Structure and Functional Analysis of Bone Morphogenetic Protein Receptor

我们利用在TGF-β家族受体如激活素和TGF-β II型受体的保守序列中设计的寡核苷酸引物，从小鼠MC 3 T3-E1细胞cDNA中分离了几个新的受体Ser/Thr激酶cDNA。其中一个被命名为mTFR 11的受体被证明是I型受体，它结合BMP-2和BMP-4。我们构建了一个mu受体cDNA，预测一个截短的受体缺乏整个丝氨酸/苏氨酸激酶，并证实，显性负性受体抑制BMP-2和BMP-4在培养的成骨细胞和早期胚胎。此外，我们通过GS结构域中Gln到Asp的点突变产生了BMP受体的组成型活性形式，这被认为是BMP信号传导所必需的。我们进一步证实了该突变体受体不依赖BMP配体而传递BMP样信号。因此，我们利用BMP受体的胞浆区作为诱饵，通过酵母双杂交方法筛选了人cDNA文库。我们成功地分离出了三个编码BMP受体结合蛋白的cDNA，目前正在研究这些蛋白在信号转导中的功能。

We have isolated several novel receptor Ser/Thr kinase cDNAs from a mouse MC3T3-E1 cells cDNA using oligonucleotide primers designed in the conserved sequence among TGF-beta family receoptors such as activin and TGF-beta type II receptors. One of them designated as mTFR11turned out to be a type I class of receptor that binds both BMP-2 and BMP-4. We constructed a mutantreceptor cDNA that predicts a truncated receptor lacking entire Ser/Thr kinase and confirmed that the dominant negative receptor inhibited both BMP-2 and BMP-4 in cultured osteoblastic cells and in early embryos. Moreover, we generated constitutively active form ofthe BMP receptor by a point mutation Gln to Asp in GS domain, which is believed to be essential for BMP signaling. We further confirmed that the mutant receptor transmits BMP-like signals independent of BMP ligands.Our effort is now focused on the identification of downstream factor that mediates BMP signaling. Therefore, we have screened a human cDNA library by yeast two-hybrid method using a cytoplasmic region of the BMP receptor as a bait. Successfully, we have isolated three cDNAs that encodes proteins that binds BMP receptor and function of the proteins in the signal transduction is currently investigated.