VLDL exits from the endoplasmic reticulum in a specialized vesicle, the VLDL transport vesicle, in rat primary hepatocytes

Abstract

The movement of VLDL [very-LDL (low-density lipoprotein)] from the ER (endoplasmic reticulum) to the Golgi is required for its eventual secretion from hepatocytes and represents a potential target in controlling elevated concentrations of its metabolite LDL, the major determinant of atherosclerosis. To study this process, an in vitro ER-budding assay was developed to examine the generation of the VTV (VLDL transport vesicle) and PTV (protein transport vesicles) using ER isolated from [(14)C]TAG (triacylglycerol) and [(3)H]protein-labelled primary rat hepatocytes. VTVs do not contain albumin, as determined by immunoblots. VTVs were distributed in light-density fractions, whereas PTVs were mainly in the mid-portion of the sucrose gradient. Electron microscopy revealed that VTVs were larger ( approximately 100-120 nm) in size than PTVs ( approximately 55-70 nm). ER from 0.4 mM OA (oleic acid)-treated hepatocytes budded VTVs of a lighter density as compared with VTVs budded from ER of 0.1 mM or 0.004 mM OA-treated hepatocytes. The generation of VTVs from rat hepatic ER required cytosol, ATP, Sar1 (a GTPase) and incubation at 37 degrees C. Proteinase K treatment did not degrade the VTV cargo protein, apoB100 (apolipoprotein 100), indicating that VTVs were sealed. Immunoblots showed that VTV concentrated apoB100, Sar1 and rSec22b, and excluded albumin and calnexin. VTVs were shown to fuse with cis-Golgi and delivered their cargo to the Golgi lumen, as determined by in vitro fusion, and acquired endoglycosidase H resistance. These results suggest that a new ER-derived transport vesicle (VTV) has been identified and characterized which transports nascent VLDL from the hepatic ER to the Golgi.

Role of Phosphatidylinositol 4-Phosphate (PI4P) and Its Binding Protein GOLPH3 in Hepatitis C Virus Secretion

“…The occurrence of VLDL-associated viral particles (lipoviroparticles) in patient sera further strengthen the notion that HCV and VLDL secretion overlap and that HCV co-opts the VLDL secretory pathway for its secretion (50). Although the intricate details of VLDL biogenesis are still unclear, it is well known that the VLDL particles traffic through the Golgi compartments, implicating a similar pathway for HCV virion egress (19,51).…”

“…These * This work was supported, in whole or in part, by National Institutes of Health pre-VLDL particles are subjected to additional lipidation to generate a mature VLDL particle. Although the mechanism and location of this second lipidation event is not clear, it is understood that the VLDL particles are transported from the ER to the Golgi in specialized vesicles (VLDL transport vesicles) and are eventually secreted via the Golgi secretory pathway (19). It has been shown that perturbation of VLDL biogenesis inhibits HCV virion secretion (3,11,12).…”

Hepatitis c virus and host cell lipids: An intimate connection

“…166 Mature VLDL is transported to the Golgi and finally secreted. 163 By using mass spectrometry analyses of affinity purified HCV particles produced in cell culture, it was found that the lipid ©2 0 1 1 L a n d e s B i o s c i e n c e .…”

“…LDs and luLDs are important for the transfer of TGs to VLDL2. 163 They are composed of a core of neutral lipids and surrounded by a monolayer lipid membrane. Although LDs and luLDs share a very similar lipid composition, they are decorated with different sets of proteins, regulating their size, mobility and localization: 152 ADRP and the 47 kDa tail interacting protein (TIP47) are found on the surface of LDs, whereas MTP, ApoE and TG hydrolase cofractionate with luLDs, but not with LDs.…”

Intestinal lipid absorption

“…Thus, chylomicrons are transported on unique particles, different from those used for protein transport, by intestinal cells. Similarly, unique particles have been shown to transport hepatic lipoproteins in the liver (164).…”