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Multicolor monitoring of dysregulated protein kinases in chronic myelogenous leukemia
Qunzhao Wang1, Eric I Zimmerman, Alexei Toutchkine
1The University of North Carolina at Chapel Hill, 27599, USA.
Abstract:
The Bcr-Abl and Lyn protein tyrosine kinases have been separately linked to the emergence of imatinib resistance in patients with chronic myelogenous leukemia. We have developed fluorescent sensors for these kinases that are enzymatically and photophysically distinct, allowing us to simultaneously, yet separately, visualize the tyrosine kinase activities of both Abl and Lyn. Multicolor monitoring revealed that an imatinib-resistant cell line (MYL-R) displays a remarkable 13-fold enhancement in Lyn kinase activity relative to its imatinib-sensitive counterpart (MYL). By contrast, both cell lines display nearly identical Abl activities. The upregulation of Lyn kinase phosphotransferase activity in MYL-R cells is linked to an overexpression of the Lyn B isoform. Furthermore, MYL-R cells possess a 4-fold higher level of activated Lyn and 5-fold lower level of autoinhibited Lyn than MYL cells. Finally, studies with an activating SH2 ligand revealed that Lyn from imatinib-resistant MYL-R cells is primed and active, whereas Lyn from imatinib-sensitive cells is dependent upon phosphorylated SH2 ligands for activity.
Insights
Imatinib resistance in chronic myelogenous leukemia involves enhanced Lyn kinase activity, not Abl. Overexpression of the Lyn B isoform drives this resistance, making Lyn kinase constitutively active in resistant cells.
Area of Science:
- Biochemistry
- Molecular Biology
- Oncology
Background:
- Imatinib resistance in chronic myelogenous leukemia (CML) is a significant clinical challenge.
- Both Bcr-Abl and Lyn protein tyrosine kinases have been implicated in imatinib resistance.
Purpose of the Study:
- To develop and utilize distinct fluorescent sensors for simultaneous visualization of Abl and Lyn kinase activities.
- To investigate the differential roles of Abl and Lyn kinases in imatinib-sensitive versus imatinib-resistant CML cells.
Main Methods:
- Development of enzymatically and photophysically distinct fluorescent sensors for Abl and Lyn kinases.
- Multicolor monitoring of kinase activities in imatinib-sensitive (MYL) and resistant (MYL-R) CML cell lines.
- Analysis of Lyn kinase isoforms, activation states, and dependence on SH2 ligands.
Main Results:
- MYL-R cells exhibit a 13-fold increase in Lyn kinase activity compared to MYL cells, while Abl activities remain similar.
- Lyn kinase upregulation in MYL-R cells is associated with overexpression of the Lyn B isoform.
- MYL-R cells show higher activated Lyn and lower autoinhibited Lyn, with Lyn being constitutively active, unlike in MYL cells.
Conclusions:
- Enhanced Lyn kinase activity, specifically the Lyn B isoform, is a key driver of imatinib resistance in CML.
- Lyn kinase in resistant cells is constitutively active and independent of SH2 ligand binding, contrasting with Lyn in sensitive cells.
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