BMP Ligands in Hepcidin Regulation

铁是一种必需的营养物质，但过量的铁是有毒的。调节全身铁稳态的主要激素是肝磷脂，它由肝脏分泌，诱导铁出口蛋白铁转运蛋白的降解，从而抑制铁从饮食中的吸收和铁从体内储存的释放。Hepcidin的产生受铁、促红细胞生成驱动和炎症的调节，为红细胞生成和其他基本功能提供足够的铁，但限制铁过量的毒性。我们已经发现骨形态发生蛋白（BMP）信号通路，通过配体BMP6和BMP2，是hepcidin转录响应大多数已知信号的中心调节器。BMP配体是一种二聚体蛋白，最初是作为骨诱导因子被发现的，但现在已知在许多器官从胚胎发生到成年组织稳态的许多生物过程中起着关键作用。bmp是由原结构域和配体结构域组成的非活性前体，通过蛋白水解裂解释放。虽然原结构域缺乏信号活性，但相关BMP/TGF-b家族成员越来越多地认识到原结构域在配体折叠和二聚化中起着关键作用，也可能调节配体/受体的相互作用。值得注意的是，BMP6原结构域突变与人类hepcidin调节的改变和铁超载有关。我们将展示最近发表的数据，BMP2和BMP6在hepcidin和铁稳态调节中共同起作用；异二聚体BMP4/7和BMP2/7，而不是同二聚体配体，是哺乳动物胚胎发生的主要介质；BMP原结构域在BMP4/7异二聚体和同二聚体的形成和功能中起重要作用。因此，我们假设BMP2/6异源二聚体是hepcidin和铁稳态调节的关键功能配体，并且原结构域在BMP2/6的成熟和功能中起关键作用。在Aim I中，我们将使用Xenopus系统、原代肝细胞、小鼠模型和新的ELISA方法来测试BMP2/6异二聚体在体内是否存在并受铁的调节，并阐明BMP6和BMP2如何进行蛋白水解加工，它们的前结构域如何促进异二聚体和/或同二聚体的形成和调节hepcidin的功能，以及BMP6前结构域突变如何影响这些过程导致铁过载。在Aim II中，我们将展示初步数据，在缺乏BMP6和/或BMP2的情况下，铁和红细胞生成驱动仍能调节hepcidin，我们将确定另外两种参与hepcidin调节的BMP配体。我们将使用遗传小鼠模型来确定这两种BMP配体在体内hepcidin和铁稳态调节中的功能作用。该项目的长期目标是了解如何调节BMP信号以控制hepcidin表达和全身铁稳态；这一过程如何在遗传性血色素沉着症、炎症性贫血和地中海贫血等铁疾病中受到干扰；并最终开发出治疗这些铁紊乱的新方法。我们还旨在获得对BMP配体成熟和原结构域功能的基本见解，这将与BMP信号重要的许多其他领域相关。

Iron is an essential nutrient, but excess iron is toxic. The master regulator of systemic iron homeostasis is the hormone hepcidin, which is secreted by the liver and induces degradation of the iron exporter ferroportin to inhibit iron absorption from the diet and iron release form body stores. Hepcidin production is regulated by iron, erythropoietic drive, and inflammation to provide adequate iron for erythropoiesis and other essential functions, but to limit the toxicity of iron excess. We have discovered that the bone morphogenetic protein (BMP) signaling pathway, via the ligands BMP6 and BMP2, is a central regulator of hepcidin transcription in response to most known signals. BMP ligands are dimeric proteins that were initially discovered as bone inducing factors, but are now known to play critical roles in many biologic processes from embryogenesis to adult tissue homeostasis in many organs. BMPs are made as inactive precursors comprised of a prodomain and a ligand domain that is released by proteolytic cleavage. Although prodomains lack signaling activity, there is increasing recognition from related BMP/TGF-b family members that prodomains play critical roles in ligand folding and dimerization, and may also regulate ligand/receptor interactions. Notably, BMP6 prodomain mutations have been linked to altered hepcidin regulation and iron overload in humans. We will show recently published data that BMP2 and BMP6 function together in hepcidin and iron homeostasis regulation; heterodimeric BMP4/7 and BMP2/7, rather than homodimeric ligands, are the major mediators mammalian embryogenesis; and BMP prodomains play essential roles in BMP4/7 heterodimer and homodimer formation and function. We therefore hypothesize that BMP2/6 heterodimers are a key functional ligand for hepcidin and iron homeostasis regulation and that prodomains have critical roles in BMP2/6 maturation and function. In Aim I, we will use our Xenopus system, primary liver cells, mouse models, and novel ELISA assay to test whether BMP2/6 heterodimers are present and regulated by iron in vivo, and to elucidate how BMP6 and BMP2 are proteolytically processed, how their prodomains contribute to heterodimer and/or homodimer formation and function to regulate hepcidin, and how BMP6 prodomain mutations impact these processes to cause iron overload. In Aim II, we will show preliminary data that both iron and erythropoietic drive still regulate hepcidin in the absence of BMP6 and/or BMP2, and we will identify two other BMP ligands that participate in hepcidin regulation. We will use genetic mouse models to establish the functional role of these two BMP ligands in hepcidin and iron homeostasis regulation in vivo. The long-term goals of this project are to understand how BMP signaling is regulated to control hepcidin expression and systemic iron homeostasis; how this process is perturbed in iron disorders such as hereditary hemochromatosis, anemia of inflammation, and b-thalassemia; and ultimately to develop new treatments for these iron disorders. We also aim to gain fundamental insights into BMP ligand maturation and prodomain function that will be relevant for many other fields where BMP signaling is important.