Cancer Letters

Cancer Letters

Volume 415, 28 February 2018, Pages 106-116
Cancer Letters

Original Articles
TAp73 inhibits cell invasion and migration by directly activating KAI1 expression in colorectal carcinoma

https://doi.org/10.1016/j.canlet.2017.12.002Get rights and content

Highlights

  • TAp73 inhibits invasion, migration and metastasis of colorectal cancer cells.

  • TAp73 directly binds to the KAI1 promoter and induces its expression.

  • KAI1 is indispensable for TAp73-mediated inhibition of cell invasion and migration.

  • TAp73 and KAI1 expression levels are positively correlated in human colorectal cancer.

Abstract

p73 is a member of the p53 family of transcription factors and, like p53, plays a role as a tumor suppressor. p73 is involved in development, proliferation, apoptosis and metastasis. However, the precise molecular mechanisms underlying its function in inhibiting metastasis remain largely unknown. Here, we show that induction of TAp73 decreased invasion and migration activity of colorectal cancer cells, whereas knockdown of TAp73 led to increased invasion and migration activity. KAI1 was identified as a transcriptional target of TAp73 and its expression is indispensable for TAp73-mediated inhibition of cell invasion and migration. Furthermore, induction of TAp73 in colorectal cancer cells elevated KAI1 expression and decreased the frequency of hepatic metastasis in vivo. Whereas, the decreased invasion and migration activities caused by TAp73 induction were abrogated by knockdown of KAI1. Interestingly, TAp73 and KAI1 are overexpressed in primary colorectal cancers and a significant correlation between TAp73 and KAI1 expression was detected, but their expressions were significantly down-regulated in metastatic cancers. Taken together, our results support a novel role for TAp73 in controlling colorectal cancer cell invasion, migration and metastasis by regulating transcription of KAI1.

Introduction

p53 is a sequence-specific transcription factor that plays a central role as a tumor suppressor by regulating the transcription of target genes in response to intracellular and extracellular stimuli, but it is frequently mutated in a variety of human cancers [1]. p73 is a member of the p53 family and shares significant similarity at the amino acid level with p53 and p63. Specifically, the highest similarity is found in the DNA-binding domain (DBD), in which p73 has more than 60% similarity with p53 and p63. Thus, p73 binds to canonical p53-responsive elements to transactivate many p53 target genes [2]. However, unlike p53, p73 is rarely mutated in human tumors [3]. Furthermore, altered expression levels of various p73 isoforms, which primarily include transactivation (TA) isoforms and transactivation-deficient N-terminally truncated dominant negative (DN) isoforms, are frequently detected in different types of cancer [3]. TA isoforms of p73 form homo- or heteromeric complexes with p53 and/or p63 and directly bind to their response elements to transactivate target gene promoters, while DN isoforms of p73 are known to act as dominant-negative inhibitors of p53 family proteins by either directly blocking their binding to target gene promoters and/or by forming inactive heteromeric complexes with TA isoforms [4]. However, the physiological roles of p73 in tumor cells are not yet fully understood. Previous studies have demonstrated that TAp73 acts as a tumor suppressor in cell proliferation, cell cycle, apoptosis, invasion, migration and tumor metastasis through direct regulation of its unique or shared target genes with p53, whereas DNp73 can serve as an antagonist of p53, TAp63 and TAp73 [5], [6], [7], [8], [9]. On the other hand, previous studies also showed that TAp73 and DNp73 can act as oncogenes or tumor suppressors in cell proliferation and invasion [10], [11].

Tetraspanins are a family of membrane proteins with four transmembrane domains, and they interact with other tetraspanins and several different types of proteins, including integrins, membrane receptors, intracellular cytoskeletal proteins and signaling molecules [12]. Due to their multiple interactions with a diverse range of molecules, tetraspanins have been implicated in a wide range of biological processes, including cell motility, invasion, morphology, metastasis, proliferation and differentiation. KAI1/CD82 (Cluster of Differentiation 82) is a member of the tetraspanin superfamily and acts as a metastasis suppressor during the progression of a variety of solid tumors [13]. KAI1 also interacts with several cell surface receptors, such as EGFR, c-Met and Integrin β1, to inhibit their signaling transduction, resulting in suppression of invasion, migration and metastasis of cancer cells [14], [15], [16], [17]. Down-regulation of KAI1 has been observed during the progression of many types of human tumors and is correlated with poor survival in patients with those cancers. However, the underlying causes of the down-regulation of KAI1 in advanced or metastatic cancers remain unknown. The available evidence does not support mechanisms that involve mutations within the coding or promoter regions of KAI1, loss of heterozygosity, or CpG hypermethylation of the KAI1 promoter [18], [19], [20], [21], [22]. Rather, down-regulation of KAI1 expression is likely due to negative and/or positive alterations of its transcriptional activity.

In this study, we found that TAp73 and KAI1 were down-regulated in metastatic cancer tissues, which inhibited invasion, migration and EMT of colorectal cancer cells. We identified TAp73 as a positive transcriptional regulator that directly regulates the expression of KAI1 and found that KAI1 is indispensable for TAp73-mediated inhibition of cancer cell invasion, migration and metastasis.

Section snippets

Experimental procedures

Plasmid construction, western blot analysis, luciferase reporter assay, chromatin immunoprecipitation (ChIP) assay, electrophoretic mobility shift assay (EMSA), hepatic metastasis model, immunofluorescence, immunohistochemistry, RNA-seq analysis and ChIP-seq analysis are described in Supplementary Materials and Methods.

Tissue samples, cell culture and transfection

The biospecimens and data used for this study were provided by the Biobank of Chonnam National University Hwasun Hospital, a member of the Korea Biobank Network. HEK293T, HT29,

TAp73 inhibits invasion and migration of colorectal cancer cells

To analyze the role of TAp73 in cancer progression, a tetracycline-inducible TAp73 expression plasmid (pTet-TAp73) was constructed and transfected into HCT116 (p53-/-) and HT29 colorectal cancer cell lines. An empty vector (pTet) was transfected as a control. The TAp73 protein levels in pTet-TAp73-transfected cells were significantly increased by treatment with the tetracycline analog doxycycline (Dox) at 100 ng/ml compared with control cells (Fig. 1A). Conversely, application of gene-specific

Discussion

TAp73 is known to function as a tumor suppressor and regulate genomic integrity, cellular proliferation, and apoptosis. Recently, it has been reported and reviewed that TAp73 and its multiple isoforms control cell invasion, migration, metastasis and EMT-mediated cancer cell stemness by regulation of EPLIN, p57Kip2, FOXF1, POSTN, miR-885-5p, MIR34A and MIR3158 in a variety of cancer cells [8], [9], [11], [29], [30], [31], [32], [33]. However, its role in colorectal cancer invasion and metastasis

Conflicts of interest

The authors declare no conflict of interest.

Acknowledgments

This work was supported by the National Research Foundation of Korea grant (MRC, 2011-0030132) funded by the Korea government (MSIP). We thank Myung-Sook Park and Ji-Na Choi for conducting the experimental studies.

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