Figure 1. ASB2α induced degradation of FLNa but not of JAK1 and JAK2 in acute myeloid leukemia cells.

(A) Expression of ASB2α, FLNa, JAK1, JAK2 and non-muscle myosin IIA heavy chain (NM myosin IIA) in differentiating leukemia cells. Promyelocytic NB4 and myeloblastic PLB985 cells were left untreated (-) or were treated with 10⁻⁶ M of all-trans retinoic acid (RA) as indicated. Differentiation was assessed by cell morphology on May-Grünwald-Giemsa-stained cytospins, and quantification of the percentage of cells with nitro blue tetrazolium deposits (not shown). (B) ASB2α E3 ubiquitin ligase activity triggers degradation of FLNa but not of JAK1 and JAK2 in myeloid leukemia cells. PLB985/MT-FlagASB2α (ASB2αWT) and PLB985/MT-FlagASB2αLA (ASB2αLA) cells were left untreated (-) or were treated for 8, 48, 96 and 144 hours with 75 μM or 85 μM ZnSO₄, respectively to induce the expression of equivalent amounts of ASB2α proteins. In A and B, 10-μg aliquots of the protein extracts were separated by SDS-PAGE and immunoblotted for ASB2α, FLNa, JAK1, JAK2 and NM myosin IIA. (C) U937 cells were imaged 20 hours after nucleofection with GFP-ASB2α or GFP-ASB2αLA expression vector. Cells were fixed and stained for JAK3 and FLNa. Scale bar represents 10 μm.