Original Article

Expression and intracellular processing of the 58 kDa sterol carrier protein-2/3-oxoacyl-CoA thiolase in transfected mouse L-cell fibroblasts

Correspondence to: Friedhelm Schroeder

Although the sterol carrier protein 2 (SCP-2) gene encodes for two proteins, almost nothing is known of the function and potential processing of the larger transcript corresponding to the 58 kDa sterol carrier protein-2/3-oxoacyl-CoA thiolase (SCP-x), in intact cells. L-cell fibroblasts transfected with cDNA encoding for the 58 kDa SCP-x protein had a 4.5-fold increase in SCP-x mRNA transcript levels. Western blot analysis showed SCP-x protein expression reached 0.011% of total protein, representing a 4.1-fold increase over basal levels. Surprisingly, the 13.2 kDa SCP-2 protein also increased 2-fold in the transfected cells. This was consistent with part of the 58 kDa SCP-x being proteolytically processed to 13.2 kDa SCP-2 as there was no evidence of an mRNA transcript corresponding to a 13.2/15.2 kDa gene product in the transfected L-cell clones. Confocal immunofluorescence microscopy of transfected L-cells showed that SCP-x/SCP-2 co-localized in highest concentration with catalase in peroxisomes, but significant amounts appeared extra-peroxisomal. Overexpression of SCP-x significantly altered cholesterol uptake and metabolism. Uptake of exogenous [³H]cholesterol and total cholesterol mass were increased 1.9- and 1.4-fold, respectively, in SCP-x expressors. Although cholesterol ester mass was unaltered, incorporation of exogenous [³H]cholesterol and [³H]oleic acid into cholesteryl esters increased 2.3- and 2.5-fold, respectively.

These results from intact cells suggest the 13.2 kDa SCP-2 can arise from the larger SCP-2 gene product and indicate a role for the 58 kDa SCP-x protein in cholesterol uptake and intracellular cycling.—Atshaves, B. P., A. D. Petrescu, O. Starodub, J. B. Roths, A. B. Kier, and F. Schroeder. Expression and intracellular processing of the 58 kDa sterol carrier protein-2/3-oxoacyl-CoA thiolase in transfected mouse L-cell fibroblasts. J. Lipid Res. 1999. 40: 610 – 622.

Supplementary key words: sterol carrier protein-2, sterol carrier protein-2/3-oxoacyl-CoA thiolase, L-cell fibroblast, cholesterol trafficking

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