Fig. 3: Generation of the ETV6::NTRK3 chromosomal translocation in hES cells and differentiation to hES-MP. | Oncogenesis

Fig. 3: Generation of the ETV6::NTRK3 chromosomal translocation in hES cells and differentiation to hES-MP.

From: Generating in vitro models of NTRK-fusion mesenchymal neoplasia as tools for investigating kinase oncogenic activation and response to targeted therapy

Fig. 3

A Breakpoints within ETV6 on chromosome 12 and NTRK3 on chromosome 15 create the ETV6::NTRK3 t(12;15)(p13;q25). B A donor template containing a puromycin selectable marker under the control of a PGK promoter is inserted by HDR at the ETV6 and NTRK3 participating loci after induction of DSBs by sgRNA (scissors). The selectable marker is removed by Cre recombinase allowing the expression of the oncogenic fusion. C Dual color FISH analysis with telomeric (red) and centromeric (green) probes on ETV6 showing the ETV6::NTRK3 translocation on clone G2. D RT-PCR time course (days) for ETV6::NTRK3 fusion after Cre expression in hES and hES-MP cells. The fusion product is detectable in cells before Cre recombination indicating a partial splicing of PGK-Puro sequence. E Western blot analysis showing the ETV6::TRKC fusion protein and the TRKC phosphorylation in hES-MP clone G2-6 and G2 after expression of Cre recombinase. The total ETV6::TRKC protein was detected by pan-TRK antibodies and the TRKC phosphorylation by phospho TRKA/B antibodies (same used for detection of LMNA::pTRKA).

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