Expression of HA-TEL-JAK2 transgene.

(A) Northern blot analysis. Total mRNA from diseased spleen (lane 1), thymus (lane 2), lymph nodes (lane 3), and from a control spleen (lane 4) were hybridized as indicated with probes specific for TEL or JAK2. The TEL-JAK2 transgene mRNA is indicated by a white arrowhead. Bands corresponding to the endogenous TEL and JAK2 transcripts are indicated by gray and black arrowheads, respectively. (B) Flow cytometric detection of HA-TEL-JAK2 protein expression in diseased thymus and lymph nodes. Double labeling with mouse anti-Thy1.2–PE and rabbit anti-HA plus antirabbit Ig-FITC antibodies revealed that Thy1.2⁺ cells express the HA epitope. For the thymus, FITC-fluorescence intensity in gated Thy1.2⁺ cells is represented for a transgenic animal (71-10, dark surface) and for a control animal (71-6, open surface). For diseased lymph nodes, labeling is with the anti-HA rabbit antibody plus antirabbit Ig-FITC (dark surface) or with antirabbit Ig-FITC alone (open surface). (C) Western blot analysis of cellular extracts from lymph nodes, thymus, and spleen from diseased and control animals. Extracts corresponding to the same number of cells from each organ were immunoprecipitated by anti-JAK2 antibody, separated by SDS-PAGE, and blotted on nitrocellulose. Blots were analyzed with either the anti-HA (12CA5, Boehringer) or anti-TEL23 antibodies (upper panel) or with the 4G10 phosphotyrosine-specific antibody (lower panel). Lanes 1 and 2: lymph nodes from transgenic animals 71-4 and 71-18. Lanes 3 and 5: thymus from transgenic mice 65 and 71-18. Lane 6: spleen from transgenic mouse 71-18. Lanes 4 and 7: control thymus and spleen, respectively. The HA-TEL-JAK2 protein is indicated by a white arrowhead.