Main author: Elín Sóley Sigurbjörnsdóttir
Institution or Company: University of Iceland
Co-Authors, Institution or Company:
Sara Sigurbjörnsdóttir, University of Iceland. Eiríkur Steingrímsson, University of Iceland.
Introduction: The hedgehog (HH) signaling pathway plays a role in regulating normal growth and differentiation of chondrocytes. Cholesterol is essential for HH signaling, especially for its second messenger role in activation of the Smoothened (SMO) protein, a key activator of the HH pathway. In vertebrates, SMO signaling occurs through primary cilium. In a large genome-wide association study, deCODE genetics discovered a rare missense variant (SMOR173C) in the human SMO gene. Patients carrying this mutation are at high risk for developing hip osteoarthritis, a painful disease causing physical disability. Hence, further understanding of the SMOR173C variant is important. We hypothesise that the amino acid substitution, caused by the mutation, results in misfolded cholesterol-binding pocket of SMO, potentially attenuating the signaling ability of SMO. Aberrant HH signaling would then affect phenotypic stability of chondrocytes in the hip joints.
Material and methods: We are testing the functional effects of this variant using assays that determine activity of the HH pathway, cholesterol-binding, and localisation to primary cilium. As controls we use wild type SMO and variants that lack cholesterol-binding. The primary focus has been on identifying cell lines that form primary cilia and are responsive to HH signaling. We have used the CRISPR and PRIME-edit technologies to knockout SMO and introduce specific mutations into SMO. The goal is to generate systems that truly represent the variant and then determine its effects on HH signaling.
Results and conclusions: Our results suggest that the degree of HH responsiveness varies between cell lines and the variant does not affect localisation to the primary cilia.