Inhibition of the c-Jun N-terminal Kinase/AP-1 and NF-κB Pathways by PICOT, a Novel Protein Kinase C-interacting Protein with a Thioredoxin Homology Domain

Stephan Witte,Martin Villalba,Kun Bi,Yuhong Liu,Noah Isakov,Amnon Altman

doi:10.1074/jbc.275.3.1902

Abstract

Protein kinase C-theta (PKCtheta) is a Ca(2+)-independent PKC isoform that is selectively expressed in T lymphocytes (and muscle), and is thought to play an important role in T cell receptor-induced activation. To gain a better understanding of the function and regulation of PKCtheta, we have employed the yeast two-hybrid system to identify PKCtheta-interacting proteins. We report the isolation and characterization of a cDNA encoding a novel 335-amino acid (37. 5-kDa) PKCtheta-interacting protein termed PICOT (for PKC-interacting cousin of thioredoxin). PICOT is expressed in various tissues, including in T cells, where it colocalizes with PKCtheta. PICOT displays an N-terminal thioredoxin homology domain, which is required for the interaction with PKC. Comparison of the unique C-terminal region of PICOT with expressed sequence tag data bases revealed two tandem repeats of a novel domain that is highly conserved from plants to mammals. Transient overexpression of full-length PICOT (but not its N- or C-terminal fragments) in T cells inhibited the activation of c-Jun N-terminal kinase (but not extracellular signal-regulated kinase), and the transcription factors AP-1 or NF-kappaB. These findings suggest that PICOT and its evolutionary conserved homologues may interact with PKC-related kinases in multiple organisms and, second, that it plays a role in regulating the function of the thioredoxin system.

Highlights

Protein kinase C-␪ (PKC␪) is a Ca2؉-independent protein kinase C (PKC) isoform that is selectively expressed in T lymphocytes, and is thought to play an important role in T cell receptor-induced activation
These findings suggest that PICOT and its evolutionary conserved homologues may interact with PKC-related kinases in multiple organisms and, second, that it plays a role in regulating the function of the thioredoxin system
Selective Inhibition of Jun N-terminal kinase (JNK) Activation by PICOT—As PICOT was identified on the basis of its interaction with PKC␪, we examined the effect of transient PICOT overexpression on the activation of the stress-activated protein kinase JNK, since it represents a selective target of PKC␪ in the T cell antigen receptor (TCR)/CD28 signaling pathway [7, 8]

Summary

Introduction

Protein kinase C-␪ (PKC␪) is a Ca2؉-independent PKC isoform that is selectively expressed in T lymphocytes (and muscle), and is thought to play an important role in T cell receptor-induced activation. Transient overexpression of full-length PICOT (but not its N- or C-terminal fragments) in T cells inhibited the activation of c-Jun N-terminal kinase (but not extracellular signal-regulated kinase), and the transcription factors AP-1 or NF-␬B These findings suggest that PICOT and its evolutionary conserved homologues may interact with PKC-related kinases in multiple organisms and, second, that it plays a role in regulating the function of the thioredoxin system. We describe the isolation of a novel PKC␪-interacting protein having a unique domain structure, which consists of an N-terminal thioredoxin (Trx)-homologous domain followed by two tandem repeats of a novel, evolutionary conserved protein domain. We have termed this protein PICOT (for PKC-interacting cousin of thioredoxin). Transient overexpression of PICOT inhibited the activation of JNK and two transcription factors i.e. AP-1 and NF-␬B, induced by PKC␪ or by combinations of T cell-activating stimuli, suggesting that this novel protein plays an important role in regulating T cell activation and the function of PKC␪

Methods

Results

Conclusion