Fig. 1. A functional truncated protein is encoded by the Bmpr2^E2 allele. (A) Targeting strategy. Exon 2 of Bmpr2 was replaced by a neomycin resistance cassette. A, AatII; B, BamHI; Bg, BglII; E, EcoRI; Rv, EcoRV; K, KpnI; X, XbaI; Xh, XhoI. (B) Southern blot showing genotyping of pups with the internal probe. (C) Amino acid sequence comparison of the extracellular domains of mouse ActRII, TGFßRII and Bmpr2. The region encoded by exon 2 is boxed and conserved residues involved in 3D-structure formation are in red. This includes three out of seven cysteine residues, which are conserved among all type II receptors, that are required to maintain the conformation of the extracellular domain, and two out of five hydrophobic amino acids thought to participate in the hydrophobic interactions that stabilize the structure. (D) Structure of the Bmpr2 protein. Exon 1 encodes the initiator methionine. Exons 2 and 3 each encode half of the extracellular ligand-binding domain. Exon 4 encodes the transmembrane domain. The serine threonine kinase domain is encoded by exons 5-11. Exons 12-13 encode an intracytoplasmic tail of unknown function. Deletion of exon 2, which is in frame, should generate a truncated protein. (E) RT-PCR on RNA from wild type (+/+), heterozygous (+/-) and E2 mutants (-/-) with primers encompassing the deletion (exons 1-5) and downstream of the deletion (exons 3-5) shows that the Bmpr2^E2 allele is transcribed. (F) Transfection of the mutant or wild-type Bmpr2 cDNA expression plasmid confers BMP responsiveness to Mv1Lu cells. BMP signaling activity was assayed with a reporter construct that contains luciferase under the control of a BMP-responsive element from the promoter of the Msx2 gene (Daluiski et al., 2001; Liu et al., 1994).

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