Nischarin Inhibits LIM Kinase To Regulate Cofilin Phosphorylation and Cell Invasion

Abstract

Nischarin is a novel protein that regulates cell migration by inhibiting p21-activated kinase (PAK). LIM kinase (LIMK) is a downstream effector of PAK, and it is known to play an important role in cell invasion. Here we show that nischarin also associates with LIMK to inhibit LIMK activation, cofilin phosphorylation, and LIMK-mediated invasion of breast cancer cells, suggesting that nischarin regulates cell invasion by negative modulation of the LIMK/cofilin pathway. The amino terminus of nischarin binds to the PDZ and kinase domains of LIMK. Although LIMK activation enhances the interaction with nischarin, only phosphorylation of threonine 508 of LIMK is crucial for the interaction. Inhibition of endogenous nischarin expression by RNA interference stimulates breast cancer cell invasion. Also, nischarin small interfering RNA (siRNA) enhances cofilin phosphorylation. In addition, knock-down of nischarin showed branched projection actin structures. Collectively these data indicate that nischarin siRNA may enhance random migration, resulting in stimulation of invasion.

ACKNOWLEDGMENTS

We thank Ora Bernard of Australia for her generosity in providing LIMK constructs, Gordon Gill for LIMK mutants, and Lawrence Quilliam for ROCK constructs. We are grateful to Val Weaver for accommodating us in her lab at PENN right after the Hurricane Katrina evacuation. We also thank M. B. Hatten at Rockefeller for financial help and for arranging a visiting professorship in her lab for a period of 9 months. We thank our LSUHSC colleagues Andy Catling, Becky Worthylake, David Worthylake, and Ashok Pullikuth for their valuable suggestions, Charles Nichols for help with confocal microscopy, and Wayne Vedeckis for critical reading of the manuscript. Also, we thank our lab colleagues Somesh Baranwal for making the Flag-tagged LIMK T508V construct and Amelia Walch for technical help.

This work was supported by grants from NIH (CA 115706) and Susan Komen (BCTR0600278) and funds from the Louisiana Cancer Research Consortium to S.K.A.