Ukr.Biochem.J. 2021; Volume 93, Issue 6, Nov-Dec, pp. 46-54


Extracellular vesicles produced by mouse breast adenocarcinoma 4T1 cells with up- or down-regulation of adaptor protein Ruk/CIN85 differentially modulate the biological properties of 4T1 WT cells

A. Yu. Zhyvolozhnyi1,2*, I. R. Horak1, D. S. Geraschenko1, M. O. Gomozkova3,
O. O. Hudkova1, S. J. Vainio2, A. A. Samoylenko2, L. B. Drobot1

1Palladin Institute of Biochemistry, National Academy of Sciences of Ukraine, Kyiv;
2Faculty of Biochemistry and Molecular Medicine, University of Oulu, Finland;
3Brigham Young University-Idaho, Rexburg, USA;

Received: 02 November 2021; Accepted: 12 November 2021

Extracellular vesicles (EVs) are secreted by most cell types under both physiological and pathological conditions and were proposed to be actively involved in intercellular communication. The mode of EVs action is dependent on their cargos composition. EVs  play an important role in tumor initiation, recurrence, metastasis and therapeutic resistance. EVs marker proteins Alix and Tsg101 and cortactin are the binding partners of adaptor protein Ruk/CIN85. The present study aims to analyze the regulatory effects of EVs produced by 4T1 cells with overexpression (RukUp) or down-regulation (RukDown) of adaptor protein Ruk/CIN85 on proliferation rate, migration and invasion activity of parental 4T1 WT cells. EVs from conditioned medium of 4T1 RukUp or RukDown cells were isolated by differential centrifugation followed by further purification using Exo-spin™ kit (Cell Guidance Systems). The number and size of EVs were characterized by NTA (Malvern Panalytical NanoSight NM300) instrument. The content of marker proteins and Ruk/CIN85 in isolated EVs was analyzed by Western-blotting. The viability, migration and invasion activity of 4T1 WT cells were studied using MTT-test, scratch-test and Boyden chamber assay, respectively. It was demonstrated for the first time that adaptor protein Ruk/CIN85 is a constitutive component of  EVs produced by 4T1 cells. It was also shown that EVs produced by 4T1 cells with different levels of Ruk/CIN85 expression are characterized by a specific profile of the content of its multiple molecular forms. It turned out that the ability of EVs to modulate the proliferative activity, motility and invasiveness of 4T1 WT cells was tightly correlated with the biological properties of 4T1 cells that produce EVs (highly aggressive 4T1 RukUp cells or weakly invasive 4T1 RukDown cells). Our data suggest that adaptor protein Ruk/CIN85 is not only a constitutive component of cargos composition of EVs produced by tumor cells but, depending on its content in EVs, plays an active role in the control of carcinogenesis.

Keywords: , , , , ,


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