Lipid regulators of Pkh2 in Candida albicans, the protein kinase ortholog of mammalian PDK1

• Published in: Biochimica et biophysica acta Vol. 1861; no. 3; pp. 249 - 259
• Main Authors: Pastor-Flores, Daniel, Schulze, Jörg O., Bahí, Anna, Süß, Evelyn, Casamayor, Antonio, Biondi, Ricardo M.
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.03.2016
• Subjects: 3-Phosphoinositide-Dependent Protein Kinases - genetics; 3-Phosphoinositide-Dependent Protein Kinases - metabolism; Allostery; Amino Acid Sequence; Binding Sites; Budding yeast; C. albicans; Candida albicans - enzymology; Candida albicans - genetics; Cell wall integrity; Computational Biology; DHS; Fungal Proteins - metabolism; HEK293 Cells; Humans; Lipid Metabolism; Models, Molecular; Molecular Sequence Data; PDK1; Phosphatidic acid; Phosphatidic Acids - metabolism; Phosphatidylcholine; Phosphatidylethanolamine; Phosphatidylinositol 4,5-Diphosphate - metabolism; Phosphatidylinositol Phosphates - metabolism; Phosphatidylserine; Phosphatidylserines - metabolism; PHS; PI(3,4)P2; PI(3,4,5)P3; PI(4,5)P2; PIP3; Pkh; Protein Binding; Protein Structure, Tertiary; Sphingosine; Sphingosine - analogs & derivatives; Sphingosine - metabolism; Structure-Activity Relationship; Sulfatide; Sulfoglycosphingolipids - metabolism; Transfection; PI(3,4)P2; Phosphatidylethanolamine; Budding yeast; Allostery; Cell wall integrity; Pkh; Sulfatide; DHS; PI(4,5)P2; C. albicans; Sphingosine; Phosphatidylcholine; PIP3; PHS; Phosphatidylserine; PI(3,4,5)P3; PDK1; Phosphatidic acid; PI(4,5)P; PI(3,4,5)P; PIP; PI(3,4)P
• ISSN: 1388-1981; 0006-3002; 1879-2618; 1878-2434
• DOI: 10.1016/j.bbalip.2015.12.016

Pkh is the yeast ortholog of the mammalian 3-phosphoinositide-dependent protein kinase 1 (PDK1). Pkh phosphorylates the activation loop of Ypks, Tpks, Sch9 and also phosphorylates the eisosome components Lsp1 and Pil1, which play fundamental roles upstream of diverse signaling pathways, including the cell wall integrity and sphingosine/long-chain base (LCB) signaling pathways. In S. cerevisiae, two isoforms, ScPkh1 and ScPkh2, are required for cell viability, while only one ortholog exists in C. albicans, CaPkh2. In spite of the extensive information gathered on the role of Pkh in the LCB signaling, the yeast Pkh kinases are not known to bind lipids and previous studies did not identify PH domains in Pkh sequences. We now describe that the C-terminal region of CaPkh2 is required for its intrinsic kinase activity. In addition, we found that the C-terminal region of CaPkh2 enables its interaction with structural and signaling lipids. Our results further show that phosphatidylserine, phosphatidic acid, phosphatidylinositol (3,4 and 4,5)-biphosphates, and phosphatidylinositol (3,4,5)-trisphosphate inhibit Pkh activity, whereas sulfatide binds with high affinity but does not affect the intrinsic activity of CaPkh2. Interestingly, we identified that its human ortholog PDK1 also binds to sulfatide. We propose a mechanism by which lipids and dihydrosphingosine regulate CaPkh2 kinase activity by modulating the interaction of the C-terminal region with the kinase domain, while sulfatide-like lipids support localization CaPkh2 mediated by a C-terminal PH domain, without affecting kinase intrinsic activity. •CaPkh2, the Candida ortholog of PDK1, requires a C-terminal region for activity•CaPkh2 has a putative PH domain and binds to structural and signaling lipids•CaPkh2 binds to sulfatide, but it does not affect kinase activity•Phospholipds inhibit CaPkh2 in vitro activity while DHS reverses inhibition•Pkh kinases and PDK1 have similarities and differences in their mechanisms of regulation by lipids