Interaction between the PH and START domains of ceramide transfer protein competes with phosphatidylinositol 4-phosphate binding by the PH domain

Prashek, J.; Bouyain, Samuel; Fu, Mingui; Li, Yong; Berkeš, Dušan; Yao, Xiaolan

doi:10.1074/jbc.m117.780007

Cited by 37 publications

(55 citation statements)

References 49 publications

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“…This model was recently supported by a structural study by Prashek et al (20), who showed that isolated PH and START domains of CERT interacted with each other and that specific regions on the surface of the PH domain, which are involved in the interaction between the two domains, are partially overlapped with the PI4P-binding site. In addition, Prashek et al (20) demonstrated that amino acid substitutions in the PH-START-interacting surface abrogated the repressive effects of the CERT 10E mutation.…”

Section: Discussionmentioning

confidence: 99%

“…In this sense, the interaction between the PH domain and phosphorylated SRM can be an "intermediate" form toward a more drastic conformational rearrangement of CERT and will lead the PH-START interdomain interaction for mutually and functionally silencing conformations. The START domain is required for the repression of the PI4P-binding activity of the CERT PH domain under these conditions (9,20). The inactive conformer is mainly distributed in the cytosol as an inactive pool of CERT, whereas inactive CERT is dephosphorylated in response to specific stimuli (e.g.…”

Section: Discussionmentioning

confidence: 99%

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Phosphoinositide binding by the PH domain in ceramide transfer protein (CERT) is inhibited by hyperphosphorylation of an adjacent serine-repeat motif

Sugiki¹,

Egawa²,

Kumagai³

et al. 2018

Journal of Biological Chemistry

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Sphingolipids such as ceramide are important constituents of cell membranes. The ceramide transfer protein (CERT) moves ceramide from the endoplasmic reticulum to the Golgi apparatus in a nonvesicular manner. Hyperphosphorylation of the serine-repeat motif (SRM) adjacent to the pleckstrin homology (PH) domain of CERT down-regulates the inter-organelle ceramide transport function of CERT. However, the mechanistic details of this down-regulation remain elusive. Using solution NMR and binding assays, we herein show that a hyperphosphorylation-mimetic CERT variant in which 10 serine/threonine residues of SRM had been replaced with glutamate residues (the 10E variant) displays an intramolecular interaction between SRM and positively charged regions of the PH domain, which are involved in the binding of this domain to phosphatidylinositol 4-monophosphate (PI4P). Of note, the binding of the PH domain to PI4P-embedded membranes was attenuated by the SRM 10E substitutions in cell-free assays. Moreover, the 10E substitutions reduced the Golgi-targeting activity of the PH-SRM construct in living cells. These results indicate that hyperphosphorylated SRM directly interacts with the surface of the PH domain in an intramolecular manner, thereby decreasing the PI4P-binding activity of the PH domain. In light of these findings, we propose that the hyperphosphorylation of SRM may trigger the dissociation of CERT from the Golgi apparatus, resulting in a functionally less active conformation of CERT.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Phosphoinositide binding by the PH domain in ceramide transfer protein (CERT) is inhibited by hyperphosphorylation of an adjacent serine-repeat motif

Sugiki¹,

Egawa²,

Kumagai³

et al. 2018

Journal of Biological Chemistry

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“…Recently, Xray crystallography with the purified CERT PH and START domains revealed that a basic groove in the PH domain snugly binds to the negatively charged loop of the START domain [43]. Mutations disrupting the PH/START interaction increase both PtdIns(4)Pbinding activity and ceramide-transfer activity of CERT 10E, thereby supporting the inter-domain steric hindrance model [43]. The CERT PH domain also electrostatically interacts with the phosphor-mimetic SRM (SRM 10E; Fig.…”

Section: Serine-repeat Motifmentioning

confidence: 95%

Structure, functions and regulation of CERT, a lipid‐transfer protein for the delivery of ceramide at the ER–Golgi membrane contact sites

Kumagai

Hanada

2019

FEBS Letters

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The inter‐organelle transport of lipids must be regulated to ensure appropriate lipid composition of each organelle. In mammalian cells, ceramide synthesised in the endoplasmic reticulum (ER) is transported to the trans‐Golgi regions, where ceramide is converted to sphingomyelin (SM) with the concomitant production of diacylglycerol. Ceramide transport protein (CERT) transports ceramide from the ER to the trans‐Golgi regions at the ER–Golgi membrane contact sites (MCS). The function of CERT is down‐regulated by multisite phosphorylation of a serine‐repeat motif (SRM) and up‐regulated by phosphorylation of serine 315 in CERT. Multisite phosphorylation of the SRM is primed by protein kinase D, which is activated by diacylglycerol. The function of CERT is regulated by a phosphorylation‐dependent feedback mechanism in response to cellular requirements of SM. CERT‐dependent ceramide transport is also affected by the pool of phosphatidylinositol (PtdIns)‐4‐phosphate (PtdIns(4)P) in the trans‐Golgi regions, while the PtdIns(4)P pool is regulated by PtdIns‐4‐kinases and oxysterol‐binding protein. The ER‐Golgi MCS may serve as inter‐organelle communication zones, in which many factors work in concert to serve as an extensive rheostat of SM, diacylglycerol, cholesterol and PtdIns(4)P.

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“…LTP targeting can be regulated by post-translational modification. CERT targeting is affected by two different phosphorylations, one of which activates the FFAT, while the other causes autoinhibitory binding of the PH and StARkin domains [70][71][72] . LTPs and their localisation can also be regulated by Ca 2+ signalling.…”

Section: Dual Membrane Targeting Domainsmentioning

confidence: 99%

Lipid transfer proteins: the lipid commute via shuttles, bridges and tubes

Wong

Gatta

Levine

2018

Nat Rev Mol Cell Biol

366

437

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Lipids are distributed in a highly asymmetric fashion in different cellular membranes. Only a minority of lipids achieve their final intracellular distribution by selection into the membranes of transport vesicles. Instead, the bulk of lipid traffic is mediated by a large group of lipid transfer proteins (LTPs), which move small numbers of lipids at a time using hydrophobic cavities that stabilise lipid outside membranes. Despite the first discoveries of LTPs almost 50 years ago, most progress has been made in the last few years, leading to considerable temporal and spatial refinement in our understanding. The number of known LTPs has increased, with exciting discoveries of multimeric assemblies. Structural studies of LTPs have progressed from static crystal structures to dynamic structural approaches that show how conformational changes contribute to lipid handling at a sub-millisecond timescale. Many intracellular LTPs localise to membrane contact sites, nanoscale zones where an LTP can form either a shuttle, bridge or tube linking donor and acceptor compartments. Understanding how each lipid achieves its final destination at the molecular level allows a better explanation of the range of defects that occur in disease, with therapies being developed to target lipid transfer.

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Interaction between the PH and START domains of ceramide transfer protein competes with phosphatidylinositol 4-phosphate binding by the PH domain

Cited by 37 publications

References 49 publications

Phosphoinositide binding by the PH domain in ceramide transfer protein (CERT) is inhibited by hyperphosphorylation of an adjacent serine-repeat motif

Phosphoinositide binding by the PH domain in ceramide transfer protein (CERT) is inhibited by hyperphosphorylation of an adjacent serine-repeat motif

Structure, functions and regulation of CERT, a lipid‐transfer protein for the delivery of ceramide at the ER–Golgi membrane contact sites

Lipid transfer proteins: the lipid commute via shuttles, bridges and tubes

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