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. 2021 Jan 19;137(23):3259–3271. doi: 10.1182/blood.2020009082

Figure 5.

Figure 5.

A feedback loop couples MEK/ERK hyperactivation and Rasgrp1 downregulation. (A-B) Representative hemoxylin and eosin-stained images (A) and flow cytometric plots of 2 cell lines (B387 and B159) (B) derived from Kras−/−; NrasQ61R/+ thymocytes. (C) Cell growth was measured using Cell TiterGlo assay. (D) B387 and B159 were treated with DMSO, 2 µM MEK inhibitor (AZD6244), 2 µM JAK inhibitor (CYT387), 100 nM SRC inhibitor (dasatinib), or 10 µM JNK inhibitor (SP600125) for 18 hours. Rasgrp1 and Ets1 mRNA levels were quantified using quantitative reverse transcription polymerase chain reaction. (E) Western blot analysis of Rasgrp1 and Ets1 protein levels in the absence or presence of AZD6244. (F-H) Jurkat cells, a human T-ALL cell line, stably expressed a scrambled shRNA or a RASGRP1 shRNA. (F) Western blot analysis of Rasgrp1, ERK1/2 and pERK1/2 protein levels. (G) Cell-cycle analysis. (H) Jurkat T cells were seeded at the same density and cell numbers were counted 48 hours later. P values were calculated with a 2-tailed Student t test. *P < .05; ***P < .001.