Introduction
The BMPs are members of the transforming growth factor-
(TGF-) superfamily of ligands, which have important roles in a myriad of biological activities1. They act (Fig. 1) by binding to complexes of two type I and two type II receptors (BMPR-I and BMPR-II) and modulating the expression of target genes through a series of signal transduction pathways. Of these, signaling through the SMAD proteins is best characterized, but the activated ligand-receptor complex can also stimulate the mitogen-activated protein (MAP) kinase pathway and possibly other pathways2. To investigate this complex regulatory network and its role in essential physiological processes, specific inhibitors of signaling would be extremely valuable, but so far the known inhibitors of this pathway are soluble antagonists that sequester the BMP ligands themselves and cannot distinguish SMAD-dependent from SMAD-independent signaling events3. In this issue, Yu et al. provide an important advance in this area by identifying dorsomorphin, the first small-molecule inhibitor of BMP signaling4.
Figure 1: The BMPs bind to type I and II receptors and facilitate their association.
The constitutively active kinase domains of type II receptors phosphorylate type I receptors, and this in turn activates the SMAD signaling pathway through phosphorylation of receptor SMADs (SMAD1, SMAD5 and SMAD8). These associate with co-SMADs (SMAD4) to form a heteromeric complex that translocates to the nucleus and stimulates the expression of a wide range of target genes, including the gene encoding the iron regulatory peptide hepcidin. BMPs can also signal through SMAD-independent pathways, notably via MAP kinases. Dorsomorphin inhibits BMP signaling through the SMAD pathway, likely by affecting BMPR-I kinase activity. Many of the previously known inhibitors of BMP signaling (such as noggin and chordin) act upstream to sequester BMPs and cannot differentiate SMAD-dependent from SMAD-independent signaling. The activation of the hepcidin gene by IL-6 requires both the JAK-STAT and BMP-SMAD pathways, but how the pathways interact is unclear. Similarly, TfR2 and the HFE–TfR1 complex can alter hepcidin expression, but it is not known whether their functions require the BMP-SMAD system. Modified from ref. 10.
Full size image (54 KB)A vexing problem in identifying inhibitors that target phosphorylation events is attaining specificity for the kinase of interest while avoiding cross-reactivity against structurally similar kinases. Yu et al.4 have addressed the problem of off-target effects by using a new screen in which the output is dorsoventral patterning in zebrafish. Because they knew that mutations in components of the BMP signaling pathway lead to a very specific phenotype in zebrafish embryos—the loss of ventral structures and relative dorsalization of the embryo5—they searched several small-molecule libraries (totaling over 7,500 compounds) for substances that would phenocopy this dorsalization. Dorsomorphin (6-[4-(2-piperidin-1-yl-ethoxy)phenyl]-3-pyridin-4-yl-pyrazolo[1,5-a] pyrimidine) was the result.
In a series of well-controlled in vitro studies, the investigators showed that dorsomorphin inhibits the phosphorylation of the receptor SMADs but does not alter MAPK p38 phosphorylation, which indicates specificity for SMAD-dependent signaling. In contrast, the BMP-sequestering inhibitor noggin downregulated both pathways. Dorsomorphin was also shown to have no or very limited effects on the action of other TGF- family ligands (such as TGF-1 and activin A), which indicates considerable specificity for BMP-mediated signals. The precise target of dorsomorphin remains unclear, but preliminary studies indicate that it most likely acts downstream of BMPR-II and inhibits the kinase activity of BMPR-I.
The characterization of dorsomorphin in zebrafish embryos has shown that the inhibitor has considerable utility in the study of developmental processes, but it should prove equally useful in many other areas in which BMP signaling is implicated. As an example of this, Yu et al.4 used dorsomorphin to glean important new information about factors controlling the expression of the iron regulatory peptide hepcidin. Hepcidin has a central role in iron homeostasis by determining how much iron enters the plasma from macrophages, enterocytes and other cells6. A number of recent studies have shown that the BMP-SMAD pathway is important for the regulation of hepcidin and that stimulation through the BMPRs increases the expression of HAMP, the gene encoding the peptide7. Yu et al.4 found that dorsomorphin is highly effective in blocking HAMP expression (stimulated by either BMP2 or the BMP coreceptor hemojuvelin), which suggests that hepcidin is regulated through BMP type I receptors.
Equally intriguing are the observations that dorsomorphin blocks the stimulation of HAMP expression either by iron or by the proinflammatory cytokine interleukin 6 (IL-6). Earlier observations suggested that an intact BMP-SMAD pathway is essential for HAMP to respond to a range of superficially disparate stimuli8, but the dorsomorphin studies have extended this to show specifically the importance of BMPR signaling. The mode of action of other upstream regulators of hepcidin, such as HFE and TfR2 (both of which, when mutated, lead to the iron loading disorder hemochromatosis9), is unknown, but studies with dorsomorphin should elucidate whether they too require BMP signaling for their activities.
Our understanding of dorsomorphin action remains in its infancy. The data suggesting that it acts by interfering with the kinase activity of type I BMP receptors are strong, but this still does not tell us how it is able to functionally separate signaling through the receptor SMADs from signaling through MAPK. Perhaps the link to MAPK does not require BMPR-I kinase activity, but detailed mechanistic studies are required to confirm this. The possibility that dorsomorphin exerts effects on other targets must also be considered. Dorsomorphin (under an earlier name of compound C) has been shown to inhibit AMP-activated kinase (AMPK) activity, and it is structurally similar to compounds that inhibit the activity of KDR kinase. Yu et al.4 conducted several experiments to show that the dorsalization effects of dorsomorphin are not related to AMPK or KDR inhibition, but the role of these additional inhibitory activities requires more careful definition.
In dorsomorphin, Yu et al.4 have provided the research community with a valuable and versatile reagent that is certain to find widespread use as a tool by those interested in the mechanisms of BMP action, and they have demonstrated its utility using developmental biology and iron metabolism as examples. The finding that dorsomorphin is able to increase plasma iron levels when administered to mice shows it may find use in the treatment of the anemia of chronic disease and demonstrates the potential of this small molecule (or its derivatives) as a therapeutic agent for modulating the activity of the many physiological pathways that depend on BMP signaling.
