Protein Tyrosine Phosphorylation in Haematopoietic Cancers and the Functional Significance of Phospho- Lyn SH2 Domain

Protein-tyrosine phosphorylation (pY) is a minor but important protein post-translational modification that modulates a wide range of cellular functions and is involved in cancer. Dysregulation of tyrosine kinases (TKs) and protein-tyrosine phosphatases (PTPs) have been observed in multiple myeloma (MM) and acute myeloid leukemia (AML) and is a subject of study. Using recently developed mass spectrometry-based proteomics techniques, quantitative PTP expression and cellular pY profiles were generated for MM cell lines and mouse xenograft tumors, as well as primary AML samples. Integrated comprehensive analyses on these data implicated a subset of TKs and PTPs in MM and AML, with valuable insights gained on the dynamic regulation of pY in biological systems. In particular, I propose a model that describes the cellular pY state as a functional output of the total activated TKs and PTPs in the cell. My results show that the global pY profile in the cancer models is quantitatively related to the cellular levels of activated TKs and PTPs. Furthermore, the identity of the implicated TK/PTPs is system-dependent, demonstrating context-dependent regulation of pY. To further understand pY regulation, I studied the phosphorylation of a conserved tyrosine in the Src homology 2 (SH2) domains of Src family kinases, which was frequently observed in human cancer specimens and regulated in cancer-derived cell lines. Using the Lyn SH2 domain as a model, I discovered that when this tyrosine (Y194) is phosphorylated, the domain has reduced ability to interact in vitro with phosphopeptides and phosphoproteins. Sequence analysis of the binding motifs revealed that phosphorylation at Y194 decreased the selectivity of Lyn SH2 for the third residue C-terminal to the pY of the ligand. Together, these data show that tyrosyl phosphorylation alters the substrate binding profile of Lyn SH2 and may potentially affect Lyn kinase signaling. This may be a general mechanism for all Src family kinases and represents another layer of the complexity of pY regulation.Ph.D