Sterol regulatory element-binding protein negatively regulates microsomal triglyceride transfer protein gene transcription

We herein report that mRNA expression of microsomal triglyceride transfer p rotein (MTP) and its protein synthesis decline in response to sterol deplet ion in HepG2 cells, and we functionally characterized the MTP gene promoter in an effort to investigate the molecular mechanisms by which MTP gene tra nscription is regulated. Luciferase assays using truncated versions of the reporter gene revealed that the region at -124 to +33 base pairs of the hum an promoter contains the elements required for the suppression of transcrip tion by sterol depletion. Enforced expression of an active form of sterol r egulatory element-binding protein (SREBP)-1 (amino acids 1-487) or -2 (amin o acids 1-481), both of which are activated under sterol-depleted condition s, is able to mimic sterol-mediated down-regulation. Either further truncat ion of the promoter region or mutation of the putative SREBP-binding sequen ce (5'-GCAGCCCAC-3', -124 to -116 base pairs) abolishes the sterol- and SRE BP-dependent transcriptional regulation. Gel mobility shift assay showed th at recombinant SREBP-2-(1-481) is able to bind the sequence. Enforced expre ssion of a truncated form of SREBP-2 (amino acids 31-481), which acts as an inhibitor of transcription of the low density lipoprotein receptor gene be cause it lacks the transcriptional activation domain, also diminishes the l uciferase activity, suggesting that direct binding to the promoter region m ight be sufficient and that the mechanism by which SREBPs inhibit MTP gene expression is distinct from that for the transcriptional stimulation of ste rol-regulated genes. Although the SREBP-binding site overlaps a negative in sulin-responsive element, insulin negatively regulates MTP gene expression even when the amount of the active form of SREBPs is quite low under the st erol-loaded conditions, indicating that SREBPs only slightly mediate, if at all, the insulin effects. Overall, we conclude that SREBPs are responsible for regulation of lipoprotein secretion via their control of MTP gene expr ession. Moreover, our results describe for the first time a novel mechanism by which SREBPs negatively regulate expression of the gene encoding the pr otein involved in lipid metabolism.