Lipid-transfer proteins as regulators of membrane trafficking

Transport of lipids between biological membranes is mediated both by vesicular and non-vesicular transport mechanisms. Lipid-transfer proteins (LTPs) mediate a non-vesicular lipid transport between membrane bilayers by facilitating the exchange of lipid monomers between a donor and acceptor membrane. Although LTPs have been implicated in the regulation of membrane trafficking, the underlying mechanisms of their action in intact cells remain largely unknown.

We previously found that the LTP, Nir2, which transfers phosphatidylinositol (PI) and phosphatidycholine (PC) between membranes in vitro, is required for maintaining the structural and functional integrity of the Golgi complex by regulating the level of a key lipid, diacylglycerol (DAG), in this organelle (Litvak et al., 2005). DAG is a strongly conical component of the bilayer that induces membrane bending and the formation of highly curved intermediates, thereby facilitating membrane budding, fusion, and fission events (Lev, 2006).The level of DAG in the Golgi membrane can, therefore, directly affect Golgi-mediated transport events. Our finding that Nir2 is involved in maintaining a critical pool of DAG in the Golgi, provides a novel mechanism for regulating membrane transport by PI/PC-transfer proteins, and demonstrates the interface between lipid homeostasis and Golgi secretory function.

More recently we found that Nir2 functions coordinately with other LTPs at the ER-Golgi membrane contact sites (MCSs) to tightly and locally control the lipid composition of the Golgi complex and thereby its secretory function (Peretti et al., 2008). This coordinated function is mediated by LTPs containing the FFAT motif (double phenylalanine in an acidic tract), including Nir2, CERT (ceramide transfer protein) and OSBP (oxysterol-binding protein). We found that the FFAT motif of Nir2 protein, as well as of its other family members, Nir1 and Nir3, targets the proteins to the ER by direct interaction with the integral ER membrane proteins of the VAP family; VAP-A and VAP-B (Amarilio et al., 2005).

The VAP are highly conserved proteins that have been identified in all eukaryotic organisms from yeast to human. They have been implicated in the regulation of a wide range of cellular processes including, membrane trafficking, lipid metabolism, the unfolded protein response (UPR), and microtubule organization (Lev et al., 2008). Recently, a single missense mutation within the human VAP-B gene, which substitutes a conserved proline residue at position 56 by a serine (P56S), was identified in three forms of familial motor neuron diseases (MNDs). This discovery has opened many questions related to the cellular functions of VAPs and the underlying mechanisms of VAP-B(P56S)-induced MNDs. Our preliminary studies indicate that the expression of the VAP-B(P56S) mutant in cultured cells results in the formation of insoluble protein aggregates. The formation of insoluble protein aggregates is a hallmark of many human neurodegenerative disorders. Yet, the molecular mechanisms underling the formation VAP-B(P56S) insoluble protein aggregates and their neurotoxicity effects are poorly understood.

The integral ER proteins VAP-A and VAP–B interact with LTPs containing a FFAT motif. Co-expression of VAP-B with either Nir2 or Nir3 affects the ER structure, while the P56S mutation in VAP-B induces the formation of protein aggregates and is involved in Amyotrophic Lateral Sclerosis (ALS) in humans.

Current ongoing studies are aimed at elucidating:

1. The mechanisms by which LTPs of the Nir/rdgB (Nir2 and Nir3) function in intact cells
2. The function of different structural domains of VAP proteins, and their potential role in regulating cellular lipid homeostasis and membrane trafficking
3. The function of VAP-A and/or –B in specialized secretory cells, such as the insulin-producing pancreatic β cells
4. The underlying mechanisms of VAP-B(P56s)-induced MNDs