Department of Neurobiology, Division of Hematology and Hematologic Malignancies, University of Utah, Salt Lake City, Utah, USA.
Nephrology Division and Endocrine Unit, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, USA.
J Biol Chem. 2024 Oct;300(10):107790. doi: 10.1016/j.jbc.2024.107790. Epub 2024 Sep 19.
Bone morphogenetic protein 2 (BMP2) and BMP6 are key regulators of systemic iron homeostasis. All BMPs are generated as inactive precursor proteins that dimerize and are cleaved to generate the bioactive ligand and inactive prodomain fragments, but nothing is known about how BMP2 or BMP6 homodimeric or heterodimeric precursor proteins are proteolytically activated. Here, we conducted in vitro cleavage assays, which revealed that BMP2 is sequentially cleaved by furin at two sites, initially at a site upstream of the mature ligand, and then at a site adjacent to the ligand domain, while BMP6 is cleaved at a single furin motif. Cleavage of both sites of BMP2 is required to generate fully active BMP2 homodimers when expressed in Xenopus embryos or liver endothelial cells, and fully active BMP2/6 heterodimers in Xenopus. We analyzed BMP activity in Xenopus embryos expressing chimeric proteins consisting of the BMP2 prodomain and BMP6 ligand domain, or vice versa. We show that the prodomain of BMP2 is necessary and sufficient to generate active BMP6 homodimers and BMP2/6 heterodimers, whereas the BMP6 prodomain cannot generate active BMP2 homodimers or BMP2/6 heterodimers. We examined BMP2 and BMP6 homodimeric and heterodimeric ligands generated from native and chimeric precursor proteins expressed in Xenopus embryos. Whereas native BMP6 is not cleaved when expressed alone, it is cleaved to generate BMP2/6 heterodimers when co-expressed with BMP2. Furthermore, BMP2-6 chimeras are cleaved to generate BMP6 homodimers. Our findings reveal an important role for the BMP2 prodomain in dimerization and proteolytic activation of BMP6.
骨形态发生蛋白 2（BMP2）和 BMP6 是全身铁稳态的关键调节因子。所有的 BMP 都是作为无活性的前体蛋白产生的，这些前体蛋白二聚化并被切割，生成有生物活性的配体和无活性的前导片段，但目前还不知道 BMP2 或 BMP6 同源二聚体或异源二聚体前体蛋白是如何被蛋白水解激活的。在这里，我们进行了体外切割实验，结果表明 BMP2 首先在前导肽域上游的一个位点，然后在靠近配体域的一个位点被 furin 连续切割，而 BMP6 则在一个 furin 基序处被切割。BMP2 的两个位点的切割对于在 Xenopus 胚胎或肝内皮细胞中表达时生成完全活性的 BMP2 同源二聚体以及在 Xenopus 中生成完全活性的 BMP2/6 异源二聚体是必需的。我们分析了在表达嵌合蛋白的 Xenopus 胚胎中 BMP 活性，该嵌合蛋白由 BMP2 前导肽和 BMP6 配体域组成，或者反之亦然。我们表明，BMP2 的前导肽是生成活性 BMP6 同源二聚体和 BMP2/6 异源二聚体所必需和充分的，而 BMP6 的前导肽不能生成活性 BMP2 同源二聚体或 BMP2/6 异源二聚体。我们检查了从天然和嵌合前体蛋白表达的 Xenopus 胚胎中生成的 BMP2 和 BMP6 同源二聚体和异源二聚体配体。虽然单独表达时天然 BMP6 不被切割，但当与 BMP2 共表达时，它会被切割生成 BMP2/6 异源二聚体。此外，BMP2-6 嵌合体被切割生成 BMP6 同源二聚体。我们的研究结果揭示了 BMP2 前导肽在 BMP6 二聚化和蛋白水解激活中的重要作用。
