Transgelin interacts with PARP1 and affects Rho signaling pathway in human colon cancer cells

Abstract

<jats:title>Abstract</jats:title> <jats:p><jats:bold>Background:</jats:bold> Transgelin, an actin-binding protein, is associated with the cytoskeleton remodeling. Our previous studies found that transgelin was up-regulated in node-positive colorectal cancer versus in node-negative disease. Over-expression<jats:italic> </jats:italic>of<jats:italic> TAGLN</jats:italic> affected the expression of 256 downstream transcripts and increased the metastatic potential of colon cancer cells in vitro and in vivo. This study aims to explore the mechanisms that transgelin participates in the metastasis of colon cancer cells.<jats:bold>Methods:</jats:bold> Immunofluorescence and immunoblotting analysis were used to determine the cellular localization of the endogenous and exogenous transgelin in colon cancer cells. Co-immunoprecipitation and subsequent high performance liquid chromatography/tandem mass spectrometry were performed to identify the proteins potentially interacting with transgelin. Bioinformatics methods were used to analyze the 256 downstream transcripts regulated by transgelin to discriminate the specific key genes and signaling pathways. GCBI tools were used to predict the potential transcription factor(s) for these key genes. The predicted transcription factors were matching to the proteins that have been identified to interact with transgelin. The interaction between transgelin and these transcription factors was verified by co-immunoprecipitation and immunofluorescence.<jats:bold>Results:</jats:bold> Transgelin was found to localize both in cytoplasm and nucleus of colon cancer cells. 297 proteins have been identified to interact with transgelin by co-immunoprecipitation and subsequent high performance liquid chromatography/mass spectrometry. Over-expression of <jats:italic>TAGLN</jats:italic> could lead to differential expression of 184 downstream genes. The network of gene-encoded proteins was constructed in which 7 genes (<jats:italic>CALM1, MYO1F, NCKIPSD, PLK4, RAC1, WAS</jats:italic> and <jats:italic>WIPF1</jats:italic>) have been discriminated as key genes by network topology analysis. They are mostly involved in the Rho signaling pathway. Poly ADP-ribose polymerase-1 (PARP1) was predicted as the unique transcription factor for the key genes and concurrently matching to the DNA-binding proteins potentially interacting with transgelin. Immunoprecipitation and immunofluorescence validated the interaction between PARP1and transgelin in human RKO colon cancer cells. <jats:bold>Conclusions:</jats:bold> The results of this study suggest that transgelin binds to PARP1 and regulates the expression of downstream key genes which are mainly involved in Rho signaling pathway, thus participates in the metastasis of colon cancer.</jats:p>