Nuclear Factor-Y is an adipogenic factor that regulates leptin gene expression

Objective: Leptin gene expression is highly correlated with cellular lipid content in adipocytes but the transcriptional mechanisms controlling leptin expression in vivo are poorly understood. In this report, we set out to identify cis- and trans-regulatory elements controlling leptin expression.

Methods: Leptin-BAC luciferase transgenic mice combining with other computational and molecular techniques were used to identify transcription regulatory elements including a CCAAT-binding protein Nuclear Factor Y (NF-Y). The function of NF-Y in adipocyte was studied in vitro with 3T3-L1 cells and in vivo with adipocyte-specific knockout of NF-Y.

Results: Using Leptin-BAC luciferase mice, we showed that DNA sequences between -22 kb and +8.8 kb can confer quantitative expression of a leptin reporter. Computational analysis of sequences and gel shift assays identified a 32 bp sequence (chr6: 28993820-2899385) consisting a CCAAT binding site for Nuclear Factor Y (NF-Y) and this was confirmed by a ChIP assay in vivo. A deletion of this 32 bp sequence in the -22 kb to +8.8 kb leptin-luciferase BAC reporter completely abrogates luciferase reporter activity in vivo. RNAi mediated knockdown of NF-Y interfered with adipogenesis in vitro and adipocyte-specific knockout of NF-Y in mice reduced expression of leptin and other fat specific genes in vivo. Further analyses of the fat specific NF-Y knockout revealed that these animals develop a moderately severe lipodystrophy that is remediable with leptin therapy.

Conclusions: These studies advance our understanding of leptin gene expression and show that NF-Y controls the expression of leptin and other adipocyte genes and identifies a new form of lipodystrophy.