Poster #010, Cedars Sinai Medical Center
Investigating GATAD2B Function during Neural Development using GAND as a model
Mentors: Molly Easter, PhD and Tyler M. Pierson, MD-PhD
GATAD2B-associated neurodevelopmental disorder (GAND) is an ultra-rare condition affecting cortical development. GATAD2B is a subunit of the Nucleosome Remodeling and Deacetylase complex (NuRD) that bridges two NuRD domains: the Histone Deacetylase Core and Chromatin-Remodeling Subcomplex. To date, GATAD2B is the one of the least studied members of the NuRD complex and its relevance to brain development is evidenced by its association with GAND. Previous data from our lab and others (from clinical and mouse studies) have demonstrated that GATAD2B haploinsufficiency is associated with behavioral, learning and facial abnormalities, altered gene expression and abnormal cortical development. Specifically, GATAD2B deficiency during corticogenesis affects cortical patterning and results in increased co-expression of SATB2 and CTIP2 (BCL11B) in layer V of the cortex. To investigate GATAD2B function during corticogenesis, we engineered a doxycycline inducible HA-tagged GATAD2B cassette that will be integrated into the AAVS1 locus of control and GAND patient derived IPSCs. To test the functionality of our doxycycline inducible HA-GATAD2B cassette, we performed transient transfections in HEK293T cells with and without doxycycline treatment and collected the cell lysates 48-hour post-transfection. Western blots and qPCR on lysates from cells treated with doxycycline had increased mRNA and protein expression of HA-GATAD2B. Our results demonstrate that we were able to make a functional doxycycline inducible HA-GATAD2B cassette. Future directions include integrating this cassette into patient-derived IPSC neurons, to investigate whether inducible GATAD2B can rescue assays of NuRD dysfunction in GAND IPSCs and cortical organoids.