학위논문 (박사)-- 서울대학교 대학원 : 치의학대학원 치의과학과, 2018. 2. 류현모.One of the most reported post-translational modifications (PTMs) regulation of protein in the cell is prolin-directed phosphorylation, these phosphorylated resides also can be the targets of enzyme for other PTMs. Pin1 is an isomerase that recognizes phosphorylated Ser/Thr-Pro residues and causes the structural change of protein. Our previous studies have shown that Pin1 deficient exhibited a wide range of abnormal bone phenotypes. Moreover, in the signaling pathways such as FGF, BMP and PTH, which are responsible for osteoblast differentiation, isomerization of the molecule by Pin1 plays important role.
Wnt/β-catenin signaling pathway, in which β-catenin nuclear localization is a critical step, also plays a major role in the control of osteoblast differentiation. However, the association of Pin1 with Wnt/β-catenin signaling pathway during bone development and detailed mechanism is not clear. In Part 1, we observed a marked reduction of β-catenin in osteoblasts of Pin1 null mice tibia. From MC3T3-E1 cells, transcription activity, the nucleus β-catenin level and the target gene expression of β-catenin were all decreased when the activity of Pin1 was reduced by si RNA or by inhibitor. We also demonstrated that Pin1 directly interacted with β-catenin, and the isomerized β-catenin which could not bind to nuclear adenomatous polyposis coli (APC) only can be retention and act as a transcription factor in the nucleus. These results might explain the decrease of β-catenin ubiquitination and showed that Pin1 is a critical factor that can regulate bone formation regulated by Wnt/β-catenin signaling pathway.
In part 2, the importance of Pin1 in osteogenesis was confirmed in craniosynostosis (CS) disease animal model. CS is characterized by the premature closure of craniofacial sutures. It is follows autosomal dominant inheritance pattern, and the mutation of the FGFR2 gene is mainly the genetic cause of it. This study focused on the change of coronal suture fusion which is the major phenotype of Apert syndrome (AS) using S252W gain of function mutation of FGFR2. We aimed to treat CS caused by over-activated FGF/FGFR signal, and it was targeted that inhibiting Pin1-induced isomerization of Runx2 in FGF signaling pathway. As a result, we showed that fusion of coronal suture can be restored to close to the normal phenotype when the FGFR2S252W+ and the Pin1+/- genotype are present simultaneously. The similar results were obtained when juglone, Pin1 inhibitor, was administered to the fetus of FGFR2S252W/+ mice for drug therapy.
Collectively, we demonstrated here that isomerization of β-catenin through direct interaction of β-catenin with Pin1 during osteoblast differentiation is an important factor controlling nuclear β-catenin retention and stability. In addition, we demonstrated the drug target of CS by proving that the inactivation of Pin1 causes the recovery of disease through gain of function mutation in FGFR2.I. Abbreviations 1
II. Literature review 4
III. Purpose of study 16
IV.Part 1. Pin1-mediated modification prolongs the nuclear retention of β-catenin in Wnt3a-induced osteoblast differentiation 17
V. Part 2. Pin1 is a new therapeutic target of craniosynostosis 71
VI. Conclusion 134
VII. References 136
VIII. 국문 초록 150Docto
