The lymphocyte tyrosine kinase Lck and its binding partners at mitosis
Abstract
The lymphocyte specific tyrosine kinase, Lck, is critical for lymphocyte development and activation. Lck is differentially phosphorylated with respect to the cell cycle. In cells arrested at mitosis, Lck exhibits an additional isoform with retarded electrophoretic mobility, a characteristic of the serine phosphorylation of Lck. Lck specifically associates with the active mitotic isoform of the cyclin-dependent kinase, Cdc2. Two other members of the Src family, the tyrosine kinases, Lyn and Fyn also associate with Cdc2 at mitosis. Lck does not bind the Cyclin B component of active Cdc2 directly. The association of Lck with Cdc2 is mediated by the SH3 domain of Lck. Complexes of Lck and Cdc2 are localized to specific cellular compartments. Phosphorylation of Lck by Cdc2 in vitro results in the phosphorylation of Lck with a resultant retardation in its electrophoretic mobility. Thus, it is likely that Cdc2 mediates the mitotic phosphorylation of Lck. Attempts to elucidate the functional significance of the mitotic phosphorylation of Lck led to the discovery of proteins that associated with Lck specifically at mitosis. Sam 68, an RNA-binding protein is tyrosine phosphorylated at mitosis and associates with the SH3 domain of Lck. Fyn and Lyn, other tyrosine kinases of the Src family also associated with Sam 68. An 85 kDa protein is tyrosine phosphorylated at mitosis and can be immunodepleted from mitotic cell lysates by virtue of its interaction with Lck. The serine/threonine kinase, Raf-1, is hyperphosphorylated at mitosis on serine residues with a resultant retardation of its electrophoretic mobility. Raf-1 is also activated at mitosis. Both, hyperphosphorylation and activation of Raf-1 are abrogated in Lck-deficient T-cell derived cell lines. This suggests the presence of an Lck-dependent pathway in the activation of Raf-1 in these cells. The study demonstrates the cell cycle-specific regulation of the lymphocyte tyrosine kinase Lck and describes a novel role for Lck, that of mediating tyrosine phosphorylation of proteins specifically at mitosis. Moreover, the results suggest the presence of distinct signaling pathways at mitosis, that involve tyrosine kinases such as Lck.
Degree
Ph.D.
Advisors
Harrison, Purdue University.
Subject Area
Cellular biology|Biochemistry|Immunology
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