MicroRNA-129-5p-regulated microglial expression of the surface receptor CD200R1 controls neuroinflammation

Vikas Singh,Shaivya Kushwaha,Jamal Ahmad Ansari,Siddhartha Gangopadhyay,Shubhendra K Mishra,Rajib K Dey,Ashok K Giri,Satyakam Patnaik,Debabrata Ghosh

doi:10.1016/j.jbc.2021.101521

Abstract

CD200R1 is an inhibitory surface receptor expressed in microglia and blood macrophages. Microglial CD200R1 is known to control neuroinflammation by keeping the microglia in resting state, and therefore, tight regulation of its expression is important. CCAAT/enhancer-binding protein β (CEBPβ) is the known regulator of CD200R1 transcription. In the present study, our specific intention was to find a possible posttranscriptional regulatory mechanism of CD200R1 expression. Here we investigated a novel regulatory mechanism of CD200R1 expression following exposure to an environmental stressor, arsenic, combining in silico analysis, in vitro, and in vivo experiments, as well as validation in human samples. The in silico analysis and in vitro studies with primary neonatal microglia and BV2 microglia revealed that arsenic demethylates the promoter of a microRNA, miR-129-5p, thereby increasing its expression, which subsequently represses CD200R1 by binding to its 3′-untranslated region and shuttling the CD200R1 mRNA to the cytoplasmic-processing body in mouse microglia. The role of miR-129-5p was further validated in BALB/c mouse by stereotaxically injecting anti-miR-129. We found that anti-miR-129 reversed the expression of CD200R1, as well as levels of inflammatory molecules IL-6 and TNF-α. Experiments with a CD200R1 siRNA-induced loss-of-function mouse model confirmed an miR-129-5p→CD200R1→IL-6/TNF-α signaling axis. These main findings were replicated in a human cell line and validated in human samples. Taken together, our study revealed miR-129-5p as a novel posttranscriptional regulator of CD200R1 expression with potential implications in neuroinflammation and related complications.

Highlights

CD200R1 is an inhibitory surface receptor expressed in microglia and blood macrophages
129-5p function is similar across the species, increase in the formation of P-bodies with we performed in silico analysis and found that GW182 immunostaining, one of the essential miR-129-5p has two binding sites
The post-transcriptional regulation of CD200R1 is of great clinical interest as it is involved in neuroinflammation (4) and other cellular functions

Summary

Introduction

Keywords Microglia, miR-129-5p, CD200R1, arsenic, miRNA, neuroinflammation, Cytokine, neuroimmunology. We not altered in brain following in vivo treatment searched for potential CpG sites 1000 bp (Fig. 5C) even though arsenic could decrease upstream of the miR-129-2 gene the expression of CD200R1 protein transcriptional start site using Methprimer significantly (~0.58 fold) as detected in western software (41). Brain sections were immunostained primers which can detect all four variants of against CD200R1 (Fig. 5E) and its quantitative CD200R1 mRNA and found a non-significant analysis (Fig.S5) supports the changes decrease (~1.4 fold in control subjects and observed in the CD200R1 protein expression ~0.71 fold in arsenicosis subjects) in their following arsenic, anti-miR, and/or siRNA levels (Fig.6A). One individual showed high neuropathy with additional symptoms like paresthesia, vibration joint sense, muscle wasting (Table S1)

Discussion

Findings

Experimental procedures