Id Genes Are Direct Targets of Bone Morphogenetic Protein Induction in Embryonic Stem Cells

Abstract

Bone morphogenetic proteins (BMPs) are morphogenetic signaling molecules essential for embryonic patterning. To obtain molecular insight into the influence of BMPs on morphogenesis, we searched for new genes directly activated by BMP signaling. In vitro cultured mouse embryonic stem (ES) cells were used, cultivated in chemically defined growth medium (CDM). CDM-cultured ES cells responded very selectively to stimulation by various mesoderm inducers (BMP2/4, activin A, and basic fibroblast growth factor). BMP2/4 rapidly induced transcript levels of the homeobox genes Msx-1 and Msx-2 and the proto-oncogene JunB, whereas c-jun transcripts displayed delayed albeit prolonged increase. Using differential display cDNA cloning, six direct BMP target genes were identified. These include Id3, which showed strong mRNA induction, and the moderately induced Cyr61, DEK, and eIF4AII genes, as well as a gene encoding a GC-binding protein. Besides Id3, also the Id1 and Id2 genes were activated by BMP4 in both ES cells and a range of different cell lines. Id genes encode negative regulators of basic helix-loop-helix transcription factors. In vivo we observed local ectopic expression of Id3 and Msx-2 mRNAs in Ft/+ embryos at overlapping regions of ectopic Bmp4 misexpression. We therefore propose that the Msx and Id genes are direct target genes of embryonic BMP4 signaling in vivo.