The RGD motif is involved in CD97/ADGRE5-promoted cell adhesion and viability of HT1080 cells

Wen-Ye Tjong,Hsi-Hsien Lin

doi:10.1038/s41598-018-38045-w

Wen-Ye Tjong, Hsi-Hsien Lin

Open Access

https://doi.org/10.1038/s41598-018-38045-w

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Abstract

CD97/ADGRE5 is an adhesion G protein-coupled receptor (aGPCR) involved in tumor cell adhesion, migration, angiogenesis, and apoptosis. CD97 has been shown previously to stimulate angiogenesis by interacting with integrins on endothelial cells via an Arginine-Glycine-Aspartic acid (RGD) motif. In this report, the role of the RGD motif in tumor cell adhesion and apoptosis was investigated using a previously-established HT1080 cell-based system. We found that the RGD motif is critical in CD97-promoted cell adhesion, in part due to the up-regulation of αvβ5 and α2β1 integrins, and that CD97 mediates its anti-apoptotic effect in extrinsic apoptosis via RGD-dependent cell adhesion. In contrast, CD97-modulated anti-apoptotic effect in intrinsic apoptosis is mediated by RGD-independent, N-cadherin-induced homotypic cell aggregation. Hence, CD97 promotes tumorigenesis via RGD-dependent and -independent mechanisms.

Highlights

Adhesion-class G protein-coupled receptors are evolutionarily conserved cell surface proteins characterized by a long N-terminal extracellular domain (ECD) linked to a seven-span transmembrane (7TM) region
EGF-like module-containing mucin-like hormone receptor-like 2 (EMR2) is included as a control because it shares with CD97 a 97% sequence identity in the epidermal growth factor (EGF)-like domains, but does not contain a RGD motif in its GPCR-Autoproteolysis INducing (GAIN) domain[26]
Flow cytometry and western blot analyses verified the expression of the recombinant CD97 and EMR2 proteins was up-regulated in the designated groups of stable HT1080 cells to a consistent and comparable level when compared to the control HT1080-Neo cells (Fig. 1B,C)

Summary

Introduction

Adhesion-class G protein-coupled receptors (aGPCRs) are evolutionarily conserved cell surface proteins characterized by a long N-terminal extracellular domain (ECD) linked to a seven-span transmembrane (7TM) region. Three CD97 receptor isoforms containing different EGF-like motifs, namely CD97(EGF/125), CD97(EGF/1235) and CD97(EGF/1–5), were identified[11] These distinct CD97 isoforms interact with four endogenous cellular ligands including CD55 (DAF), α5β1 integrin, CD90 (Thy-1), and chondroitin sulphate mostly in an isoform-restricted manner[12,13,14,15]. As the RGD peptide itself is a well-known cell-adhesion motif capable of modulating numerous cellular functions[24,25], we aim to delineate the possible role of the RGD motif in CD97-modulated tumor cell adhesion and apoptosis. To this end, we adapted the previously-established HT1080 cell-based experimental system utilizing site-directed mutants, chimeric receptors, and specific function-blocking peptides. These findings contribute to the functional insights of CD97-regulated tumorigenesis

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Results

Conclusion