Subcellular localization of glypican-5 is associated with dynamic motility of the human mesenchymal stem cell line U3DT.

Masao Takeuchi,Ritsuko Yamaguchi,Jun K Takeuchi,Tomoyo Takai,Ken-Ichi Akagi,Kikuko Takeuchi,Tetsushi Furukawa,Feng Zhao

doi:10.1371/journal.pone.0226538

Abstract

Glypican-5 (GPC5) is a heparan sulfate proteoglycan (HSPG) localized to the plasma membrane. We previously reported that in the human mesenchymal stem cell line UE6E7T-3, GPC5 is overexpressed in association with transformation and promotes cell proliferation by acting as a co-receptor for Sonic hedgehog signaling. In this study, we found using immunofluorescence microscopy that in transformed cells (U3DT), GPC5 localized not only at primary cilia on the cell surface, but also at the leading edge of migrating cells, at the intercellular bridge and blebs during cytokinesis, and in extracellular vesicles. In each subcellular region, GPC5 colocalized with fibroblast growth factor receptor (FGFR) and the small GTPases Rab11 and ARF6, indicating that GPC5 is delivered to these regions by Rab11-associated recycling endosomes. These colocalizations suggest that GPC5 plays an important role in FGF2 stimulation of cell migration, which was abrogated by knockdown of GPC5. Our findings indicate that GPC5 plays a role in regulation of U3DT cell migration and provides several insights into the functions of GPC5 that could be elucidated by future studies.

Highlights

Glypicans (GPCs) and syndecans (SDCs), which are heparan sulfate proteoglycans (HSPGs) displayed on the surface of most mammalian cells, have long been thought to act as co-receptors for cell-surface receptors in several signaling pathways, including Hedgehog (Hh), Wnt, bone morphogenetic protein (BMP), and fibroblast growth factor (FGF) signaling [1]
We demonstrated that GPC5 participates in U3DT cell proliferation, HSPGs have numerous cellular functions related to modulation of the Hh, Wnt, BMP, and FGF signaling pathways, depending on cell and tissue type
GPC5 localized to primary cilia on cell surface, and to the leading edge of migrating cells, the intercellular bridge, and blebs during cytokinesis, and extracellular vesicles (EVs). These localizations suggest that GPC5 play an important role in cell migration, which was abrogated by knockdown of GPC5

Summary

Introduction

Glypicans (GPCs) and syndecans (SDCs), which are heparan sulfate proteoglycans (HSPGs) displayed on the surface of most mammalian cells, have long been thought to act as co-receptors for cell-surface receptors in several signaling pathways, including Hedgehog (Hh), Wnt, bone morphogenetic protein (BMP), and fibroblast growth factor (FGF) signaling [1]. Glypican-5 (GPC5) is dramatically overexpressed in association with transformation after prolonged culture of the human mesenchymal stem cell line UE6E7T-3, and that knockdown of GPC5 expression decreases cell proliferation [3]. GPC5 is overexpressed in rhabdomyosarcomas (RMS), and down-regulation of GPC5 expression by RNAi decreases the proliferation rate of RMS cells [4]. Subsequent work showed that GPC5 stimulates RMS cell proliferation by activating Hh signaling by promoting the binding of the ligand Sonic hedgehog (Shh) to Patched (Ptc), the Hh receptor on the cell surface [5]. Overexpression of GPC5 inhibits prostate [8] and lung cancer [9] cell proliferation. In non-small cell lung cancer, some reports have suggested that GPC5 is a tumor promoter [10], whereas others insist that it is a tumor suppressor [11, 12]

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