The effect of C/EBPα on the expression of LIPE gene encoding hormone-sensitive lipase/cholesteryl esterase (HSL) was investigated in Y-1 CCL79 cells.
It was found that transfection of these cells with the vector overexpressing C/EBPα increased both the level of LIPE transcript, measured by RT-qPCR, and the luminesce emitted by luciferase reporter gene fused to the −2150 fragment of LIPE promoter.
Activation of adenylyl cyclase by forskolin resulted in 2.5-fold increase in the intensity of luminescence and over 3-fold increase in luminescence was observed when the cells were cotransfected with the vector overexpressing C/EBP. The incubation of C/EBP-cotransfected cells with forskolin caused over 6-fold increase in the intensity of luminescence, suggesting that the effects of C/EBPα and forskolin are additive.
The analysis of sequence of the proximal LIPE promoter showed multiple binding sites for various transcription factors including C/EBPα site, which is located between nucleotides −46 bp and −59 bp.
When the Y-1 cells were transfected with the recombinant vector containing −60 bp fragment of LIPE promoter fused to the luciferase reporter gene and were cotransfected with the vector overexpressing C/EBPα, the luminescence increases about 9-fold indicating that C/EBPα stimulates the expression of LIPE by reacting with its response element.
The results indicate that C/EBPα stimulates the expression of LIPE independently of the PKA pathway by binding to a response element situated within the −60 bp fragment of LIPE promoter. This suggests that C/EBPα might be involved in the regulation of LIPE expression and thus cholesterol supply for steroid hormone synthesis.