HAS3-induced extracellular vesicles from melanoma cells stimulate IHH mediated c-Myc upregulation via the hedgehog signaling pathway in target cells

Uma Thanigai Arasu,Pia R-M Siljander,Sanna Pasonen-Seppänen,Riikka Kärnä,Elisa Lázaro-Ibáñez,Maciej Lalowski,Sanna Oikari,Kirsi Rilla,Kai Härkönen,Ashik Jawahar Deen,Petri Mäkinen,Anna-Liisa Levonen,Sami Heikkinen

doi:10.1007/s00018-019-03399-5

Uma Thanigai Arasu, Pia R-M Siljander + Show 11 more

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https://doi.org/10.1007/s00018-019-03399-5

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Abstract

Intercellular communication is fundamental to the survival and maintenance of all multicellular systems, whereas dysregulation of communication pathways can drive cancer progression. Extracellular vesicles (EVs) are mediators of cell-to-cell communication that regulate a variety of cellular processes involved in tumor progression. Overexpression of a specific plasma membrane enzyme, hyaluronan synthase 3 (HAS3), is one of the factors that can induce EV shedding. HAS3, and particularly its product hyaluronan (HA), are carried by EVs and are known to be associated with the tumorigenic properties of cancer cells. To elucidate the specific effects of cancerous, HAS3-induced EVs on target cells, normal human keratinocytes and melanoma cells were treated with EVs derived from GFP-HAS3 expressing metastatic melanoma cells. We found that the HA receptor CD44 participated in the regulation of EV binding to target cells. Furthermore, GFP-HAS3-positive EVs induced HA secretion, proliferation and invasion of target cells. Our results suggest that HAS3-EVs contains increased quantities of IHH, which activates the target cell hedgehog signaling cascade and leads to the activation of c-Myc and regulation of claspin expression. This signaling of IHH in HAS3-EVs resulted in increased cell proliferation. Claspin immunostaining correlated with HA content in human cutaneous melanocytic lesions, supporting our in vitro findings and suggesting a reciprocal regulation between claspin expression and HA synthesis. This study shows for the first time that EVs originating from HAS3 overexpressing cells carry mitogenic signals that induce proliferation and epithelial-to-mesenchymal transition in target cells. The study also identifies a novel feedback regulation between the hedgehog signaling pathway and HA metabolism in melanoma, mediated by EVs carrying HA and IHH.

Highlights

Cancer cells depend on bidirectional communication between other cells and the extracellular environment for sustained growth, invasion and metastasis
We demonstrated that Extracellular vesicles (EVs) released from hyaluronan synthase 3 (HAS3) overexpressing metastatic melanoma cells (MV3) interacted with keratinocyte (HaCaT) and melanoma (WM115) cells and induced proliferation and expression of EMT markers, similar to that in tumorigenesis (Fig. 8)
HAS3-EVs increased the expression of c-Myc, an oncogenic factor activated in many different cancers types [21, 54]

Summary

Introduction

Cancer cells depend on bidirectional communication between other cells and the extracellular environment for sustained growth, invasion and metastasis. Extracellular vesicles (EVs) represent one of the numerous pathways for cellular communication [1]. EVs are membrane-enclosed particles that are shed from the cell surface, carry bioactive molecules [2] and contribute to, for example, the communication between tumor cells and the extracellular matrix [3]. Extensive research has demonstrated a role for EVs as effective intercellular messengers in cancer [4,5,6]. EVs can stimulate malignant transformation of target cells either by paracrine signaling or by fusion and internalization with the recipient cells [5]. The importance of EVs in cancer progression was first observed in 1980 when vesicles from highly metastatic melanoma cells (B16 mouse-F10) enabled poorly metastatic cells (mouse B16-F1) to metastasize to the lungs [7]

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