Additional file 2: Figure S2. of Calreticulin-mutant proteins induce megakaryocytic signaling to transform hematopoietic cells and undergo accelerated degradation and Golgi-mediated secretion

Regulation of megakaryocytic factors by CALR mutants in murine and human cell lines. a Detection of Fli-1 and Ets-1 or b Gata1 mRNA expression by RT-qPCR in the indicated 32D and HL60e cells. The experiments were performed in triplicates. Mean and SD are indicated. ***P < 0.001. c 32D expressing WT CALR, CALR del52, CALR ins5 cells, and outgrown 32D del52 cells were used to prepare lysates. SDS-PAGE and Western blotting were performed. An antibody for the detection of GATA-1 protein was used for immunostaining. GAPDH served as loading control and was used for the calculation of GATA-1 expression ratios. d RT-qPCR was used to detect Mpl, EpoR, and G-csfR mRNA amounts after RNA isolation of the indicated 32D cells followed by cDNA synthesis. Expression is depicted in percentage to Gapdh. Measurements were done in triplicates. Mean and SD are indicated. *P < 0.05. e Analysis of endogenous Mpl expression by RT-qPCR to compare 32D EV, WT CALR, CALR del52, and CALR ins5 expressing cells +/− ectopic MPL expression. Experiments were performed in triplicates. Mean and SD are indicated. **P < 0.01, ***P < 0.001. (PDF 55 kb