Autophagy Stimulus Promotes Early HuR Protein Activation and p62/SQSTM1 Protein Synthesis in ARPE-19 Cells by Triggering Erk1/2, p38MAPK, and JNK Kinase Pathways.

Nicoletta Marchesi,Alessandro Provenzani,Eveliina Korhonen,Natthakan Thongon,Marialaura Amadio,Anu Kauppinen,Adrian Smedowski,Alessia Pascale,Stefano Govoni,Kai Kaarniranta,Ali Koskela

doi:10.1155/2018/4956080

Abstract

RNA-binding protein dysregulation and altered expression of proteins involved in the autophagy/proteasome pathway play a role in many neurodegenerative disease onset/progression, including age-related macular degeneration (AMD). HuR/ELAVL1 is a master regulator of gene expression in human physiopathology. In ARPE-19 cells exposed to the proteasomal inhibitor MG132, HuR positively affects at posttranscriptional level p62 expression, a stress response gene involved in protein aggregate clearance with a role in AMD. Here, we studied the early effects of the proautophagy AICAR + MG132 cotreatment on the HuR-p62 pathway. We treated ARPE-19 cells with Erk1/2, AMPK, p38MAPK, PKC, and JNK kinase inhibitors in the presence of AICAR + MG132 and evaluated HuR localization/phosphorylation and p62 expression. Two-hour AICAR + MG132 induces both HuR cytoplasmic translocation and threonine phosphorylation via the Erk1/2 pathway. In these conditions, p62 mRNA is loaded on polysomes and its translation in de novo protein is favored. Additionally, for the first time, we report that JNK can phosphorylate HuR, however, without modulating its localization. Our study supports HuR's role as an upstream regulator of p62 expression in ARPE-19 cells, helps to understand better the early events in response to a proautophagy stimulus, and suggests that modulation of the autophagy-regulating kinases as potential therapeutic targets for AMD may be relevant.

Highlights

Posttranscriptional mechanisms are key determinants in the modulation of gene expression by allowing a punctual, localized adaptation of protein levels to changing environmental conditions
We previously showed that under proteasome inhibition, HuR posttranscriptionally affects the expression of p62/sequestosome 1 (SQSTM1) in a retinal pigment epithelial (RPE) cell line. p62 is a key factor to regulate protein aggregate clearance via autophagy and proteasome pathways that are involved in the pathology of age-related macular degeneration (AMD) [18]
We previously showed that in ARPE-19 cell line under 24 hr proteasomal inhibitor MG132, HuR protein binds p62 mRNA; the specific involvement of HuR in p62 expression regulation at the posttranscriptional level was confirmed by the finding that the MG132-induced increase of p62 protein is counteracted in HuR-silenced ARPE-19 cells [18]

Summary

Introduction

Posttranscriptional mechanisms are key determinants in the modulation of gene expression by allowing a punctual, localized adaptation of protein levels to changing environmental conditions. RNA-binding proteins (RBPs) are predicted to regulate up to 90% of human genes, and their physiological role is critical for the maintenance of health conditions in all tissues, including the eye [1,2,3]. Recent evidence has shown that the dysregulation of RBPs controlling the expression of proteins involved in the autophagy/proteasome pathway has a role in the onset and the progression of many neurodegenerative diseases [4]. The RBP HuR (human antigen R or HuA) is a master regulator of gene expression in several physiological and pathological conditions. HuR’s targets include mRNAs coding proteins involved in the cellular stress response and survival, inflammation, and cell cycle progression [12,13,14,15,16,17]. We previously showed that under proteasome inhibition, HuR posttranscriptionally affects the expression of p62/sequestosome 1 (SQSTM1) in a retinal pigment epithelial (RPE) cell line. p62 is a key factor to regulate protein aggregate clearance via autophagy and proteasome pathways that are involved in the pathology of age-related macular degeneration (AMD) [18]

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