Department of Oral Biology, School of Dental Medicine, University of Buffalo, State University of New York, Buffalo, NY 14214, USA.
Bone. 2012 Sep;51(3):407-17. doi: 10.1016/j.bone.2012.06.021. Epub 2012 Jul 4.
Intraflagellar transport (IFT) proteins are essential for the assembly and maintenance of cilia, which play important roles in development and homeostasis. IFT80 is a newly defined IFT protein. Partial mutation of IFT80 in humans causes diseases such as Jeune asphyxiating thoracic dystrophy (JATD) and short rib polydactyly (SRP) type III with abnormal skeletal development. However, the role and mechanism of IFT80 in osteogenesis is unknown. Here, we first detected IFT80 expression pattern and found that IFT80 was highly expressed in mouse long bone, skull, and during osteoblast differentiation. By using lentivirus-mediated RNA interference (RNAi) technology to silence IFT80 in murine mesenchymal progenitor cell line-C3H10T1/2 and bone marrow derived stromal cells, we found that silencing IFT80 led to either shortening or loss of cilia and the decrease of Arl13b expression - a small GTPase that is localized in cilia. Additionally, silencing IFT80 blocked the expression of osteoblast markers and significantly inhibited ALP activity and cell mineralization. We further found that IFT80 silencing inhibited the expression of Gli2, a critical transcriptional factor in the hedgehog signaling pathway. Overexpression of Gli2 rescued the deficiency of osteoblast differentiation from IFT80-silenced cells, and dramatically promoted osteoblast differentiation. Moreover, introduction of Smo agonist (SAG) promotes osteoblast differentiation, which was partially inhibited by IFT80 silencing. Thus, these results suggested that IFT80 plays an important role in osteogenesis through regulating Hedgehog/Gli signal pathways.
鞭毛内运输（IFT）蛋白对于纤毛的组装和维持至关重要，纤毛在发育和稳态中发挥着重要作用。IFT80 是一种新定义的 IFT 蛋白。人类 IFT80 的部分突变导致 Jeune 窒息性胸廓发育不良（JATD）和短肋多指（SRP）III 型等疾病，这些疾病伴有骨骼发育异常。然而，IFT80 在成骨中的作用和机制尚不清楚。在这里，我们首先检测了 IFT80 的表达模式，发现 IFT80 在小鼠长骨、颅骨中高度表达，并在成骨细胞分化过程中高表达。通过使用慢病毒介导的 RNA 干扰（RNAi）技术沉默小鼠间充质祖细胞系 C3H10T1/2 和骨髓基质细胞中的 IFT80，我们发现沉默 IFT80 导致纤毛缩短或缺失，并且 Arl13b 表达减少 - Arl13b 是一种定位于纤毛中的小 GTPase。此外，沉默 IFT80 阻断了成骨细胞标志物的表达，并显著抑制了碱性磷酸酶（ALP）活性和细胞矿化。我们进一步发现，IFT80 沉默抑制了 Hedgehog 信号通路中的关键转录因子 Gli2 的表达。Gli2 的过表达挽救了 IFT80 沉默细胞中成骨分化的缺陷，并显著促进了成骨分化。此外，引入 Smo 激动剂（SAG）促进成骨分化，而 IFT80 沉默部分抑制了这一过程。因此，这些结果表明，IFT80 通过调节 Hedgehog/Gli 信号通路在成骨中发挥重要作用。
