The anoikis effector Bit1 displays tumor suppressive function in lung cancer cells.

Xin Yao,Shubha Kale Ireland,Brandi Temple,Renwei Chen,Tri Pham,Scott Jennings,Hector Biliran,Mya Davis,Ian Davenport,Lucia R Languino

doi:10.1371/journal.pone.0101564

Abstract

The mitochondrial Bit1 (Bcl-2 inhibitor of transcription 1) protein is a part of an apoptotic pathway that is uniquely regulated by integrin-mediated attachment. As an anoikis effector, Bit1 is released into the cytoplasm following loss of cell attachment and induces a caspase-independent form of apoptosis. Considering that anoikis resistance is a critical determinant of transformation, we hypothesized that cancer cells may circumvent the Bit1 apoptotic pathway to attain anchorage-independence and tumorigenic potential. Here, we provide the first evidence of the tumor suppressive effect of Bit1 through a mechanism involving anoikis induction in human lung adenocarcinoma derived A549 cells. Restitution of Bit1 in anoikis resistant A549 cells is sufficient to induce detachment induced-apoptosis despite defect in caspase activation and impairs their anchorage-independent growth. Conversely, stable downregulation of Bit1 in these cells significantly enhances their anoikis resistance and anchorage-independent growth. The Bit1 knockdown cells exhibit significantly enhanced tumorigenecity in vivo. It has been previously shown that the nuclear TLE1 corepressor is a putative oncogene in lung cancer, and we show here that TLE1 blocks Bit1 mediated anoikis in part by sequestering the pro-apoptotic partner of Bit1, the Amino-terminal Enhancer of Split (AES) protein, in the nucleus. Taken together, these findings suggest a tumor suppressive role of the caspase-independent anoikis effector Bit1 in lung cancer. Consistent with its role as a tumor suppressor, we have found that Bit1 is downregulated in human non-small cell lung cancer (NSCLC) tissues.

Highlights

A critical determinant of a malignant epithelial phenotype is anchorage independence
Following loss of cell attachment, Bit1 is released to the cytosol to induce a caspase-independent form of apoptosis, which can only be suppressed by integrin-mediated cell adhesion
Since anoikis resistance is a critical determinant of transformation, we hypothesize that suppression of Bit1 function is an alternative mechanism by which malignant cells resist anoikis and endow themselves with enhanced anchorage-independent growth potential in vitro and tumorigenecity in vivo

Summary

Introduction

A critical determinant of a malignant epithelial phenotype is anchorage independence. Following detachment-induced loss of integrin-mediated survival signalling, the intrinsic apoptotic pathway is triggered through activation of the pro-apoptotic members of the BCL family of proteins (including Bax, Bad, Bid and Bim) These proteins trigger permeabilization of the outer mitochondrial membrane leading to cytosolic release of cytochrome c and downstream activation of caspase enzymes [5,6]. The death receptor (extrinsic) pathway depends upon the binding of death ligands (Fas or TRAIL) to death receptor and utilizes the formation of a deathinducing signaling complex (DISC) in activating the downstream caspase-8 This death receptor-mediated caspase 8 activation may effect destabilization of the mitochondrial membrane leading to apoptosis [8,9]. The alternative caspase-independent mode of anoikis is an important therapeutic target in tumors that exhibit a disabled caspase-dependent apoptosis pathway

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