1. Oxysterol binding proteins and PIPs are required for non-vesicular sterol transport from the plasma membrane to the ER. The proper intracellular distribution of sterols such as cholesterol is critical for numerous cellular functions, including signal transduction and protein trafficking. Sterols are moved among cellular compartments by both vesicular and poorly understood nonvesicular pathways. We have previously shown that a non-vesicular pathway moves sterols from the plasma membrane (PM) to the endoplasmic reticulum in the yeast S. cerevisiae. We found that this transport requires oxysterol-binding proteins (OSBPs), a large family of lipid-binding proteins that is conserved from yeast to humans. In addition, we have been able to show that some of these yeast OSBPs transport sterols and other lipids in vitro. Thus, lipid transport is one of the functions of this important class of proteins. We also found that sterol transport by OSBPs is regulated by particular phosphoinositides (PIPs). Because different PIP-species are enriched in various cellular compartments, PIP-stimulation of ORPs likely serves to regulate the movement of sterols (and possibility other lipids) to particular organelles by ORPs, perhaps in response to cell signaling events. This work is described in a paper in the Journal of Cell Biology. [unreadable] [unreadable] 2. Proteins required for ER tubule formation. In yeast and higher eukaryotes, portions of the ER form a dynamic tubular network, often closely apposed to the plasma membrane. This structure is thought to be important for proper organelle function, but how it is established and maintained is not well understood. In collaboration with Gia Voeltz and Tom Rapoport at Harvard Medical School we have recently found a role for a large family of proteins known as reticulons in determining ER structure. We are working to understand how these proteins shape membranes and function in cells. This work is described in a publication in Cell.[unreadable] [unreadable] 3. Identification of a RAN-independent, calmodulin-dependent nuclear localization signal (NLS) in the high mobility group (HMG) transcription factor Nhp6ap. We have identified the NLS required for the RAN-independent import of Nhp6ap and demonstrated that it can bind calmodulin, a protein that may play a role in a novel nuclear import pathway. Mutants with calmodulin defects fail to localize a protein containing the Nhp6ap NLS to the nucleus. These mutants do not affect the localization of a reporter with a RAN-dependent NLS. Thus, Nhp6ap is imported into the nucleus by a novel, calmodulin-dependent, RAN-independent nuclear pathway. We have demonstrated a direct interaction between calmodulin and Nhp6ap and have isolated mutations in Nhp6ap that fail to interact with calmodulin. These mutants are not imported into the nucleus We are currently using a combinatin of biochemical and genetic approaches to identify other components of this novel import pathway. A manuscript describing this work is under preparation.
NIH ExPorter
NIH ExPorter