Serum provides a large source of choline phospholipids that animal cells can use for the assembly of membranes. We propose to ezamine the uptake and metabolism of serum phospholipids by Chinese hamster ovary (CHO) cells grown in tissue culture in order to elucidate how animal cells coordinate the salvage of serum phospholipids with the formation of choline lipids by de novo synthesis. Already, we have established conditions that permit serum lysoecithin, enriched in lipoprotein-deficient serum or added to delipidated serum, satisfies the choline requriement of CHO cells. We will study the mechanism of lysolecithin uptake by defining its chemical specificity and by examining the role played by serum proteins. We will examine the detail the metabolism of the added lysolecithins, quantitating choline metabolite levels in cells and growth medium. We will determine whether the addition of lysolecithin affects the de novo synthesis of lecithin by measuring eh acitivites of the biosynthetic enzymes. Unlike wild-type CHO cells, recently isolated CHO cell mutants utilize low density lipoprotein (LDL) as the sole source of dietary choline in medium containing delipidated serum. This observation should permit us to test the hypothesis that LDL phosphlipid is mostly degraded inside lysosomes. We will examine if some of the phospholipid is utilized intact for membrane assembly, and we will examine the type and quntiti of choline metabolites derived from lipoprotein metabolism. We will also develop strategies for isolating other CHO cell mutants in order to define the genes and gene products that mediate LDL phospholipid metabolism. Together, these studies will shed light on the regulation of choline phospholipid metabolism. Together, these studies will shed light on the regulation of choline phospholipid metabolism in animal cells and will provide a foundation for future studies of phospholipid assembly into membranes. Furthermore, this information may suggest how the metabolism of serum phospholipids affects the onset and progression of human diseases which result from the abnormal deposition of lipids, such as atherosclerosis and in-born errors of lipid metabolism.
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