Targeting sequences of UBXD8 and AAM-B reveal that the ER has a direct role in the emergence and regression of lipid droplets

Abstract

Lipid droplets are sites of neutral lipid storage thought to be actively involved in lipid homeostasis. A popular model proposes that droplets are formed in the endoplasmic reticulum (ER) by a process that begins with the deposition of neutral lipids between the membrane bilayer. As the droplet grows, it becomes surrounded by a monolayer of phospholipid derived from the outer half of the ER membrane, which contains integral membrane proteins anchored by hydrophobic regions. This model predicts that for an integral droplet protein inserted into the outer half of the ER membrane to reach the forming droplet, it must migrate in the plane of the membrane to sites of lipid accumulation. Here, we report the results of experiments that directly test this hypothesis. Using two integral droplet proteins that contain unique hydrophobic targeting sequences (AAM-B and UBXD8), we present evidence that both proteins migrate from their site of insertion in the ER to droplets that are forming in response to fatty acid supplementation. Migration to droplets occurs even when further protein synthesis is inhibited or dominant-negative Sar1 blocks transport to the Golgi complex. Surprisingly, when droplets are induced to disappear from the cell, both proteins return to the ER as the level of neutral lipid declines. These data suggest that integral droplet proteins form from and regress to the ER as part of a cyclic process that does not involve traffic through the secretory pathway.

Identification of Ubxd8 protein as a sensor for unsaturated fatty acids and regulator of triglyceride synthesis

“…Ubxd8 is known to be an intrinsic membrane protein (16). To study the interactions of FAs with Ubxd8 we first sought to prepare a soluble form of the protein free of membrane lipids.…”

Murine Diacylglycerol Acyltransferase-2 (DGAT2) Can Catalyze Triacylglycerol Synthesis and Promote Lipid Droplet Formation Independent of Its Localization to the Endoplasmic Reticulum

“…DGAT2 has a potential proline knot with 5 proline residues from amino acids 329 and 350. Other proteins, including perilipin, caveolin, AAM-B, and viperin are directed to lipid droplets by distinct hydrophobic sequences (33,(63)(64)(65). DGAT2 has a moderately hydrophobic region between amino acids 230 and 250 that may interact with the lipid droplet surface (20).…”

“…The first is a relatively hydrophobic region between amino acids 232 and 250. Other studies have demonstrated that certain hydrophobic regions can serve as lipid droplet targeting signals (33)(34)(35). The second region is between amino acids 327 and 350, which contains four proline residues that could be part of a proline knot motif.…”

Not Just Fat: The Structure and Function of the Lipid Droplet

“…The bidirectional movement of some proteins between LDs and the ER membrane suggests a close structural relationship (Pol et al 2005;Turro et al 2006;Zehmer et al 2009). However, it has not been shown unequivocally that normal LDs retain membrane continuity with the ER as plastoglobules do with the thylakoid membrane ( Fig.…”