Skip to main content
. 2018 Jun 4;128(7):2877–2893. doi: 10.1172/JCI98765

Figure 4. NEK2 activates the canonical NF-κB signaling pathway in primary multiple myeloma samples.

Figure 4

(A) Primary myeloma samples from 16 patients (Pts) were lysed and analyzed by Western blot using NEK2, p-p65-S536, total p65 (p65), USP7, and GAPDH antibodies. (B) CD138-positive myeloma cells isolated from 4 primary myeloma patients were mounted on cytospin slides and analyzed by immunofluorescence using NEK2 and p-p65-S536 antibodies. DAPI staining was used to visualize nuclei. Yellow arrowheads indicate myeloma cells coexpressing NEK2 and p-p65-S536. Blue arrowheads show myeloma cells expressing p-p65-S536 with undetectable NEK2 levels. (C) EV and NEK2-OE ARP1 cells were treated with vehicle, BAY11-7082 (0.5 or 1.0 μM), and bortezomib (5 nM) alone or in combination. After 48 hours, cell viability was assessed by trypan blue staining and Dunnett’s method was used to calculate the multiplicity-adjusted P values for each treatment and control group pair. **P = 0.0023; ****P = 0.0001. NS, no significance. Experiment was performed in triplicate. (D) A model for NEK2 deubiquitination and stabilization by interacting with USP7. USP7 prevents E3 ligase APC/C (30) to ubiquitinate NEK2 resulting in its stabilization.