ERK-DBP induction of senescence depends upon interaction with Egr1. (A) Real-time quantitative polymerase chain reaction analysis of Egr1 expression in pMIG or pMIG-JAK2V617F–transduced HSPCs from ERK2-WT–, -KO–, and -DBP–mutant mice that were cultured for 3 days. (B) The physical interaction between ERK2-DBP domain and the DEF motif of Egr1. (C) Physical association of ERK2 and Egr1 after PMA stimuli. Anti-FLAG anti-ERK immunoblots were performed on input and anti-FLAG immunoprecipitates (immppt) from protein extracts of SCID.adh cells transduced with pMICherry empty vector (EV), FLAG-Egr1 (WT), and FLAG-Egr1^Y253A mutant. (D-E) Colony-forming assays were performed on WT and Egr1^−/− HSPCs transduced with the indicated constructs (pMIG, EV; pMIG-JAK2V617F; pMICherry, EV; pMICherry-Egr1; pMICherry-Egr1Y253A). Sorted cells (1 × 10³) were plated on M3434 medium and scored after 7 days in culture. (D) Representative images of cultures and a graphic depiction of the mean ± standard deviation (SD) colonies are shown. **P < .01; ***P < .001. (F-G) Representative images of SA-β-gal staining and the percentages of SA-β-gal⁺ cells were determined in cultures equivalent to those in panel D. All results are from ≥3 independent experiments. Data are expressed as the mean ± SD. **P < .01.