Charles E. Sparks, M.D.

Apo B exists as B48 and B100 produced through a novel posttranscriptive mechanism of apo B mRNA editing producing protein products of differing size with differing metabolisms. The goal of our research is to understand apo B triglyceride-rich lipoprotein (TRL) assembly contrasting B100 and B48 synthetic pathways. McArdle RH-7777 cells (McA) have provided many useful insights into inherent differences in B100 and B48 in TRL biogenesis. Recent studies indicate that B100-TRL assembly occurs by lipidation of conformationally "acceptable" B100. B48 undergoes initial lipidation similar to B100 followed by neutral lipidation of the precursor high density lipoprotein particle (B48-HDL) to form B48-TRL. More than 70% of the triglyceride (TG) fatty acids (FA) arise from cytosolic, stored TG through a process involving lipolysis/reesterification. Lipolyzed FAs, possibly through acylation reactions with PL intermediates, traverse the ER membrane by unknown means prior to fusion with B48-HDL. Unlike B100, that is synthesized and rapidly secreted by rat hepatocytes (RH), there is a kinetically distinct pool of B48 that has a long cellular retention time. We hypothesize that this pool may provide a ready supply of B48-HDL as a precursor for the rapid assembly of TRL via a later assembly step. In cells that make predominantly B48, such as intestine, the cellular pool of B48 allows TRL secretion to occur without a requirement for de novo apo B synthesis. We recently identified a novel protein factor, an apo B secretion enhancer (BSE), whose expression correlates with apo B mRNA abundance in McA cells transfected with BSE. Apo B mRNA abundance, however, does not correspond with a proportional increase in apo B secretion. Studies explore the role of BSE in the observed changes in apo B mRNA stability/transcription. Apo B degradation studies address the non-proportional secretory rate. A phospholipid (PL)-dependent pathway for the stimulation of apo B secretion by BSE is suggested as BSE is homologous with betaine homocysteine methyltransferase (BHMT), a methylation enzyme. An hypothesis explored is that, apart from its ability to stabilize apo B mRNA, an ER form of BSE may provide newly synthesized phosphatidylcholine (PC) via phosphatidylethanolamine (PE) methylation of phosphatidylserine (PS) and increased supply of PL may be a mechanism for the enhancing secretion of apo B.

Importance of the research is related to the role of apo B in lipid transport and as a factor associated with risk of arterial disease. Additional research focuses on the role of apo B as a risk factor in developing heart attacks, strokes, and in recurrent heart attacks in human populations.