Examining the Role of FAM109 in an Undiagnosed Disease

Student: Liz Urbanski
Mentor: Graydon Gonsalvez (Augusta University)

A patient’s undiagnosed disease has similar characteristics to the ciliopathies Lowe Syndrome and Dent Disease. A mutation in the patient’s FAM109 gene was found, and may be causative of the disease. Since very little is known about FAM109, studying its localization, function, and relationship to other genes associated with Lowe Syndrome and Dent Disease is necessary to understand the role of this gene in the patient’s undiagnosed disease.

The National Institutes of Health Undiagnosed Disease Program identified a de novo arginine to cysteine mutation in a patient’s FAM109A gene (R6C). The human genome contains two FAM109 genes, FAM109A and FAM109B. The patient presents with disease characteristics reminiscent of a ciliopathy, a type of disease that results from mutations in OCRL, an interacting partner of FAM109. The aim of our project was to identify the functions and localizations of the FAM109 paralogs and determine if the R6C mutation is causative of the patient’s disease. HeLa cells were treated with antibodies against various cellular components to determine the localization of FAM109A and FAM109B. Immunofluorescence showed co-localization between FAM109A and the midbody marker MKLP1, as well as with the Golgi marker 58K. OCRL was found to co-localize with FAM109A and MKLP1 at the midbody. GFP-tagged wildtype and mutant constructs of FAM109A were transfected into HeLa cells and the phenotype of these cells was analyzed by immunofluorescence. Transfected mutant isoforms of FAM109A did not affect the ability of FAM109A to localize to the midbody. Immunofluorescence with FAM109B showed localization with the mitochondrial marker Cox IV, and transfection with FAM109B siRNA led to cell death. We hypothesize that the cell death is occurring via apoptosis. Our findings reveal that FAM109A localizes to both the Golgi and the midbody, while FAM109B localizes to the mitochondria. Mutant isoforms of FAM109A localize to the midbody in an identical manner to the wild type, and co-localization at the midbody with OCRL has been validated. The specific functions of the FAM109 paralogs remain largely unknown and will be the topic of future studies.