Deletion of a GA-repeat in human GPM6B disrupts differentiation of NT2 cells to neural cells

Across multiple primates and tissues, GPM6B (Glycoprotein membrane 6B) reaches maximum levels of expression in the human brain. This gene contains a strictly monomorphic short tandem repeat (STR) of (GA)9. We used a CRISPR-based tool to delete this GA-repeat in NT2 cells, and analyzed the consequence of this deletion on GPM6B expression, using qRT-PCR and western blot. Subsequently, the edited cells were induced to differentiate into neural cells, using retinoic acid (RA) treatment, and sorted by flowcytometry. Deletion of the GA-repeat significantly decreased the expression of GPM6B at the RNA level ( p  < 0.05) and 40% at the protein level. Compared to the control cells, the edited cells showed a dramatic decrease of the astrocyte and neural cell markers at the protein level, including GFAP (0.77-fold), TUBB3 (0.57-fold), and MAP2 (0.2-fold). Subsequent sorting of the edited cells, using the cell surface markers showed an increased number of NES ( p  < 0.01), but decreased number of GFAP ( p  < 0.001), TUBB3 ( p  < 0.05), and MAP2 ( p  < 0.01), compared to the control cells. In conclusion, we provide prime evidence of the biological role of a STR, exemplified in the regulation of GPM6B expression and optimal differentiation of human neural cell lineages.