Deciphering the physiological function(s) of Panx3, a gene displaying specific expression in bone-forming osteoblasts

成骨细胞介导的骨形成对骨骼的发育、生长、重塑和再生是必不可少的。这一过程依赖于大量能量的供应，而成骨细胞最近被发现主要依赖于葡萄糖的供应。此外，近二十年获得的遗传学证据清楚地表明，骨也是一个内分泌器官，成骨细胞系细胞释放血清磷和血糖稳态的激素调节器。然而，成骨细胞衍生的分子对能量代谢调控的相关性仍然存在争议，潜在的复杂性仍然没有被完全理解。我们先前已经确定了PAnnexin-3(Panx3)，一种主要由成骨细胞表达的跨膜蛋白，在连接骨形成和能量代谢中的潜在作用。特别是，我们可以证明，新生小鼠缺乏Panx3不仅会导致骨骼发育受损，还会导致低血糖。进一步的实验表明，Panx3基因缺陷的成骨细胞表现出更高的细胞内ATP水平，潜在地模仿了能量过剩，并减少了代谢相关基因的表达。另一个关键发现是，尽管Panx3在肝脏中没有表达，但Panx3基因缺陷的新生儿显示出更高的肝糖原含量。最后，我们可以确定Panx3与骨再生有关，这一过程与血糖水平的变化以及肝脏和脂肪组织中代谢标记物的表达有关。综上所述，我们的研究结果强烈表明，Panx3缺乏会在骨骼发育/生长和骨折愈合期间损害成骨细胞的活性，并且Panx3缺乏还会通过改变肝脏的代谢功能来影响葡萄糖的稳态。由于我们之前的数据也提出了几个问题，我们现在打算进行额外的分析，以确定Panx3在骨骼发育和能量代谢中的分子功能。首先，我们将对在骨骼发育和生长过程中普遍存在或仅在特定细胞类型中缺失Panx3的小鼠进行深入的表型鉴定。其次，由于我们没有在成年Panx3基因缺陷小鼠中检测到主要异常，我们将通过不同的方式对它们进行挑战，包括基因激活骨形成，但也诱导骨再生。第三，为了了解观察到的表型的分子基础，我们将分析原代野生型和Panx3缺陷细胞，其中包括无偏见的转录方法。

Bone formation mediated by osteoblasts is essential for development, growth, remodeling and regeneration of the skeleton. This process depends on the provision of large amounts of energy, and osteoblasts were recently found to rely mostly on glucose availability. Moreover, genetic evidence obtained in the last two decades has clearly demonstrated that bone is also an endocrine organ, and that osteoblast lineage cells release hormonal regulators of serum phosphate and glucose homeostasis. The relevance of energy metabolism regulation by osteoblast-derived molecules is however still debated, and the underlying complexities are still not fully understood.We have previously identified a potential role of Pannexin-3 (Panx3), a transmembrane protein predominantly expressed by osteoblasts, in linking bone formation to energy metabolism. In particular, we could demonstrate that Panx3 deficiency in neonatal mice not only results in impaired skeletal development, but also in hypoglycemia. Additional experiments revealed that Panx3-deficient osteoblasts display higher intracellular ATP levels, potentially mimicking energy excess and reducing expression of metabolically relevant genes. Another key finding was that Panx3-deficient neonates displayed a higher hepatic glycogen content despite absence of Panx3 expression in the liver. Finally, we could identify Panx3 to be relevant for bone regeneration, a process found associated with alterations in blood glucose levels and metabolic marker expression in liver and adipose tissue.Collectively, our findings strongly suggest that Panx3 deficiency impairs osteoblast activity in states of excessive bone formation, i.e. during skeletal development/growth and fracture healing and that Panx3 deficiency additionally impacts glucose homeostasis by altering metabolic functions of the liver. Since our previous data also raise several questions, we now intend to perform additional analyses to define the molecular functions of Panx3 in skeletal development and energy metabolism. First, we will perform in depth-phenotyping of mice lacking Panx3, ubiquitously or only in specific cell types, during skeletal development and growth. Second, since we did not detect major abnormalities in adult Panx3-deficient mice, we will challenge them by different means, which includes genetic activation of bone formation, but also induction of bone regeneration. Third, in order to understand the molecular bases of the observed phenotypes we will analyze primary wildtype and Panx3-deficient cells, which includes unbiased transcriptomic approaches.