biostudies-otherUnknownJosephina SampsonNational Research Foundation of Korea (NRF)Cancer Research UK (CRUK)Wellcome TrustEMBO Reportshttps://www.ebi.ac.uk/biostudies/studies/S-SCDT-EMBOR-2021-53693V1Variants of the oncogenic EML4-ALK fusion protein contain a similar region of ALK encompassing the kinase domain, but different portions of EML4. Here we show that EML4-ALK V1 and V3 proteins form cytoplasmic foci that contain components of the MAPK, PLC and PI3K signalling pathways. The ALK inhibitors ceritinib and lorlatinib dissolve these foci and EML4-ALK V3 but not V1 protein re-localises to microtubules, an effect recapitulated in a catalytically-inactive EML4-ALK mutant. Mutations that promote a constitutively active ALK stabilise the cytoplasmic foci even in the presence of these inhibitors. In contrast, the inhibitor alectinib increases foci formation of both wild-type and catalytically-inactive EML4-ALK V3 proteins, but not a Lys-Glu salt-bridge mutant. We propose that EML4-ALK biostudies-otherC24461/A23302WT104918MANo.2020R1A2C2006815Prof. Richard BaylissJene ChoiJosephina SampsonDr. Mark, William RichardsDr. Andrew, M. FryfalsePhase-separated foci of EML4-ALK facilitate signalling and depend upon an active kinase conformationVariants of the oncogenic EML4-ALK fusion protein contain a similar region of ALK encompassing the kinase domain, but different portions of EML4. Here we show that EML4-ALK V1 and V3 proteins form cytoplasmic foci that contain components of the MAPK, PLC and PI3K signalling pathways. The ALK inhibitors ceritinib and lorlatinib dissolve these foci and EML4-ALK V3 but not V1 protein re-localises to microtubules, an effect recapitulated in a catalytically-inactive EML4-ALK mutant. Mutations that promote a constitutively active ALK stabilise the cytoplasmic foci even in the presence of these inhibitors. In contrast, the inhibitor alectinib increases foci formation of both wild-type and catalytically-inactive EML4-ALK V3 proteins, but not a Lys-Glu salt-bridge mutant. We propose that EML4-ALK 2022-03-01T00:00:00Z2022-03-01T21:00:34.622Z2022-03-01T21:00:34.622ZS-SCDT-EMBOR-2021-53693V110.15252/embr.202153693