The phosphatidylinositol transfer protein, RdgBβ binds 14-3-3 via its unstructured C-terminus whereas its lipid binding domain interacts with the integral membrane protein, ATRAP (Angiotensin II Type I Receptor-associated Protein), for membrane recruitment

Kathryn L Garner , Michelle Li , Natalie Ugwuanya , S Cockcroft (1)

1 Department of Neuroscience, Physiology and Pharmacology

Phosphatidylinositol transfer proteins (PITPs) bind and transfer phosphatidylinositol between intracellular membranes and participate in many cellular processes including signalling, lipid metabolism and membrane traffic. The largely uncharacterised PITP, RdgBb (PITPNC1), contains a long C-terminal disordered region following its defining N-terminal PITP domain. We report that the C-terminus contains two tandem phosphorylated binding sites (Ser274 and Ser299) for 14-3-3. The C-terminus also contains PEST sequences which are shielded by 14-3-3 binding. Like many proteins containing PEST sequences, the levels of RdgBβ are regulated by proteolysis. RdgBb is degraded with a half-life of 4hrs following ubiquitination via the proteasome. A mutant RdgBβ which is unable to bind 14-3-3 is degraded even faster with a half-life of 2hrs. In vitro, RdgBβ is 100-fold less active than PITPα for PI transfer and RdgBβ proteins (wild-type and a mutant that cannot bind 14-3-3) either expressed in COS-7 cells or endogenous proteins from heart cytosol do not exhibit transfer activity. When cells are treated with PMA, the PITP domain of RdgBβ interacts with the integral membrane protein, ATRAP (Angiotensin II receptor-associated protein; Alt name: AGTRAP) causing membrane recruitment. We suggest that RdgBβ executes its function following recruitment to membranes via its PITP domain and the C-terminal end of the protein could regulate entry to the hydrophobic cavity.

Mots clés: Life Sciences

Kathryn L Garner, Michelle Li, Natalie Ugwuanya, S Cockcroft. The phosphatidylinositol transfer protein, RdgBβ binds 14-3-3 via its unstructured C-terminus whereas its lipid binding domain interacts with the integral membrane protein, ATRAP (Angiotensin II Type I Receptor-associated Protein), for membrane recruitment. Biochemical Journal, 2011, 439 (1), pp.97-111. ⟨10.1042/BJ20110649⟩. ⟨hal-00628672⟩