Reconstitution of ORP-mediated lipid exchange coupled to PI4P metabolism.

Oxysterol-binding protein-related proteins (ORPs) play key roles in the distribution of lipids in eukaryotic cells by exchanging sterol or phosphatidylserine for PI4P between the endoplasmic reticulum (ER) and other cell regions. However, it is unclear how their exchange capacity is coupled to PI4P metabolism. To address this question quantitatively, we analyze the activity of a representative ORP, Osh4p, in an ER/Golgi interface reconstituted with ER- and Golgi-mimetic membranes functionalized with PI4P phosphatase Sac1p and phosphatidylinositol (PI) 4-kinase, respectively.

Tumor protein D54 binds intracellular nanovesicles via an extended amphipathic region.

Tumor protein D54 (TPD54) is an abundant cytosolic protein that belongs to the TPD52 family, a family of four proteins (TPD52, 53, 54, and 55) that are overexpressed in several cancer cells. Even though the functions of these proteins remain elusive, recent investigations indicate that TPD54 binds to very small cytosolic vesicles with a diameter of ca. 30 nm, half the size of classical (e.

In situ artificial contact sites (ISACS) between synthetic and endogenous organelle membranes allow for quantification of protein-tethering activities.

Membrane contact sites are specialized areas where the membranes of two distinct organelles are physically connected and allow for the exchange of molecules and for signaling processes. Understanding the mechanisms whereby proteins localize to and function in these structures is of special interest; however, methods allowing for reconstitution of these contact sites are few and only based on synthetic membranes and recombinant proteins. Here, we devised a strategy to create in situ artificial contact sites between synthetic and endogenous organelle membranes.

Functional analyses of phosphatidylserine/PI(4)P exchangers with diverse lipid species and membrane contexts reveal unanticipated rules on lipid transfer.

Background: Lipid species are accurately distributed in the eukaryotic cell so that organelle and plasma membranes have an adequate lipid composition to support numerous cellular functions. In the plasma membrane, a precise regulation of the level of lipids such as phosphatidylserine, PI(4)P, and PI(4,5)P, is critical for maintaining the signaling competence of the cell. Several lipid transfer proteins of the ORP/Osh family contribute to this fine-tuning by delivering PS, synthesized in the endoplasmic reticulum, to the plasma membrane in exchange for PI(4)P.

Fluorescence-Based Measurements of Phosphatidylserine/Phosphatidylinositol 4-Phosphate Exchange Between Membranes.

Several members of the evolutionarily conserved oxysterol-binding protein (OSBP)-related proteins(ORP)/OSBP homologs (Osh) family have recently been found to represent a novel lipid transfer protein (LTP) group in yeast and human cells. They transfer phosphatidylserine (PS) from the endoplasmic reticulum (ER) to the plasma membrane (PM) via PS/phosphatidylinositol 4-phosphate (PI(4)P) exchange cycles. This finding allows a better understanding of how PS, which is critical for signaling processes, is distributed throughout the cell and the investigation of the link between this process and phosphoinositide (PIP) metabolism.

Oncogenic cooperation between H-Twist and N-Myc overrides failsafe programs in cancer cells.

N-Myc oncogene amplification is a frequent event in neuroblastoma and is strongly correlated with advanced disease stage and treatment failure. Similarly to c-Myc oncogenic activation, N-Myc deregulation promotes both cell proliferation and p53-dependent apoptosis by sensitizing cells to a variety of insults. Intriguingly, p53 mutations are uncommon in neuroblastomas, strongly suggesting that an alternative cooperating event circumvents this safeguard against oncogene-driven neoplasia.