Since the discovery in the early 1980's that PKC, or Protein Kinase C, was the major receptor for tumor-promoting phorbol esters1, the biological activity and regulation of PKC has been actively researched. The regulation of PKC activity is of fundamental importance to cancer biology, and PKC has emerged as an important therapeutic target.

The human PIN1 gene encodes an essential nuclear peptidyl-prolyl cis/trans isomerase, which is involved in the regulation of mitosis. Work from other labs has shown that deletion of PIN1 from HeLa cells induces mitotic arrest, whereas overexpression of PIN1 leads to a G2 phase arrest. The potential importance of Pin1 in cancer is also demonstrated by the observation that loss of Pin1 function (in mouse knockouts) leads to a phenotype resembling a cyclin D1-null phenotype. Furthermore, there is a requirement of Pin1 for the replication checkpoint to function, which is of major importance in allowing proper repair of DNA lesions prior to replication.

Pin1 has been shown to interact with the C-terminal domain of PKC. In addition to continuing to investigate the interplay of Pin1 and HSP70 controlling the isomerization state of the PKC-CT, I am in the process of determining the structure of the Pin1:PKC-CT complex. Importantly, which residues are the main determinants for interaction with Pin1, are the binding sites of HSP70 and Pin1 overlapping? Why are we seeing isomerization at Pro637 instead of Pro642? In addition to this providing more information about PKC, this will be the first structure of Pin1 with a substrate longer than 5 residues and may provide information about the communication between domains of Pin1. Futher experiments will examine the effects of the isomerization on the catalytic activity and biological signaling of PKC.

This work leads will thus lead to a greater understanding of these two enzymes, PKC and Pin1, which are both known to play important roles in cell regulation and tumor promotion. My work has discovered Pin1 as a new interaction partner for PKC, and also helps to explain the role of HSP70. I will expand upon these initial findings and demonstrate that both the phosphorylation sites and proline cis-trans isomerization states act as regulatory switches and the interplay between HSP70 and Pin1 are crucial to regulate the potential carcinogenic activity of PKC.

PUBLICATIONS (resulting from this training)

Abrahamsen H, Kruse N, Jennings PA, Newton AC. (2007) The Peptidly-Prolyl Isomerase Pin1 Controls the Down-reglation of Protein Kinase. Molecular Cell, In Review (revised manuscript).

Kruse N, Capraro D, Jennings PA (2007) The carboxyl tail of PKC is dynamically unstructured and undergoes a pin1 catalyzed isomerization. In Preparation.