An aberrant spliced transcript of focal adhesion kinase is exclusively expressed in human breast cancer.

Ling Yao,Zhiming Shao,Xinmin Zheng,Shan Li,Xin Hu,Wenting Peng,Kai Li,Qiang Lin

doi:10.1186/1479-5876-12-136

Abstract

PurposeTo clarify the roles of a new aberrantly spliced transcript of FAK that lacks exon 26 (denoted -26-exon FAK) in human breast cancers.MethodsTranscripts of FAK expressed in 102 human breast tumor tissues and 52 corresponding normal tissues were analyzed by RT-PCR and DNA sequencing, as well as agarose gel electrophoresis. The cDNA of -26-exon FAK was cloned and expressed in MCF-10A cells, and then the kinase activity, cellular localization and migration capability of FAK were examined by western blotting, immunofluorescent staining and migration assays, respectively. The expression levels of FAK were analyzed by western blotting in MCF-7 cells treated with TNF-α or in MCF-10A cells upon serum deprivation. The MCF-10A cells transfected with a plasmid expressing -26-exon FAK were cultured in serum-free medium and cell apoptosis was analyzed by flow cytometry.ResultsThe -26-exon FAK transcript was exclusively present in human breast tumor tissues and the encoded protein possessed the same kinase activity, cellular localization and cell migration-promoting ability as wild-type FAK. In MCF-7 cells treated with TNF-α, and in MCF-10A cells upon serum deprivation, the -26-exon FAK was resistant to proteolysis while wild-type FAK was largely cleaved. In addition, the -26-exon FAK, but not wild-type FAK, inhibited cell apoptosis.ConclusionsThe -26-exon FAK transcript, which is exclusively expressed in human breast tumor tissues, encodes a protein that possesses the same kinase activity and biological function as the wild-type FAK, but because it is resistant to the caspase-mediated cleavage that induces the proteolysis of the wild-type form, it ultimately prevents apoptosis.

Highlights

FAK is a non-receptor tyrosine kinase that plays a key role at focal adhesion sites by promoting cell spreading, migration, and the transmission of anchorage-dependent anti-apoptotic signals [1]
The expression level and activity of FAK is closely associated with tumorigenesis, which indicates that FAK may be an important and useful cancer marker for future cancer diagnosis and therapy [1,2,8,9]
We found that an aberrantly spliced transcript of FAK kinase missing the exon 26 segment is exclusively expressed in human breast cancer and that this FAK mutant is resistant to caspase-mediated proteolysis, even though it possesses the same kinase activity as wild-type FAK

Summary

Introduction

FAK is a non-receptor tyrosine kinase that plays a key role at focal adhesion sites by promoting cell spreading, migration, and the transmission of anchorage-dependent anti-apoptotic signals [1]. Its autophosphorylation at the Tyr-397 site is an important event for maintaining the biological function of FAK, directly maintaining tumor growth [4], preventing apoptosis and promoting the survival of tumor cells [5,6,7], and modulating focal adhesion dynamics and the cellular cytoskeleton to facilitate cancer cell invasion and metastasis [8,9]. We analyzed the FAK expression at the RNA level in human breast cancer, aiming to explore whether there are alternatively spliced transcripts of FAK in tumors and to dissect the roles of these FAK transcripts in tumorigenesis

Methods

Results

Conclusion