The effect of the peptidyl-prolyl isomerase pin1 on the expression and function of the tyrosine kinase lck

Judith Mikolajczak, Purdue University

Abstract

Lymphocyte cell kinase (Lck) is a 56 kDa non-receptor tyrosine kinase of the Src family. It plays an essential role in T cell development, signaling and differentiation. In addition to regulatory phosphorylation sites common to all Src kinases, the activity of Lck is regulated by phosphorylation of serine 59. Serine 59 is phosphorylated by Erk1/2, which prevents the binding of the tyrosine phosphatase SHP-1 that deactivates Lck. Phospho-serine/threonine-proline pS/T-P motifs cannot only act as regulatory sites through phosphorylation status but can also be isomerized by the peptidyl-prolyl isomerase Pin1, adding another layer of control to the system. Isomerization of pS/T-P peptide bonds by Pin1 can lead to changes in stability (enhance or decrease degradation) or activity in the target protein. Lck has two potential Pin1 target sites, S59-P60 and S194-P195, that have been shown to be phosphorylated in cells. Pin1 interacts with Lck in a stimulation-dependent manner as evidenced by co-immunoprecipitation and pull-down assays. Expression levels of Pin1 in HEK293T cells expressing serine to alanine mutants or WT-Lck leads to a Pin1-dose-dependent protein levels of WT-Lck protein levels while the S59A/S194-Lck mutant is unaffected by Pin1 levels. Knockdown of Pin1 increased WT-Lck levels while overexpression decreased Lck levels. In these cells, the half-life of WT-Lck depends on expression levels of Pin1. The decrease in WT-Lck protein levels is phosphorylation-dependent as blocking phosphorylation by small molecule inhibitors rescued Lck protein levels. In JCaM cells expressing Lck, overexpression of Pin1 shortens phospho-Erk1/2 signaling. In summary, these results indicate that there is a functional interaction between Lck and Pin1

Degree

Ph.D.

Advisors

Harrison, Purdue University.

Subject Area

Molecular biology|Cellular biology|Biochemistry

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