Elucidating The Developmetal Defect In Werner Syndrome Using Stem Cells
Refereed conference paper presented and published in conference proceedings


摘要Werner Syndrome (WS) is an autosomal recessive genetic disorder characterized by premature aging. This disease is caused by mutations in the WRN gene. The first sign of WS is short stature. Individuals with WS have an abnormally slow growth rate, and growth stops at puberty. As a result, affected individuals have short stature. However, the mechanism is still not clear. To find out the cause of short stature in WS, we reprogrammed the WS patients fibroblasts and the isogenic normal control into mesenchymal stem cells and compared the transcriptome by RNA-Seq. Among the top 10 candidates, SHOX (short stature homeobox) was noted. SHOX plays an important role in chondrogenesis. SHOX deficiency is a frequent cause of short stature. So, what is the role of SHOX in WS pathogenesis? To answer this question, we induced hESCs towards chondrocytes. We analyzed the WRN and SHOX expression during chondrogenesis and noticed that the expression of both genes increased gradually, indicating a high correlation during chondrogenesis. Then, we induced the chondrocytes by the same protocol in WRN knockout (WRN-/-) or SHOX knockdown cells. We noticed that WRN loss led to the decrease of SHOX during chondrogenesis. However, SHOX knockdown has no effect on WRN, suggesting that WRN may regulate SHOX as an upstream target. To further assess the role of WRN and SHOX during chondrogenesis, we immunostained the cells for specific chondrogenic markes, SOX9 and COL2A1. Again, we noticed that WRN knockout or SHOX knockdown suppressed SOX9 and COL2A1 expression. We conclude that the loss of WRN and SHOX blocked the chondrocyte differentiation. We next asked how WRN regulated SHOX. It is reported that the WRN helicase is able to recognize G-quadruplex (G4) motif and thus altering gene transcription. So we did immunostaining to detect the prevalence of G4 in vitro. Interestingly, we found that the signals of G4 staining in WRN-/- cells was much stronger than WRN+/+ cells, indicating that WRN is able to unwind some of the G4. Then, binding of WRN to G4-containing SHOX promoter was confirmed by ChIP, whereas luciferase assays suggested that WRN could unwind SHOX G4 complex and promoted its expression. In summary, we found that the downregulation of SHOX in WS inhibited chondrogenesis at the early developmental stage, which may account for the short stature.
著者TIAN Yuyao, MIU Kai Kei, WANG Zhangting, CHEUNG Hoi Hung, CHAN Wai Yee
會議名稱2019 ISSCR/KSSCR International Symposium

上次更新時間 2020-01-06 於 15:13