Role of PARP1 in the Osteoclast Lineage

多聚(ADP-核糖)聚合酶1(PARP1)是一种含有催化等多种结构域的蛋白质 结构域，其中一些参与DNA与其他分子的结合和相互作用。它催化了 将多个ADP-核糖单元从NAD+转移到聚ADP-核糖(PAR)聚合物 受体蛋白，一种翻译后修饰，称为PAR化。PARP1在细胞凋亡过程中被切割， 但对细胞死亡不是必需的，因为表达不可切割的PARP1的小鼠发育正常，而缺乏PARP1的小鼠 这种蛋白质。我们发现，在全球范围内表达不可切割的PARP1突变体(结构性)的敲入小鼠 活性突变体)由于OC分化和骨吸收减少而表现出高骨量表型， 而骨形成不受影响。相反，胚系切除小鼠的PARP1会导致骨量减少 源于破骨细胞的增加，而成骨作用没有改变。因此，PARP1的函数增益 突变阻碍了破骨细胞的形成，而功能丧失则促进了这一过程。从机制上讲，我们发现 PARP1在OC前体细胞中高度表达，并在OC分化过程中降解，并且 主要的OC转录因子NFATc1在破骨细胞形成过程中被PAR化，这意味着翻译后 由PARP1进行监管。我们还发现PARP1抑制了OC的整体抑制因子的表达 抑制者，Blimp1。有趣的是，虽然PARP1酶活性的抑制剂可以促进OC的形成 小鼠骨髓巨噬细胞在M-CSF和RANKL存在下的体外表达 缺乏催化结构域的PARP1突变体抑制破骨细胞的生成。这些观察结果表明 PARP1通过PAR化和其他不需要其催化活性的机制抑制OC的分化。 然而，PARP1对破骨细胞形成的复杂调控作用的相对作用尚不清楚。这个 这一建议的中心假设是PARP1通过依赖PAR化抑制骨吸收，并且- 破骨细胞生成的独立拮抗作用。我们将在以下两个目标中检验这一假设。目标1： 确定PARP1的骨架作用。目标2：确定PARP1调节的机制 破骨细胞生成。这项拟议的工作将确定PARP1在调节骨骼方面的生理重要性 动态平衡及其作用机制。因此，这项提议将在骨重建方面开辟新的天地， 专注于OC监管的新参与者。了解PARP1在骨重建中的作用将有助于澄清 一类新药物未被认识到的潜在副作用，并确定了一条可能的研究新途径 旨在抑制骨吸收的治疗进展。

Poly(ADP-ribose) polymerase 1 (PARP1) is a multi-domain protein containing a catalytic and other domains, some of which are involved in DNA binding and interaction with other molecules. It catalyzes the formation of poly-ADP-ribose (PAR) polymers by transferring multiple ADP-ribose units from NAD+ onto acceptor proteins, a post-translational modification termed PARylation. PARP1 is cleaved during apoptosis, but is not essential for cell death as mice expressing uncleavable PARP1 develop normally as do mice lacking this protein. We find that knock-in mice globally expressing a non-cleavable PARP1 mutant (constitutively active mutant) exhibit a high bone mass phenotype owing to decreased OC differentiation and bone resorption, while bone formation is unaffected. Conversely, germline ablation of Parp1 in mice causes osteopenia stemming from increased osteoclastogenesis, while osteogenesis is unaltered. Thus, PARP1 gain-of-function mutation hinders osteoclastogenesis whereas loss-of-function promotes this process. Mechanistically, we find that PARP1 is highly expressed by OC precursors and it is degraded during OC differentiation, and that the master OC transcription factor, NFATc1, is PARylated during osteoclastogenesis, implying post-translational regulation by PARP1. We also find that PARP1 inhibits the expression of the global repressor of OC suppressors, Blimp1. Intriguingly, while an inhibitor of PARP1 enzymatic activity enhances OC formation from mouse bone marrow macrophages cultured in the presence of M-CSF and RANKL, in vitro expression of a PARP1 mutant lacking the catalytic domain inhibits osteoclastogenesis. These observations suggest that PARP1 inhibits OC differentiation via both PARylation and other mechanisms not requiring its catalytic activity. However, the relative roles of PARP1 complex regulatory effects on osteoclastogenesis are unknown. The central hypothesis of this proposal is that PARP1 inhibits bone resorption via PARylation -dependent and - independent antagonism of osteoclastogenesis. We will test this hypothesis in the following two aims. Aim 1: Determine the skeletal actions of PARP1. Aim 2: Define the mechanisms of PARP1 regulation of osteoclastogenesis. The proposed work will determine PARP1 physiologic importance in modulating bone homeostasis, and its mechanism of action. Therefore, this proposal will break new ground on bone remodeling, focusing on a new player in OC regulation. Understanding PARP1 role in bone remodeling will clarify unrecognized, potential side effects of a new class of drugs and define a possible new avenue of research for therapeutic development with the purpose of inhibiting bone resorption.