Sensing BMP Pathway Activity by Immune Detection of Phosphorylated R-Smad Proteins in Mouse Embryonic Kidney

At the onset of mammalian kidney development, the ureteric bud invades the surrounding metanephric mesenchyme, and genetic studies in the mouse have shown that BMP pathway activity has to be antagonized in the vicinity of the epithelium, a task performed by the secreted BMP antagonist Grem1. Here, we describe a short protocol that allows for detection of the pattern of BMP canonical signal transduction by using antibodies that specifically recognize the phosphorylated forms of R-Smad proteins (Smad1, Smad5, and Smad8), which provides a way to monitor overall pathway activity in the mammalian embryonic kidney.