Inhibition of p38 mitogen-activated protein kinase enhances adrenergic-stimulated arylalkylamine N-acetyltransferase activity in rat pinealocytes.

We have previously shown that inhibition of p38(MAPK) increases adrenergic-stimulated p42/44(MAPK) activation in rat pinealocytes. In this study we investigated whether p38(MAPK) played a role in the adrenergic regulation of arylalkylamine-N-acetyltransferase (AA-NAT) induction and melatonin (MT) synthesis. Treatment of pinealocytes with norepinephrine (NE) caused a time-dependent increase in the levels of AA-NAT mRNA, AA-NAT protein, and enzymatic activity as well as MT production.

Microsomal triacylglycerol transfer protein is required for lumenal accretion of triacylglycerol not associated with ApoB, as well as for ApoB lipidation.

The assembly of very low density lipoproteins in hepatocytes requires the microsomal triacylglycerol transfer protein (MTP). This microsomal lumenal protein transfers lipids, particularly triacylglycerols (TG), between membranes in vitro and has been proposed to transfer TG to nascent apolipoprotein (apo) B in vivo. We examined the role of MTP in the assembly of apoB-containing lipoproteins in cultured murine primary hepatocytes using an inhibitor of MTP.

The assembly and secretion of apolipoprotein B-48-containing very low density lipoproteins in McA-RH7777 cells.

We have used an extraction procedure, which released membrane-bound apoB-100, to study the assembly of apoB-48 VLDL (very low density lipoproteins). This procedure released apoB-48, but not integral membrane proteins, from microsomes of McA-RH7777 cells. Upon gradient ultracentrifugation, the extracted apoB-48 migrated in the same position as the dense apoB-48-containing lipoprotein (apoB-48 HDL (high density lipoprotein)) secreted into the medium.

Assembly of very low density lipoprotein: a two-step process of apolipoprotein B core lipidation.

The liver plays a primary role in lipid metabolism. Important functions include the synthesis and incorporation of hydrophobic lipids, triacylglycerols and cholesteryl esters into the core of water-miscible particles called lipoproteins and the secretion of these particles into the circulation for transport to distant tissues. In this article, we present a brief overview of one aspect of the assembly process of very low density lipoproteins, namely, possible mechanisms for combining core lipids with apolipoprotein B.

The microsomal triglyceride transfer protein catalyzes the post-translational assembly of apolipoprotein B-100 very low density lipoprotein in McA-RH7777 cells.

In cells in which the lipoprotein assembly process had been inactivated by brefeldin A (BFA), membrane-associated apoB-100 disappeared without forming lipoproteins or being secreted, indicating that it was degraded. Reactivation of the assembly process by chasing the cells in the absence of BFA, gave rise to a quantitative recovery of the membrane-associated apoB-100 in the very low density lipoprotein (VLDL) fraction in the medium. These results indicate that the membrane-associated apoB-100 can be converted to VLDL.

Brefeldin A reversibly inhibits the assembly of apoB containing lipoproteins in McA-RH7777 cells.

BFA inhibited in a dose dependent way the assembly of apoB-48 very low density lipoprotein (VLDL) but allowed a normal rate of biosynthesis of the apolipoprotein and of the assembly of the dense ("high density lipoprotein (HDL)-like") apoB-48 particle (apoB-48 HDL). The inhibition of the assembly of apoB-48 VLDL occurred at BFA levels that allowed a major secretion of both transferrin and apoB-48 HDL. The assembly of apoB-100 containing lipoproteins was also inhibited by BFA but could be reactivated by a 30-60 min chase in the absence of BFA, which agreed with the time that was estimated to be needed to restore the secretory pathway (approximately 60 min).

Studies on the assembly of apolipoprotein B-100- and B-48-containing very low density lipoproteins in McA-RH7777 cells.

The mechanisms by which apolipoprotein B-100 (apoB-100) and apoB-48 assemble lipoproteins have been studied in the McA-RH7777 cell line. After incubation for 2 h with 360 microM oleic acid, the McA-RH7777 cells secreted apoB-100 mainly on very low density lipoprotein (VLDL) particles, as judged from sucrose gradient and sequential ultracentrifugation. ApoB-48 was secreted on both VLDL and on denser, high density lipoprotein (HDL)-like lipoproteins.

Purification of diacylglycerol:acyltransferase from rat liver to near homogeneity.

A method to isolate a protein related to the diacylglycerol:acyltransferase (DGAT) activity in rat liver microsomes has been developed. The microsomes were treated with sodium deoxycholate (DOC; 0.1 mg/mg protein) at a concentration of 1 mM, i.

Influence of triacylglycerol biosynthesis rate on the assembly of apoB-100-containing lipoproteins in Hep G2 cells.

Apolipoprotein B-100 (apoB-100) appears in three forms in the endoplasmic reticulum of Hep G2 cells: (1) tightly bound to the membrane, ie, not extractable by sodium carbonate. This form is glycosylated but protease sensitive when present in intact microsomes, suggesting that it is only partially translocated to the microsomal lumen; (2) extractable by sodium carbonate and present on low-density lipoprotein-very-low-density lipoprotein (LDL-VLDL)-like particles. This form is glycosylated and secreted into the medium; and (3) extractable by sodium carbonate but having a higher density than the LDL-VLDL-like particles.