BCR-JAK2 is constitutively phosphorylated, triggering STAT5 activation which translocates to the nucleus and induces STAT5 target gene expression on transduced Ba/F3 cells.

<p>(<b>a</b>) Western blot analysis of Ba/F3 cells infected with BCR-JAK2, TEL-JAK2 or vector alone (mock). Cellular lysates were immunoprecipitated with anti-pTyr Ab and immunoblotted with anti-JAK2 Ab to see pJAK2 (upper panel: endogenous JAK2: 130 kDa, BCR-JAK2: 90 kDa, TEL-JAK2: 72 kDa). TEL-JAK2 protein expression is higher than BCR-JAK2 in transduced Ba/F3 cells. Whole cell lysates were probed with anti-pSTAT5 and anti-STAT5 Ab's (bottom). The expression levels of Tubulin were used as a loading control. (<b>b</b>) Enriched cytoplasmic (C) and nuclear (N) extracts from Ba/F3-mock,Ba/F3-BCR-JAK2 and Ba/F3-TEL-JAK2 cells were prepared from total cell lysates and blotted with anti-pSTAT5 Ab. TATA-Binding protein (TBP) and IkBα were used as loading controls for enriched nuclear and cytoplasmic fractions, respectively. (<b>c</b>) Expression of <i>Bcl-xL, Osm and Socs2</i> in Ba/F3-mock, Ba/F3-BCR-JAK2 and Ba/F3-TEL-JAK2 cells by qPCR. For comparative purposes, mRNA levels in untreated cells were normalized to 1. Bars represent fold changes of each gene normalized using <i>GADPH</i> levels. Results are given as mean ± SEM (n = 3).