Fig. 7.
GSK3β phosphorylates emerin and regulates its intracellular location. a Emerin 1-221His6 (5 μg) and emerin 1-221His6 m175 (5 μg) were incubated in the absence (−) or presence (+) of 250U recombinant GSK3β in 1× reaction buffer for 30 min at 30°C. Several phospho-emerin bands are evident (M r 34–37 kDa) as is an auto-phosphorylated GSK3β (M r ~ 50 kDa). Inclusion of the GSK3β inhibitor SB216763 in the reaction completely abolished emerin phosphorylation. b Cultures of HEK293s and NRCs were transfected with cDNA encoding HA-tagged wild type GSK3β or HA-tagged GSK3β S9A. After 24 h cells were fixed. Immunostaining with anti-HA demonstrates exogenous expression of GSK3β whilst counterstaining with either APS20 (NRCs, red) or AP8 (HEK293s, green) demonstrates the loss of emerin from the NE in GSK3β expressing cells. Scale bars 10 μm