Cell-Specific Post-Transcriptional Regulation of γ-Synuclein Gene by Micro-RNAs

Irina Surgucheva,H Shanker Rao,Sumedha Gunewardena,Andrei Surguchov,Sanjoy Bhattacharya

doi:10.1371/journal.pone.0073786

Abstract

γ-Synuclein is a member of the synucleins family of small proteins, which consists of three members:α, β- and γ-synuclein. γ-Synuclein is abnormally expressed in a high percentage of advanced and metastatic tumors, but not in normal or benign tissues. Furthermore, γ-synuclein expression is strongly correlated with disease progression, and can stimulate proliferation, induce invasion and metastasis of cancer cells. γ-Synuclein transcription is regulated basically through the binding of AP-1 to specific sequences in intron 1. Here we show that γ-synuclein expression may be also regulated by micro RNAs (miRs) on post-transcriptional level. According to prediction by several methods, the 3′-untranslated region (UTR) of γ-synuclein gene contains targets for miRs. Insertion of γ-synuclein 3′-UTR downstream of the reporter luciferase (LUC) gene causes a 51% reduction of LUC activity after transfection into SKBR3 and Y79 cells, confirming the presence of efficient targets for miRs in this fragment. Expression of miR-4437 and miR-4674 for which putative targets in 3′-UTR were predicted caused a 61.2% and 60.1% reduction of endogenous γ-synuclein expression confirming their role in gene expression regulation. On the other hand, in cells overexpressing γ-synuclein no significant effect of miRs on γ-synuclein expression was found suggesting that miRs exert their regulatory effect only at low or moderate, but not at high level of γ-synuclein expression. Elevated level of γ-synuclein differentially changes the level of several miRs expression, upregulating the level of some miRs and downregulating the level of others. Three miRs upregulated as a result of γ-synuclein overexpression, i.e., miR-885-3p, miR-138 and miR-497 have putative targets in 3′-UTR of the γ-synuclein gene. Some of miRs differentially regulated by γ-synuclein may modulate signaling pathways and cancer related gene expression. This study demonstrates that miRs might provide cell-specific regulation of γ-synuclein expression and set the stage to further evaluate their role in pathophysiological processes.

Highlights

Introduction cSynuclein was first identified by a differential cDNA sequencing approach and was termed the breast-cancer-specific gene 1 (BCSG1)
After identifying putative micro RNAs (miRs) targets in c-synuclein 39-untranslated region (UTR) we examined whether these targets are involved in the regulation of gene expression
We examined whether miR-4437 and miR-4674 for which targets were predicted in the 39-untranslated region (39-UTR) by bioinformatics search affected c-synuclein expression in cultured cells

Summary

Introduction

Introduction cSynuclein was first identified by a differential cDNA sequencing approach and was termed the breast-cancer-specific gene 1 (BCSG1). The gene is expressed in high abundance in a breast cancer (BC) cDNA library but scarcely in a normal breast cDNA library [1]. Later the overexpression of c-synuclein has been shown in several types of cancer [2,3,4,5,6]. In cancer cells csynuclein significantly increases the ligand-dependent transcriptional activity of estrogen receptor-alpha (ER-alpha). Overexpression of c-synuclein stimulates the ligand-dependent cell proliferation and estrogen-induced activation of ERK1/2, whereas suppression of endogenous c-synuclein expression considerably inhibits cell growth in response to estrogen. Highly elevated level of c-synuclein is documented in many types of cancer, the triggers initiating its high expression as well as the downstream molecular events of such upregulation are poorly understood. Deregulation of c-synuclein accompanied by the formation of aberrant inclusions is associated with neurodegenerative diseases [9,10,11,12,13]

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