Analysis of furin (FURIN) gene expression in the U-87 MG cell line as a potential target for gene inhibiting therapies in (auto-) immune diseases

Karolina Zapletal-Pudełko; Grzegorz Machnik; Katarzyna Stec-Grosman; Łukasz Bułdak; Bogusław Okopień

doi:10.18778/1730-2366.18.16

Authors

Karolina Zapletal-Pudełko Medical University of Silesia, Faculty of Medical Science in Katowice, Department of Internal Medicine and Clinical Pharmacology, Poland https://orcid.org/0009-0003-2211-1764
Grzegorz Machnik Medical University of Silesia, Faculty of Medical Science in Katowice, Department of Internal Medicine and Clinical Pharmacology, Poland https://orcid.org/0000-0002-9081-0984
Katarzyna Stec-Grosman Medical Laboratory Synevo, Poland
Łukasz Bułdak Medical University of Silesia, Faculty of Medical Science in Katowice, Department of Internal Medicine and Clinical Pharmacology, Poland
Bogusław Okopień Medical University of Silesia, Faculty of Medical Science in Katowice, Department of Internal Medicine and Clinical Pharmacology, Poland

DOI:

https://doi.org/10.18778/1730-2366.18.16

Keywords:

neuroinflammation, FURIN, HERVs, neurodegenerative diseases, gene therapies

Abstract

Purpose: Furin is a proprotein convertase commonly found in the human body. The enzymatic activity of furin is necessary for the activation of numerous substrates including e.g. hormones and growth factors. Nevertheless, furin is involved in various pathological conditions caused by, among others, chronic inflammation. Therefore furin is considered as a potential target in autoimmune diseases therapy.

We performed an experiment in which the expression of FURIN gene in U-87 MG astrocytoma cells was investigated. Additionally, this cell line contains some sequences coding human endogenous retroviruses (HERVs), including ERVW-1 and its receptor- SLC1A5. Deregulation of HERV expression has been observed in some neurodegenerative diseases as well as in inflammatory process.

Material and Methods: Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot (WB) techniques were utilized for analysis. Phorbol 12-myristate 13-acetate (PMA) were used for cell stimulation. Short interfering RNAs (siRNA) were used for gene expression inhibition in U-87 MG cells in vitro.

Results: Cell stimulation by PMA strongly increased FURIN expression, simultaneously downregulating ERVW-1 (p<0.01). Moreover addition of PMA significantly stimulates the autocatalytic action of cellular furin itself resulting in the dissociation of its propeptide that was clearly visible in a time-dependent manner. SiRNA-mediated expression inhibition of ERVW-1 and FURIN influenced the mRNA level for SLC1A5 (ASCT2) - primary syncytin-1 receptor, that was significantly lower. FURIN inhibition by siRNA caused strong upregulation of ERVW-1 expression (p<0.01).

Conclusion: Our results showed that stimulation by PMA and inhibition expression by siRNA affects the expression of FURIN in U-87 MG astrocytoma cells. Moreover, furin shows a complex relationship on the expression of ERVW-1 and SLC1A5 genes, as well as on the form (precursor or mature) and the amount of the final translation products of the transcripts. The regulation of FURIN may pose a potential therapeutic strategy in the treatment of neurodegenerative diseases caused by autoimmunity.

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