Targeting G-quadruplex for rescuing impaired chondrogenesis in WRN-deficient stem cells.

BACKGROUND: Pathogenic mutations in WRN are a cause of premature aging disease Werner syndrome (WS). Besides accelerated aging phenotypes and cancer predisposition, patients with WS also display underdevelopment in the skeletal system, characterized by short stature, light body weight and unusually thin extremities. The reasons for these developmental defects are not completely understood and the underlying molecular mechanism remains to be elucidated. RESULTS: In this study, WRN was found to modulate transcription of short stature homeobox gene SHOX. Loss of WRN resulted in insufficient expression of SHOX, the gene dose of which is critical for driving chondrocyte differentiation. WRN could bind the G-quadruplex (G4) structures in the SHOX promoter and stimulate transcription. Aberrant formation of G4 structures in WRN-deficient cells impeded normal transcription of SHOX, thus resulting in impaired chondrogenesis. Chondrogenesis could be rescued by overexpression of WRN helicase or SHOX, suggesting that SHOX is a downstream target of WRN. Gene editing of the G4 structures in the SHOX promoter could increase SHOX expression, therefore rescuing the impaired chondrogenesis in WRN-deficient cells. CONCLUSIONS: Our data suggest that dysgenesis of the developing bone in WS might be caused by SHOX insufficiency. Aberrant formation of G4 structures in SHOX promoter suppresses SHOX expression and impairs chondrogenesis. Targeted mutagenesis in the G4 structures enhances SHOX expression and thus providing an opportunity to rescue the chondrogenic defect.

In vivo genome editing thrives with diversified CRISPR technologies.

Prokaryotic type II adaptive immune systems have been developed into the versatile CRISPR technology, which has been widely applied in site-specific genome editing and has revolutionized biomedical research due to its superior efficiency and flexibility. Recent studies have greatly diversified CRISPR technologies by coupling it with various DNA repair mechanisms and targeting strategies. These new advances have significantly expanded the generation of genetically modified animal models, either by including species in which targeted genetic modification could not be achieved previously, or through introducing complex genetic modifications that take multiple steps and cost years to achieve using traditional methods. Herein, we review the recent developments and applications of CRISPR-based technology in generating various animal models, and discuss the everlasting impact of this new progress on biomedical research.