Determining if Golga4 Regulates Notch2 in Kidney Epithelial Cells — 98p — Renee Virtue, Ayesha Anwer, Jenn DeRiso, Alicia Simmons, Matthew McKillop

Alagille syndrome is a dominant, multisystemic, disorder that affects approximately 1:30,000-100,000 people with a mortality rate of around 10%. The symptoms are highly variable between individuals and may consist of cholestasis, cardiac disease, ocular abnormalities, skeletal abnormality, renal abnormalities, and characteristic facial features. 

Notch signaling is a highly conserved pathway that plays a crucial role in organismal development. Mutations in JAG1 and NOTCH2, a ligand and a receptor of the notch pathway, have been linked to Alagille syndrome. These mutations are associated with symptoms such as small cystic kidneys. Yeast 2 Hybrid and BioID assays used to identify mediators of Notch2 function in the kidney revealed that Notch2 may interact with Golga4, a trans-golgi protein involved in endosomal trafficking. Recent genomic screening in patients with Alagille syndrome identified a mutation in GOLGA4.

In this study, we tested if Golga4 regulates Notch2 activity. In order to study the relationship between Golga4 and Notch2, we attempted to clone and characterize the effect of ectopically expressing N and C-terminal parts of Golga4. Using confocal microscopy and western blotting, we confirmed the expression of previously cloned Crp46 and 1936 Golga4 C-terminal fragments. By dual luciferase assays we determined that Notch2 dependent transcriptional activity is reduced in the absence of Golga4. In addition, we observed that transfection of Golga4 fragments in MDCK wild type cells led to a decrease in Notch2 activity. The C-terminal fragments of Golga4 were unable to rescue Notch2 activity in the Golga4 null cells. Overall, our findings indicate that Golga4 regulates Notch2 activity.

Sanford Research
Kamesh Surendran