Fig. 8. SCD1 is naturally short-lived and is constitutively degraded irrespective of cellular lipid levels that regulate SCD1 gene expression. (A) HEK-293 cells (untransfected) were grown for 36 hours in standard culture medium containing untreated FBS (delipidated, -) or delipidated FBS (delipidated, +). Total RNAs were isolated from the cells and the mRNA amount for the indicated proteins was determined by real-time quantitative PCR. Levels of mRNAs are shown relative to the amounts of mRNA from the cells cultured in untreated FBS-containing medium. The GAPDH gene was used as a negative control. (B) HEK-293 cells were transfected with plasmid carrying SCD1-HA cDNA, grown for 48 hours, and subjected to pulse-chase analysis with [³⁵S]methionine under the indicated conditions. At the indicated chase times, cells were solubilized and subjected to immunoprecipitation with anti-HA antibodies. The immunoprecipitates were analyzed by SDS-PAGE and subsequent digital autoradiography. (C) Band intensities in B were quantified. Relative radioactivity is shown with the levels at time zero defined as 100%. The results shown are the average of three independent experiments.