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bioRxiv · 10.1101/2022.12.05.519126

De novo linear phosphorylation site motifs for BCR-ABL kinase revealed by phospho-proteomics in yeast

Smolnig, M.·Fasching, S.·Stelzl, U.
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Abstract

BCR-ABL is the oncogenic fusion product of tyrosine kinase ABL1 and a highly frequent driver of acute lymphocytic leukemia (ALL) and chronic myeloid leukemia (CML). The kinase activity of BCR-ABL is strongly elevated, however changes of substrate specificity in comparison to wild-type ABL1 kinase are less well characterized. Here, we heterologously expressed full-length BCR-ABL kinases in yeast. We exploited the proteome of living yeast as in vivo phospho-tyrosine substrate for assaying human kinases specificity. Phospho-proteomic analysis of ABL1 and BCR-ABL isoforms p190 and p210 yielded a high-confidence dataset of 1127 phospho-tyrosine sites on 821 yeast proteins. We used this data set to generate linear phosphorylation site motifs for ABL1 and the oncogenic ABL1 fusion proteins. The oncogenic kinases yielded a substantially different linear motif when compared to ABL1. Kinase set enrichment analysis with human pY-sites that have high linear motif scores well recalled BCR-ABL driven cancer cell lines from human phospho-proteome data sets. HighlightsO_LIFull-length BCR-ABL kinase expression in yeast C_LIO_LI1,127 pY-sites on 821 yeast proteins originating from ABL1 and BCR-ABL kinase activity C_LIO_LIDistinct linear kinase motif forABL1 and BCR-ABL p190 and p210 isoforms C_LIO_LIValidation of the kinase motifs through KSEA of human cancer cell line phospho-proteome data C_LI SignificanceThe fusion protein BCR-ABL is a predominant oncogene in leukemia observed at very high frequency in chronic myeloid leukemia (BCR-ABL p210) and B-cell acute lymphoid (BCR-ABL p190) leukemia. In both cases the malignant transformation is reliant on elevated tyrosine kinase activity, however ABL1 kinase activation as such is not sufficient. At least in part, leukemias may therefore be driven by a specificity switch of the fusion kinase in comparison to the wild-type ABL1. We investigated BCR-ABL kinase substrates in yeast through phospho-proteomics and established linear kinase phosphorylation-motifs for the full length BCR-ABL isoforms p210 and p190. These novel motifs differ substantially from the corresponding specificity determinants of wild-type ABL1 kinase, providing direct experimental evidence for altered phosphorylation-specificity of the oncogenic kinase fusion proteins.

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Smolnig, M., Fasching, S., Stelzl, U.. 2022-12-05. De novo linear phosphorylation site motifs for BCR-ABL kinase revealed by phospho-proteomics in yeast. https://doi.org/10.1101/2022.12.05.519126

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