Combined analysis of oligonucleotide microarray data from transgenic and knockout mice identifies direct SREBP target genes

Abstract

The synthesis of fatty acids and cholesterol, the building blocks of membranes, is regulated by three membrane-bound transcription factors: sterol regulatory element-binding proteins (SREBP)-1a, -1c, and -2. Their function in liver has been characterized in transgenic mice that overexpress each SREBP isoform and in mice that lack all three nuclear SREBPs as a result of gene knockout of SREBP cleavage-activating protein (SCAP), a protein required for nuclear localization of SREBPs. Here, we use oligonucleotide arrays hybridized with RNA from livers of three lines of mice (transgenic for SREBP-1a, transgenic for SREBP-2, and knockout for SCAP) to identify genes that are likely to be direct targets of SREBPs in liver. A total of 1,003 genes showed statistically significant increased expression in livers of transgenic SREBP-1a mice, 505 increased in livers of transgenic SREBP-2 mice, and 343 showed decreased expression in Scap-/- livers. A subset of 33 genes met the stringent combinatorial criteria of induction in both SREBP transgenics and decreased expression in SCAP-deficient mice. Of these 33 genes, 13 were previously identified as direct targets of SREBP action. Of the remaining 20 genes, 13 encode enzymes or carrier proteins involved in cholesterol metabolism, 3 participate in fatty acid metabolism, and 4 have no known connection to lipid metabolism. Through application of stringent combinatorial criteria, the transgenic/knockout approach allows identification of genes whose activities are likely to be controlled directly by one family of transcription factors, in this case the SREBPs.

Fig. 1.

SREBP-2 mediated activation of the entire pathway for cholesterol synthesis in mammalian cells. Key biosynthetic intermediates are highlighted in yellow. Red italics denotes the fold increase in mRNA levels for each enzyme in TgSREBP-2 liver. Each value is the average of the two microarray hybridizations listed in Tables 1 and 3. *, Desmosterol reductase was not represented on the microarray. The fold-increase value is from real-time PCR. †,17β-Hydroxysteroid dehydrogenase-7 has not been confirmed as a cholesterol biosynthetic enzyme.

Fig. 2.

SREBP-1a mediated activation of pathways for monounsaturated and polyunsaturated fatty acid synthesis in mammalian cells. Fatty acid products are highlighted in yellow. Red italics denotes the fold increase in mRNA levels for each enzyme in TgSREBP-1a liver. Each value is the average of the two microarray hybridizations listed in Tables 1 and 2 except as indicated. *, Results are from real-time PCR.