We found more than 400 proteins, including proteins with previously known acidic phospholipid-binding properties, and confirmed that several candidates, such as Coronin 1A, mDia1 (Diaphanous-related formin-1), PIR121/CYFIP2, EB2 (end plus binding protein-2), KIF21A (kinesin family member 21A), eEF1A1 (translation elongation factor 1alpha1), and TRIM2, directly bind to acidic phospholipids. Among such novel proteins, we provide evidence that Coronin 1A activity, which disassembles Arp2/3-containing actin filament branches, is spatially and temporally regulated by phosphatidylinositol 4,5-bisphosphate (PI(4,5)P(2)). Whereas Coronin 1A co-localizes with PI(4,5)P(2) at the plasma membrane in resting cells, it is dissociated from the plasma membrane during lamellipodia formation where the PI(4,5)P(2) signal is significantly reduced. Our in vitro experiments show that Coronin 1A preferentially binds to PI(4,5)P(2)-containing liposomes and that PI(4,5)P(2) antagonizes the ability of Coronin 1A to disassemble actin filament branches, indicating a spatiotemporal regulation of Coronin 1A via a direct interaction with the plasma membrane lipid. Collectively, our proteomics data provide a list of potential acidic phospholipid-binding protein candidates ranging from the actin regulatory proteins to translational regulators.
Tsujita
Kazuya
K
Division of Lipid Biochemistry, Department of Biochemistry and Molecular Biology, Kobe University Graduate School of Medicine, Kobe 650-0017, Japan.
Itoh
Toshiki
T
Kondo
Akihiro
A
Oyama
Masaaki
M
Kozuka-Hata
Hiroko
H
Irino
Yasuhiro
Y
Hasegawa
Junya
J
Takenawa
Tadaomi
T
D008841:Actin Cytoskeleton / Q000378:metabolism; D000818:Animals; D001923:Brain Chemistry; D002462:Cell Membrane / Q000378:metabolism; D002853:Chromatography, Liquid; D008840:Microfilament Proteins / Q000378:metabolism*; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D010716:Phosphatidylinositols / Q000378:metabolism*; D010743:Phospholipids / Q000378:metabolism*; D011485:Protein Binding; D011506:Proteins / Q000032:analysis / Q000378:metabolism; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods; D051381:Rats; D053719:Tandem Mass Spectrometry
D008840:Microfilament Proteins; D019269:Phosphatidylinositol 4,5-Diphosphate; D018129:Phosphatidylinositol Phosphates; D010716:Phosphatidylinositols; D010743:Phospholipids; D011506:Proteins; D020543:Proteome; C072061:coronin proteins