Figure 2. KDM4A overexpression promotes EGFR copy gains.

A) Transient overexpression of GFP-tagged KDM4A drives EGFR copy number gain in RPE cells. B) Catalytic activity of KDM4A and the Tudor domains are required for EGFR copy gains. C) Representative DNA FISH image of a stable KDM4A overexpressing RPE nucleus with EGFR DNA copy number gain (red). D) RPE cells with stable KDM4A overexpression have increased EGFR DNA copies. E) Upper panel: Analysis of publicly available ChIP-sequencing data reveals that KDM4A is recruited to the EGFR locus (40). Lower panel: RNA sequencing analysis showed increased EGFR transcripts in RPE cells stably overexpressing KDM4A. F) KDM4A overexpressing RPE cells have increased EGFR transcripts as measured by qPCR. G) KDM4A overexpressing RPE cells have increased sensitivity to the EGFR-family inhibitor, Lapatinib, as measured by trypan blue exclusion assay. H) KDM4A overexpressing RPE cells have a dose-dependent increase in sensitivity to the specific EGFR inhibitor, Gefitinib, as measured by trypan blue exclusion assay. I) KDM4A overexpressing RPE cells migrate faster following 24 hours of 50ng/ml EGF stimulation as measured by scratch assays. J) KDM4A overexpressing RPE cells proliferate faster in response to a 48 hour treatment with 50ng/ml EGF. K) siRNA-mediated depletion of EGFR prevents increased EGF-stimulated (50ng/ml) cell growth in KDM4A overexpressing RPE cells. Error bars represents S.E.M. The * represents p=≤0.05 by two-tailed Student’s t-test. The arrowheads mark DNA FISH foci. The scale bars are 5um.