Liver-Specific Silencing of the Human Gene Encoding Succinyl-CoA: 3-Ketoacid CoA Transferase

Abstract

The human succinyl-CoA: 3-ketoacid CoA transferase (SCOT) gene encodes the ketolytic enzyme that functions in the mitochondrial matrix. The activation of acetoacetate to acetoacetyl-CoA by SCOT is essential for the use of ketone bodies as an energy source. The ketolytic capacity of tissues is proportional to their level of SCOT activity. Normal hepatocytes, the site of ketone body synthesis, have no detectable SCOT protein. The absence of SCOT in hepatocytes is an important element in energy metabolism, suppressing ketolysis in the liver. To study the tissue-specific silencing of SCOT expression, we analyzed the promoter function of SCOT gene in three different human cell lines. Immunoblot analysis showed that SCOT protein was detectable in HeLa cervical cancer cells and Chang liver cells. However, SCOT protein was not detected in HepG2 hepatoma cells and liver tissues, indicating that HepG2 hepatoma cells maintain the characteristics of liver cells in the ketone body metabolism. Luciferase reporter assays in HeLa and Chang liver cells showed that the 361-bp proximal region of the SCOT gene was responsible for the basal promoter activity and contained two GC boxes, each of which was bound in vitro by Sp1, a ubiquitously expressed transcription factor. These results suggest that these GC boxes may be important for SCOT gene expression. Moreover, the region between -2168 and -361 appeared to inhibit the SCOT promoter activity in HepG2 cells. Thus, liver-specific silencing of the SCOT gene expression may be mediated in part by its 5'-flanking sequence.