Possible role of the Golgi apparatus in the assembly of very low density lipoprotein.

Abstract

Transit of newly synthesized triacyl[3H]-glycerol through organelles of the secretory system leading to assembly into nascent very low density lipoproteins (VLDLs) or to cytoplasmic storage was investigated in chick hepatocytes. Cells in monolayer culture were pulse-labeled with [2-3H]glycerol, and after different periods of chase with unlabeled glycerol, the movement of triacyl[3H~glycerol through the endoplasmic reticulum (ER) and Golgi and the incorporation into nascent VLDL and cytoplasmic triacylglycerol-rich vesi- Very low density lipoprotein (VLDL), secreted by the hepatocyte, is a major carrier of triacylglycerol in the bloodstream of higher animals. The VLDL particle consists of a neutral hydrophobic core of triacylglycerol and cholesterol ester surrounded by an amphipathic shell of phospholipid, cholesterol, and specific apolipoproteins. Each ofthese components is synthesized by a distinct membrane-bound enzyme system associated with the endoplasmic reticulum (ER) (1, 2). The apolipoproteins, like other secretory proteins, are synthesized and cotranslationally translocated through the membrane of the ER into its lumen (3,4). Since the lipid components of VLDL are synthesized by enzymes situated on the cytoplasmic face of the ER (2), the question is raised as to how these lipids cross the ER membrane to gain access to the lumenal apolipoproteins with which they must assemble? Also unknown is the site within the secretory system at which this assembly process occurs.Recently we determined that three apolipoproteins (apoB, apoII, and apoA-I) of nascent VLDL move through the ER of the chick hepatocyte at markedly different rates (5). In each case transit through the Golgi was found to be ratelimiting for the overall process of apolipoprotein secretion. Thus, it can be concluded that these apolipoproteins do not associate with each other or with the same particle during movement through the ER; apoB and apoll are hydrophobic and difficult to dissociate from the triacylglycerol core of VLDL once this association has occurred (6, 7). It would be expected, therefore, that upon binding to the nascent triacylglycerol particle, these apolipoproteins would remain tightly bound to this lipid during the remainder of their transit through the secretory system. Taken together, these observations and the fact that the apolipoproteins traverse the Golgi at comparably slow rates (5) Cells were scraped into SH buffer containing protease inhibitors (5) and then were homogenized by 10 passages through the ball-bearing homogenizer (5) described by Balch and Rothman (10). One-half milliliter of 0.22 M sucrose was layered over the cell homogenate, and the sample was centrifuged for 10 min at 10,000 x g. Nuclei and debris from the cell homogenate were in the pellet, whereas cytoplasmic triacylglycerol-rich vesicles (TGRVs; see ref. 11) were in the Abbreviations: VLDL, very low density lipoprotein; ER, endoplasmic reticulum; TGRVs, triacylglycerol-rich vesicles; apoB, apoll, and apoA-I, apolipoproteins B, II, and A-I.

N-3 fatty acids stimulate intracellular degradation of apoprotein B in rat hepatocytes.

“…There are divergent views on such matters as the site of initial lipid transfer (the rough endoplasmic reticulum [31,32] vs. the smooth endoplasmic reticulum [25,33]) and the organelle in which apoB resides the longest (endoplasmic reticulum [15,34] vs. Golgi apparatus [35,36]); however, in recent reports there is good evidence that during translation, apoB is not completely translocated across the endoplasmic reticulum membrane and remains membrane-bound (31,32,37-39). Presumably, this apoB could be targeted to either a lipoprotein assembly/secretion pathway or to degradation.…”

CideB Protein Is Required for the Biogenesis of Very Low Density Lipoprotein (VLDL) Transport Vesicle

“…Moreover, it has been proposed that additional triglycerides are added to the nascent VLDL in the Golgi lumen (11,(15)(16)(17). Based on a number of biochemical and histological data, it has been suggested that Golgi is the site of VLDL maturation; however, this supposition is still a subject of debate (11)(12)(13)(14)(15)(16)(17)(18)(19)(20). Regardless of their maturation site, the transport of nascent VLDL particles from the ER to the Golgi is imperative and determines the rate of VLDL secretion from the liver (10).…”

“…Their structural protein, apolipoprotein B100 (apoB100), gets further glycosylated and phosphorylated (12)(13)(14). Moreover, it has been proposed that additional triglycerides are added to the nascent VLDL in the Golgi lumen (11,(15)(16)(17). Based on a number of biochemical and histological data, it has been suggested that Golgi is the site of VLDL maturation; however, this supposition is still a subject of debate (11)(12)(13)(14)(15)(16)(17)(18)(19)(20).…”

Substrate delivery as a determinant of hepatic apoB secretion.

“…The evidence that VLDL particles can be larger in the Golgi than in the endoplasmic reticulum has been noted above. 45 - 46 If cholesterol can move directly from the lysosome to the Golgi and then promptly leave the cell, it would never come into equilibrium with the critical sites within the cell at which LDL receptor synthesis is determined. As well, cholesterol that does penetrate the interior compartments of the cell more fully can stimulate increased apoB secretion, and so cholesterol export from the cell can also increase by virtue of this effect.…”